WorldWideScience

Sample records for alu rna editing

  1. Alu sequences in undifferentiated human embryonic stem cells display high levels of A-to-I RNA editing.

    Directory of Open Access Journals (Sweden)

    Sivan Osenberg

    Full Text Available Adenosine to Inosine (A-to-I RNA editing is a site-specific modification of RNA transcripts, catalyzed by members of the ADAR (Adenosine Deaminase Acting on RNA protein family. RNA editing occurs in human RNA in thousands of different sites. Some of the sites are located in protein-coding regions but the majority is found in non-coding regions, such as 3'UTRs, 5'UTRs and introns - mainly in Alu elements. While editing is found in all tissues, the highest levels of editing are found in the brain. It was shown that editing levels within protein-coding regions are increased during embryogenesis and after birth and that RNA editing is crucial for organism viability as well as for normal development. In this study we characterized the A-to-I RNA editing phenomenon during neuronal and spontaneous differentiation of human embryonic stem cells (hESCs. We identified high editing levels of Alu repetitive elements in hESCs and demonstrated a global decrease in editing levels of non-coding Alu sites when hESCs are differentiating, particularly into the neural lineage. Using RNA interference, we showed that the elevated editing levels of Alu elements in undifferentiated hESCs are highly dependent on ADAR1. DNA microarray analysis showed that ADAR1 knockdown has a global effect on gene expression in hESCs and leads to a significant increase in RNA expression levels of genes involved in differentiation and development processes, including neurogenesis. Taken together, we speculate that A-to-I editing of Alu sequences plays a role in the regulation of hESC early differentiation decisions.

  2. Consistent levels of A-to-I RNA editing across individuals in coding sequences and non-conserved Alu repeats

    Directory of Open Access Journals (Sweden)

    Osenberg Sivan

    2010-10-01

    Full Text Available Abstract Background Adenosine to inosine (A-to-I RNA-editing is an essential post-transcriptional mechanism that occurs in numerous sites in the human transcriptome, mainly within Alu repeats. It has been shown to have consistent levels of editing across individuals in a few targets in the human brain and altered in several human pathologies. However, the variability across human individuals of editing levels in other tissues has not been studied so far. Results Here, we analyzed 32 skin samples, looking at A-to-I editing level in three genes within coding sequences and in the Alu repeats of six different genes. We observed highly consistent editing levels across different individuals as well as across tissues, not only in coding targets but, surprisingly, also in the non evolutionary conserved Alu repeats. Conclusions Our findings suggest that A-to-I RNA-editing of Alu elements is a tightly regulated process and, as such, might have been recruited in the course of primate evolution for post-transcriptional regulatory mechanisms.

  3. Selective stimulation of translational expression by Alu RNA

    OpenAIRE

    Rubin, Carol M.; Kimura, Richard H.; Schmid, Carl W.

    2002-01-01

    Human Alu and adenovirus VA1 RNAs each stimulate the translational expression of reporter genes in co-transient transfection assays without affecting either the rate of global protein synthesis or the abundance of the reporter mRNA. This selective, post-transcriptional stimulation of expression, which is observed in human and mouse cell lines and for three reporters, acts through a PKR- independent mechanism. The activity of Alu and VA1 RNAs in this assay is transient, causing a reduction in ...

  4. 3'-UTR-located inverted Alu repeats facilitate mRNA translational repression and stress granule accumulation.

    Science.gov (United States)

    Fitzpatrick, Terry; Huang, Sui

    2012-07-01

    Alu repeats within human genes may potentially alter gene expression. Here, we show that 3'-UTR-located inverted Alu repeats significantly reduce expression of an AcGFP reporter gene. Mutational analysis demonstrates that the secondary structure, but not the primary nucleotide sequence, of the inverted Alu repeats is critical for repression. The expression levels and nucleocytoplasmic distribution of reporter mRNAs with or without 3'-UTR inverted Alu repeats are similar; suggesting that reporter gene repression is not due to changes in mRNA levels or mRNA nuclear sequestration. Instead, reporter gene mRNAs harboring 3'-UTR inverted Alu repeats accumulate in cytoplasmic stress granules. These findings may suggest a novel mechanism whereby 3'-UTR-located inverted Alu repeats regulate human gene expression through sequestration of mRNAs within stress granules. PMID:22688648

  5. Iron Toxicity in the Retina Requires Alu RNA and the NLRP3 Inflammasome

    Directory of Open Access Journals (Sweden)

    Bradley D. Gelfand

    2015-06-01

    Full Text Available Excess iron induces tissue damage and is implicated in age-related macular degeneration (AMD. Iron toxicity is widely attributed to hydroxyl radical formation through Fenton’s reaction. We report that excess iron, but not other Fenton catalytic metals, induces activation of the NLRP3 inflammasome, a pathway also implicated in AMD. Additionally, iron-induced degeneration of the retinal pigmented epithelium (RPE is suppressed in mice lacking inflammasome components caspase-1/11 or Nlrp3 or by inhibition of caspase-1. Iron overload increases abundance of RNAs transcribed from short interspersed nuclear elements (SINEs: Alu RNAs and the rodent equivalent B1 and B2 RNAs, which are inflammasome agonists. Targeting Alu or B2 RNA prevents iron-induced inflammasome activation and RPE degeneration. Iron-induced SINE RNA accumulation is due to suppression of DICER1 via sequestration of the co-factor poly(C-binding protein 2 (PCBP2. These findings reveal an unexpected mechanism of iron toxicity, with implications for AMD and neurodegenerative diseases associated with excess iron.

  6. A structural determinant required for RNA editing

    Science.gov (United States)

    Tian, Nan; Yang, Yun; Sachsenmaier, Nora; Muggenhumer, Dominik; Bi, Jingpei; Waldsich, Christina; Jantsch, Michael F.; Jin, Yongfeng

    2011-01-01

    RNA editing by adenosine deaminases acting on RNAs (ADARs) can be both specific and non-specific, depending on the substrate. Specific editing of particular adenosines may depend on the overall sequence and structural context. However, the detailed mechanisms underlying these preferences are not fully understood. Here, we show that duplex structures mimicking an editing site in the Gabra3 pre-mRNA unexpectedly fail to support RNA editing at the Gabra3 I/M site, although phylogenetic analysis suggest an evolutionarily conserved duplex structure essential for efficient RNA editing. These unusual results led us to revisit the structural requirement for this editing by mutagenesis analysis. In vivo nuclear injection experiments of mutated editing substrates demonstrate that a non-conserved structure is a determinant for editing. This structure contains bulges either on the same or the strand opposing the edited adenosine. The position of these bulges and the distance to the edited base regulate editing. Moreover, elevated folding temperature can lead to a switch in RNA editing suggesting an RNA structural change. Our results indicate the importance of RNA tertiary structure in determining RNA editing. PMID:21427087

  7. Altered A-to-I RNA editing in human embryogenesis.

    Directory of Open Access Journals (Sweden)

    Ronit Shtrichman

    Full Text Available Post-transcriptional events play an important role in human development. The question arises as to whether Adenosine to Inosine RNA editing, catalyzed by the ADAR (Adenosine Deaminase acting on RNA enzymes, differs in human embryogenesis and in adulthood. We tested the editing of various target genes in coding (FLNA, BLCAP, CYFIP2 and non-coding sequences at their Alu elements (BRCA1, CARD11, RBBP9, MDM4, FNACC, as well as the transcriptional levels of the ADAR1 enzymes. This analysis was performed on five fetal and adult human tissues: brain, heart, liver, kidney, and spleen, as well as on human embryonic stem cells (hESCs, which represent the blastocyst stage in early human development. Our results show substantially greater editing activity for most adult tissue samples relative to fetal ones, in six of the eight genes tested. To test the effect of reduced A-to-I RNA editing activity in early human development we used human embryonic stem cells (hESCs as a model and tried to generate hESC clones that overexpress the ADAR1-p110 isoform. We were unable to achieve overexpression of ADAR1-p110 by either transfection or lentiviral infection, though we easily generated hESC clones that expressed the GFP transgene and overexpressed ADAR1-p110 in 293T cells and in primary human foreskin fibroblast (HFF cells. Moreover, in contrast to the expected overexpression of ADAR1-p110 protein following its introduction into hESCs, the expression levels of this protein decreased dramatically 24-48 hr post infection. Similar results were obtained when we tried to overexpress ADAR1-p110 in pluripotent embryonal carcinoma cells. This suggests that ADAR1 protein is substantially regulated in undifferentiated pluripotent hESCs. Overall, our data suggest that A-to-I RNA editing plays a critical role during early human development.

  8. Exploring the RNA editing potential of RNA-Seq data by ExpEdit.

    Science.gov (United States)

    D'Antonio, Mattia; Picardi, Ernesto; Castrignanò, Tiziana; D'Erchia, Anna Maria; Pesole, Graziano

    2015-01-01

    Revealing the impact of A-to-I RNA editing in RNA-Seq experiments is relevant in humans because RNA editing can influence gene expression. In addition, its deregulation has been linked to a variety of human diseases. Exploiting the RNA editing potential in complete RNA-Seq datasets, however, is a challenging task. Indeed, no dedicated software is available, and sometimes deep computational skills and appropriate hardware resources are required. To explore the impact of known RNA editing events in massive transcriptome sequencing experiments, we developed the ExpEdit web service application. In the present work, we provide an overview of ExpEdit as well as methodologies to investigate known RNA editing in human RNA-Seq datasets. PMID:25577388

  9. Statistical Physics Approaches to RNA Editing

    Science.gov (United States)

    Bundschuh, Ralf

    2012-02-01

    The central dogma of molecular Biology states that DNA is transcribed base by base into RNA which is in turn translated into proteins. However, some organisms edit their RNA before translation by inserting, deleting, or substituting individual or short stretches of bases. In many instances the mechanisms by which an organism recognizes the positions at which to edit or by which it performs the actual editing are unknown. One model system that stands out by its very high rate of on average one out of 25 bases being edited are the Myxomycetes, a class of slime molds. In this talk we will show how the computational methods and concepts from statistical Physics can be used to analyze DNA and protein sequence data to predict editing sites in these slime molds and to guide experiments that identified previously unknown types of editing as well as the complete set of editing events in the slime mold Physarum polycephalum.

  10. Novel modes of RNA editing in mitochondria.

    Science.gov (United States)

    Moreira, Sandrine; Valach, Matus; Aoulad-Aissa, Mohamed; Otto, Christian; Burger, Gertraud

    2016-06-01

    Gene structure and expression in diplonemid mitochondria are unparalleled. Genes are fragmented in pieces (modules) that are separately transcribed, followed by the joining of module transcripts to contiguous RNAs. Some instances of unique uridine insertion RNA editing at module boundaries were noted, but the extent and potential occurrence of other editing types remained unknown. Comparative analysis of deep transcriptome and genome data from Diplonema papillatum mitochondria reveals ∼220 post-transcriptional insertions of uridines, but no insertions of other nucleotides nor deletions. In addition, we detect in total 114 substitutions of cytosine by uridine and adenosine by inosine, amassed into unusually compact clusters. Inosines in transcripts were confirmed experimentally. This is the first report of adenosine-to-inosine editing of mRNAs and ribosomal RNAs in mitochondria. In mRNAs, editing causes mostly amino-acid additions and non-synonymous substitutions; in ribosomal RNAs, it permits formation of canonical secondary structures. Two extensively edited transcripts were compared across four diplonemids. The pattern of uridine-insertion editing is strictly conserved, whereas substitution editing has diverged dramatically, but still rendering diplonemid proteins more similar to other eukaryotic orthologs. We posit that RNA editing not only compensates but also sustains, or even accelerates, ultra-rapid evolution of genome structure and sequence in diplonemid mitochondria. PMID:27001515

  11. The art of editing RNA structural alignments

    DEFF Research Database (Denmark)

    Andersen, Ebbe Sloth

    2014-01-01

    rewarded by great insight into the evolution of structure and function of your favorite RNA molecule. In this chapter I will review the methods and considerations that go into constructing RNA structural alignments at the secondary and tertiary structure level; introduce software, databases, and algorithms......Manual editing of RNA structural alignments may be considered more art than science, since it still requires an expert biologist to take multiple levels of information into account and be slightly creative when constructing high-quality alignments. Even though the task is rather tedious, it is...

  12. Pentatricopeptide repeat proteins involved in plant organellar RNA editing

    OpenAIRE

    Yagi, Yusuke; Tachikawa, Makoto; Noguchi, Hisayo; Satoh, Soichirou; Obokata, Junichi; Nakamura, Takahiro

    2013-01-01

    C-to-U RNA editing has been widely observed in organellar RNAs in terrestrial plants. Recent research has revealed the significance of a large, plant-specific family of pentatricopeptide repeat (PPR) proteins for RNA editing and other RNA processing events in plant mitochondria and chloroplasts. PPR protein is a sequence-specific RNA-binding protein that identifies specific C residues for editing. Discovery of the RNA recognition code for PPR motifs, including verification and prediction of t...

  13. Analysis of RNA editing in non-human primates

    OpenAIRE

    Marjan Bozinoski; Chris Mason

    2015-01-01

    Adenosine deaminases acting on RNA (ADAR) mediated RNA editing is the prevalent form of post-transcriptional RNA modification in higher organisms. Deaminated Adenosine creates Inosine, which is recognized as Guanosine thereby causing changes in the coding sequence, splicing and miRNA binding. RNA editing is mostly primate specific phenomenon and few studies of limited scope have shown that humans possess more editing sites than other primates. Here, we present the extent and intensity of ...

  14. Ebola virus RNA editing depends on the primary editing site sequence and an upstream secondary structure.

    Science.gov (United States)

    Mehedi, Masfique; Hoenen, Thomas; Robertson, Shelly; Ricklefs, Stacy; Dolan, Michael A; Taylor, Travis; Falzarano, Darryl; Ebihara, Hideki; Porcella, Stephen F; Feldmann, Heinz

    2013-01-01

    Ebolavirus (EBOV), the causative agent of a severe hemorrhagic fever and a biosafety level 4 pathogen, increases its genome coding capacity by producing multiple transcripts encoding for structural and nonstructural glycoproteins from a single gene. This is achieved through RNA editing, during which non-template adenosine residues are incorporated into the EBOV mRNAs at an editing site encoding for 7 adenosine residues. However, the mechanism of EBOV RNA editing is currently not understood. In this study, we report for the first time that minigenomes containing the glycoprotein gene editing site can undergo RNA editing, thereby eliminating the requirement for a biosafety level 4 laboratory to study EBOV RNA editing. Using a newly developed dual-reporter minigenome, we have characterized the mechanism of EBOV RNA editing, and have identified cis-acting sequences that are required for editing, located between 9 nt upstream and 9 nt downstream of the editing site. Moreover, we show that a secondary structure in the upstream cis-acting sequence plays an important role in RNA editing. EBOV RNA editing is glycoprotein gene-specific, as a stretch encoding for 7 adenosine residues located in the viral polymerase gene did not serve as an editing site, most likely due to an absence of the necessary cis-acting sequences. Finally, the EBOV protein VP30 was identified as a trans-acting factor for RNA editing, constituting a novel function for this protein. Overall, our results provide novel insights into the RNA editing mechanism of EBOV, further understanding of which might result in novel intervention strategies against this viral pathogen. PMID:24146620

  15. Ebola virus RNA editing depends on the primary editing site sequence and an upstream secondary structure.

    Directory of Open Access Journals (Sweden)

    Masfique Mehedi

    Full Text Available Ebolavirus (EBOV, the causative agent of a severe hemorrhagic fever and a biosafety level 4 pathogen, increases its genome coding capacity by producing multiple transcripts encoding for structural and nonstructural glycoproteins from a single gene. This is achieved through RNA editing, during which non-template adenosine residues are incorporated into the EBOV mRNAs at an editing site encoding for 7 adenosine residues. However, the mechanism of EBOV RNA editing is currently not understood. In this study, we report for the first time that minigenomes containing the glycoprotein gene editing site can undergo RNA editing, thereby eliminating the requirement for a biosafety level 4 laboratory to study EBOV RNA editing. Using a newly developed dual-reporter minigenome, we have characterized the mechanism of EBOV RNA editing, and have identified cis-acting sequences that are required for editing, located between 9 nt upstream and 9 nt downstream of the editing site. Moreover, we show that a secondary structure in the upstream cis-acting sequence plays an important role in RNA editing. EBOV RNA editing is glycoprotein gene-specific, as a stretch encoding for 7 adenosine residues located in the viral polymerase gene did not serve as an editing site, most likely due to an absence of the necessary cis-acting sequences. Finally, the EBOV protein VP30 was identified as a trans-acting factor for RNA editing, constituting a novel function for this protein. Overall, our results provide novel insights into the RNA editing mechanism of EBOV, further understanding of which might result in novel intervention strategies against this viral pathogen.

  16. An RNA editing fingerprint of cancer stem cell reprogramming

    OpenAIRE

    Crews, Leslie A; Jiang, Qingfei; Zipeto, Maria A; de Lazzari, Elisa; Court, Angela C.; Ali, Shawn; Barrett, Christian L.; Frazer, Kelly A; Jamieson, Catriona HM

    2015-01-01

    Background Deregulation of RNA editing by adenosine deaminases acting on dsRNA (ADARs) has been implicated in the progression of diverse human cancers including hematopoietic malignancies such as chronic myeloid leukemia (CML). Inflammation-associated activation of ADAR1 occurs in leukemia stem cells specifically in the advanced, often drug-resistant stage of CML known as blast crisis. However, detection of cancer stem cell-associated RNA editing by RNA sequencing in these rare cell populatio...

  17. Ebola Virus RNA Editing Depends on the Primary Editing Site Sequence and an Upstream Secondary Structure

    OpenAIRE

    Mehedi, Masfique; Hoenen, Thomas; Robertson, Shelly; Ricklefs, Stacy; Dolan, Michael A.; Taylor, Travis; Falzarano, Darryl; Ebihara, Hideki; Porcella, Stephen F.; Feldmann, Heinz

    2013-01-01

    Ebolavirus (EBOV), the causative agent of a severe hemorrhagic fever and a biosafety level 4 pathogen, increases its genome coding capacity by producing multiple transcripts encoding for structural and nonstructural glycoproteins from a single gene. This is achieved through RNA editing, during which non-template adenosine residues are incorporated into the EBOV mRNAs at an editing site encoding for 7 adenosine residues. However, the mechanism of EBOV RNA editing is currently not understood. I...

  18. Deletions in cox2 mRNA result in loss of splicing and RNA editing and gain of novel RNA editing sites.

    Directory of Open Access Journals (Sweden)

    Stefanie Grüttner

    Full Text Available As previously demonstrated, the maize cox2 RNA is fully edited in cauliflower mitochondria. Use of constructs with a deleted cox2 intron, however, led to a loss of RNA editing at almost all editing sites, with only a few sites still partially edited. Likewise, one deletion in exon 1 and three in exon 2 abolish RNA editing at all cox2 sites analyzed. Furthermore, intron splicing is abolished using these deletions. Mutation of a cytosine residue, which is normally edited and localized directly adjacent to the intron, to thymidine did not result in restoration of splicing, indicating that the loss of splicing was not due to loss of RNA editing. One deletion in exon 2 did not lead to loss of splicing. Instead, most editing sites were found to be edited, only three were not edited. Unexpectedly, we observed additional RNA editing events at new sites. Thus it appears that deletions in the cox2 RNA sequence can have a strong effect on RNA processing, leading to loss of splicing, loss of editing at all sites, or even to a gain of new editing sites. As these effects are not limited to the vicinity of the respective deletions, but appear to be widespread or even affect all editing sites, they may not be explained by the loss of PPR binding sites. Instead, it appears that several parts of the cox2 transcript are required for proper RNA processing. This indicates the roles of the RNA sequence and structural elements in the recognition of the editing sites.

  19. RNA Editing and its Control in Hepatitis Delta Virus Replication

    Directory of Open Access Journals (Sweden)

    John L. Casey

    2010-01-01

    Full Text Available The hepatitis delta virus genome is a small circular RNA, similar to viroids. Although HDV contains a gene, the protein produced (HDAg is encoded by less than half the genome and possesses no RNA polymerase activity. Because of this limited coding capacity, HDV relies heavily on host functions and on structural features of the viral RNA—very much like viroids. The virus’ use of host RNA editing activity to produce two functionally distinct forms of HDAg is a particularly good example of this reliance. This review covers the mechanisms and control of RNA editing in the HDV replication cycle.

  20. A model for codon position bias in RNA editing

    CERN Document Server

    Liu, T; Liu, Tsunglin; Bundschuh, Ralf

    2005-01-01

    RNA editing can be crucial for the expression of genetic information via inserting, deleting, or substituting a few nucleotides at specific positions in an RNA sequence. Within coding regions in an RNA sequence, editing usually occurs with a certain bias in choosing the positions of the editing sites. In the mitochondrial genes of {\\it Physarum polycephalum}, many more editing events have been observed at the third codon position than at the first and second, while in some plant mitochondria the second codon position dominates. Here we propose an evolutionary model that explains this bias as the basis of selection at the protein level. The model predicts a distribution of the three positions rather close to the experimental observation in {\\it Physarum}. This suggests that the codon position bias in {\\it Physarum} is mainly a consequence of selection at the protein level.

  1. Hepatocellular carcinoma: transcriptome diversity regulated by RNA editing.

    Science.gov (United States)

    Li, Yan; Chen, Leilei; Chan, Tim Hon Man; Guan, Xin-Yuan

    2013-08-01

    Hepatocellular carcinoma (HCC) can be envisioned as a prolonged multi-stage process accumulating genetic and epigenetic changes. In the past years, DNA alterations lent us important clues to the comprehension of molecular pathways involved in HCC. However, as an increasing number of RNAs were identified to be subject to A-to-I modifications, it has become apparent that RNA editing might be the causal basis of various human diseases. Recent evidence has strengthened this notion by correlating hyper-edited AZIN1 (antizyme inhibitor 1) protein with HCC onset and the mechanisms that regulate cell transformation. As we continue to demystify it, RNA editing astonishes us with its diverse substrates, esoteric functions, elaborate machinery and complex interaction with HBV/HCV viral infection. In this review, we examine the contribution of A-to-I RNA editing to caner onset/progression and explore its potential implications for cancer treatment advances. PMID:23748106

  2. Alu element-containing RNAs maintain nucleolar structure and function.

    Science.gov (United States)

    Caudron-Herger, Maïwen; Pankert, Teresa; Seiler, Jeanette; Németh, Attila; Voit, Renate; Grummt, Ingrid; Rippe, Karsten

    2015-11-12

    Non-coding RNAs play a key role in organizing the nucleus into functional subcompartments. By combining fluorescence microscopy and RNA deep-sequencing-based analysis, we found that RNA polymerase II transcripts originating from intronic Alu elements (aluRNAs) were enriched in the nucleolus. Antisense-oligo-mediated depletion of aluRNAs or drug-induced inhibition of RNA polymerase II activity disrupted nucleolar structure and impaired RNA polymerase I-dependent transcription of rRNA genes. In contrast, overexpression of a prototypic aluRNA sequence increased both nucleolus size and levels of pre-rRNA, suggesting a functional link between aluRNA, nucleolus integrity and pre-rRNA synthesis. Furthermore, we show that aluRNAs interact with nucleolin and target ectopic genomic loci to the nucleolus. Our study suggests an aluRNA-based mechanism that links RNA polymerase I and II activities and modulates nucleolar structure and rRNA production. PMID:26464461

  3. RNA editing changes the identity of a mitochondrial tRNA in marsupials.

    OpenAIRE

    Börner, G V; Mörl, M.; Janke, A.; Pääbo, S

    1996-01-01

    In the mitochondrial genome of marsupials, the tRNA gene located at the position where in other mammals an aspartyl-tRNA is encoded carries the glycine anticodon GCC. Post-transcriptionally, an RNA editing mechanism affects the second position of the anticodon such that the aspartate anticodon GUC is created in approximately 50% of the mature tRNA pool. We show that the unedited version of this tRNA'Asp' (GCC) can be specifically aminoacylated with glycine in vitro, while the edited version b...

  4. Organelle RNA recognition motif-containing (ORRM) proteins are plastid and mitochondrial editing factors in Arabidopsis.

    Science.gov (United States)

    Shi, Xiaowen; Bentolila, Stephane; Hanson, Maureen R

    2016-05-01

    Post-transcriptional C-to-U RNA editing occurs at specific sites in plastid and plant mitochondrial transcripts. Members of the Arabidopsis pentatricopeptide repeat (PPR) motif-containing protein family and RNA-editing factor Interacting Protein (RIP, also known as MORF) family have been characterized as essential components of the RNA editing apparatus. Recent studies reveal that several organelle-targeted RNA recognition motif (RRM)-containing proteins are involved in either plastid or mitochondrial RNA editing. ORRM1 (Organelle RRM protein 1) is essential for plastid editing, whereas ORRM2, ORRM3 and ORRM4 are involved in mitochondrial RNA editing. The RRM domain of ORRM1, ORRM3 and ORRM4 is required for editing activity, whereas the auxiliary RIP and Glycine-Rich (GR) domains mediate the ORRM proteins' interactions with other editing factors. The identification of the ORRM proteins as RNA editing factors further expands our knowledge of the composition of the editosome. PMID:27082488

  5. Kinetoplastid RNA-editing-associated protein 1 (REAP-1): a novel editing complex protein with repetitive domains.

    OpenAIRE

    Madison-Antenucci, S; Sabatini, R S; Pollard, V W; Hajduk, S L

    1998-01-01

    Kinetoplastid RNA editing consists of the addition or deletion of uridines at specific sites within mitochondrial mRNAs. This unusual RNA processing event is catalyzed by a ribonucleoprotein (RNP) complex that includes editing site-specific endoribonuclease, RNA ligase and terminal uridylnucleotidyl transferase (Tutase) among its essential enzymatic activities. To identify the components of this RNP, monoclonal antibodies were raised against partially purified editing complexes. One antibody ...

  6. Comprehensive analysis of RNA-Seq data reveals extensive RNA editing in a human transcriptome

    DEFF Research Database (Denmark)

    Peng, Zhiyu; Cheng, Yanbing; Tan, Bertrand Chin-Ming;

    2012-01-01

    RNA editing is a post-transcriptional event that recodes hereditary information. Here we describe a comprehensive profile of the RNA editome of a male Han Chinese individual based on analysis of ∼767 million sequencing reads from poly(A)(+), poly(A)(-) and small RNA samples. We developed a...

  7. Identification of RNA editing sites in the SNP database

    OpenAIRE

    Eisenberg, Eli; Adamsky, Konstantin; Cohen, Lital; Amariglio, Ninette; Hirshberg, Abraham; Rechavi, Gideon; Levanon, Erez Y.

    2005-01-01

    The relationship between human inherited genomic variations and phenotypic differences has been the focus of much research effort in recent years. These studies benefit from millions of single-nucleotide polymorphism (SNP) records available in public databases, such as dbSNP. The importance of identifying false dbSNP records increases with the growing role played by SNPs in linkage analysis for disease traits. In particular, the emerging understanding of the abundance of DNA and RNA editing c...

  8. RNA-guided genome editing à la carte

    OpenAIRE

    Horvath, Philippe; Barrangou, Rodolphe

    2013-01-01

    Two recent papers in Science illustrate how the prokaryotic CRISPR-Cas immune system machinery, which typically targets invasive genetic elements such as viruses and plasmids, can be converted into a sophisticated molecular tool for next-generation human genome editing. The versatile Cas9 RNA-guided endonuclease can be readily reprogrammed using customizable small RNAs for sequence-specific single- or double-stranded DNA cleavage.

  9. The moss Physcomitrella patens, a model plant for the study of RNA editing in plant organelles

    OpenAIRE

    Tasaki, Eiji; Sugita, Mamoru

    2010-01-01

    RNA editing is an enigmatic phenomenon in which specific cytidines (C) in the transcripts are changed to uridines (U). In flowering plants, over 500 editing sites have been identified in the mitochondria and plastids. By contrast, in a moss Physcomitrella patens, only 12 editing sites are found in both organelles. Recent extensive genetics studies have revealed involvement of the pentatricopeptide repeat (PPR) proteins with a C-terminal DYW domain (PPR-DYW) in site-specific RNA editing events...

  10. CURE-Chloroplast: A chloroplast C-to-U RNA editing predictor for seed plants

    Directory of Open Access Journals (Sweden)

    Li Yanda

    2009-05-01

    Full Text Available Abstract Background RNA editing is a type of post-transcriptional modification of RNA and belongs to the class of mechanisms that contribute to the complexity of transcriptomes. C-to-U RNA editing is commonly observed in plant mitochondria and chloroplasts. The in vivo mechanism of recognizing C-to-U RNA editing sites is still unknown. In recent years, many efforts have been made to computationally predict C-to-U RNA editing sites in the mitochondria of seed plants, but there is still no algorithm available for C-to-U RNA editing site prediction in the chloroplasts of seed plants. Results In this paper, we extend our algorithm CURE, which can accurately predict the C-to-U RNA editing sites in mitochondria, to predict C-to-U RNA editing sites in the chloroplasts of seed plants. The algorithm achieves over 80% sensitivity and over 99% specificity. We implement the algorithm as an online service called CURE-Chloroplast http://bioinfo.au.tsinghua.edu.cn/pure. Conclusion CURE-Chloroplast is an online service for predicting the C-to-U RNA editing sites in the chloroplasts of seed plants. The online service allows the processing of entire chloroplast genome sequences. Since CURE-Chloroplast performs very well, it could be a helpful tool in the study of C-to-U RNA editing in the chloroplasts of seed plants.

  11. Integrity of the core mitochondrial RNA-binding complex 1/nis vital for trypanosome RNA editing

    Czech Academy of Sciences Publication Activity Database

    Huang, Zhenqiu; Faktorová, Drahomíra; Křížová, A.; Kafková, L.; Read, L. K.; Lukeš, Julius; Hashimi, Hassan

    2015-01-01

    Roč. 21, č. 12 (2015), s. 2088-2102. ISSN 1355-8382 R&D Projects: GA ČR GA15-21974S EU Projects: European Commission(XE) 289007 Institutional support: RVO:60077344 Keywords : RNA editing * mitochondrion * trypanosome Subject RIV: EB - Genetics ; Molecular Biology Impact factor: 4.936, year: 2014

  12. Plant-type mitochondrial RNA editing in the protist Naegleria gruberi

    OpenAIRE

    Rüdinger, Mareike; Fritz-Laylin, Lillian; Polsakiewicz, Monika; Knoop, Volker

    2011-01-01

    RNA editing converts hundreds of cytidines into uridines in plant organelles. Recognition of editing sites requires nuclear encoded RNA-binding pentatricopeptide repeat (PPR) proteins. Previous studies have documented the presence of PPR proteins encoded by the genome of the protist Naegleria gruberi, raising the possibility that plant-like RNA editing might occur in this organism. This paper shows that this is, indeed, the case. Because Naegleria and plants diverged ∼1.5 billion years ago, t...

  13. Genome-wide A-to-I RNA editing in fungi independent of ADAR enzymes

    Science.gov (United States)

    Liu, Huiquan; Wang, Qinhu; He, Yi; Chen, Lingfeng; Hao, Chaofeng; Jiang, Cong; Li, Yang; Dai, Yafeng; Kang, Zhensheng; Xu, Jin-Rong

    2016-01-01

    Yeasts and filamentous fungi do not have adenosine deaminase acting on RNA (ADAR) orthologs and are believed to lack A-to-I RNA editing, which is the most prevalent editing of mRNA in animals. However, during this study with the PUK1 (FGRRES_01058) pseudokinase gene important for sexual reproduction in Fusarium graminearum, we found that two tandem stop codons, UA1831GUA1834G, in its kinase domain were changed to UG1831GUG1834G by RNA editing in perithecia. To confirm A-to-I editing of PUK1 transcripts, strand-specific RNA-seq data were generated with RNA isolated from conidia, hyphae, and perithecia. PUK1 was almost specifically expressed in perithecia, and 90% of transcripts were edited to UG1831GUG1834G. Genome-wide analysis identified 26,056 perithecium-specific A-to-I editing sites. Unlike those in animals, 70.5% of A-to-I editing sites in F. graminearum occur in coding regions, and more than two-thirds of them result in amino acid changes, including editing of 69 PUK1-like pseudogenes with stop codons in ORFs. PUK1 orthologs and other pseudogenes also displayed stage-specific expression and editing in Neurospora crassa and F. verticillioides. Furthermore, F. graminearum differs from animals in the sequence preference and structure selectivity of A-to-I editing sites. Whereas A's embedded in RNA stems are targeted by ADARs, RNA editing in F. graminearum preferentially targets A's in hairpin loops, which is similar to the anticodon loop of tRNA targeted by adenosine deaminases acting on tRNA (ADATs). Overall, our results showed that A-to-I RNA editing occurs specifically during sexual reproduction and mainly in the coding regions in filamentous ascomycetes, involving adenosine deamination mechanisms distinct from metazoan ADARs. PMID:26934920

  14. RNA editing in Drosophila melanogaster: new targets and functionalconsequences

    Energy Technology Data Exchange (ETDEWEB)

    Stapleton, Mark; Carlson, Joseph W.; Celniker, Susan E.

    2006-09-05

    Adenosine deaminases that act on RNA (ADARs) catalyze the site-specific conversion of adenosine to inosine in primary mRNA transcripts. These re-coding events affect coding potential, splice-sites, and stability of mature mRNAs. ADAR is an essential gene and studies in mouse, C. elegans, and Drosophila suggest its primary function is to modify adult behavior by altering signaling components in the nervous system. By comparing the sequence of isogenic cDNAs to genomic DNA, we have identified and experimentally verified 27 new targets of Drosophila ADAR. Our analyses lead us to identify new classes of genes whose transcripts are targets of ADAR including components of the actin cytoskeleton, and genes involved in ion homeostasis and signal transduction. Our results indicate that editing in Drosophila increases the diversity of the proteome, and does so in a manner that has direct functional consequences on protein function.

  15. Mitochondrial tRNA 5'-editing in Dictyostelium discoideum and Polysphondylium pallidum.

    Science.gov (United States)

    Abad, Maria G; Long, Yicheng; Kinchen, R Dimitri; Schindel, Elinor T; Gray, Michael W; Jackman, Jane E

    2014-05-30

    Mitochondrial tRNA (mt-tRNA) 5'-editing was first described more than 20 years ago; however, the first candidates for 5'-editing enzymes were only recently identified in a eukaryotic microbe (protist), the slime mold Dictyostelium discoideum. In this organism, eight of 18 mt-tRNAs are predicted to be edited based on the presence of genomically encoded mismatched nucleotides in their aminoacyl-acceptor stem sequences. Here, we demonstrate that mt-tRNA 5'-editing occurs at all predicted sites in D. discoideum as evidenced by changes in the sequences of isolated mt-tRNAs compared with the expected sequences encoded by the mitochondrial genome. We also identify two previously unpredicted editing events in which G-U base pairs are edited in the absence of any other genomically encoded mismatches. A comparison of 5'-editing in D. discoideum with 5'-editing in another slime mold, Polysphondylium pallidum, suggests organism-specific idiosyncrasies in the treatment of U-G/G-U pairs. In vitro activities of putative D. discoideum editing enzymes are consistent with the observed editing reactions and suggest an overall lack of tRNA substrate specificity exhibited by the repair component of the editing enzyme. Although the presence of terminal mismatches in mt-tRNA sequences is highly predictive of the occurrence of mt-tRNA 5'-editing, the variability in treatment of U-G/G-U base pairs observed here indicates that direct experimental evidence of 5'-editing must be obtained to understand the complete spectrum of mt-tRNA editing events in any species. PMID:24737330

  16. Parallel Evolution and Lineage-Specific Expansion of RNA Editing in Ctenophores.

    Science.gov (United States)

    Kohn, Andrea B; Sanford, Rachel S; Yoshida, Masa-aki; Moroz, Leonid L

    2015-12-01

    RNA editing is a process of targeted alterations of nucleotides in all types of RNA molecules (e.g., rRNA, tRNA, mRNA, and miRNA). As a result, the transcriptional output differs from its genomic DNA template. RNA editing can be defined both by biochemical mechanisms and by enzymes that perform these reactions. There are high levels of RNA editing detected in the mammalian nervous system, suggesting that nervous systems use this mechanism to increase protein diversity, because the post-transcription modifications lead to new gene products with novel functions. By re-annotating the ctenophore genomes, we found that the number of predicted RNA-editing enzymes is comparable to the numbers in mammals, but much greater than in other non-bilaterian basal metazoans. However, the overall molecular diversity of RNA-editing enzymes in ctenophores is lower, suggesting a possible "compensation" by an expansion of the ADAT1-like subfamily in this lineage. In two genera of ctenophores, Pleurobrachia and Mnemiopsis, there are high levels of expression for RNA-editing enzymes in their aboral organs, the integrative center involved in control of locomotion and geotaxis. This finding supports the hypothesis that RNA editing is correlated with the complexity of tissues and behaviors. Smaller numbers of RNA-editing enzymes in Porifera and Placozoa also correlates with the primary absence of neural and muscular systems in these lineages. In ctenophores, the expansion of the RNA-editing machinery can also provide mechanisms that support the remarkable capacity for regeneration in these animals. In summary, despite their compact genomes, a wide variety of epigenomic mechanisms employed by ctenophores and other non-bilaterian basal metazoans can provide novel insights into the evolutionary origins of biological novelties. PMID:26089435

  17. Canonical A-to-I and C-to-U RNA editing is enriched at 3'UTRs and microRNA target sites in multiple mouse tissues.

    Directory of Open Access Journals (Sweden)

    Tongjun Gu

    Full Text Available RNA editing is a process that modifies RNA nucleotides and changes the efficiency and fidelity of the central dogma. Enzymes that catalyze RNA editing are required for life, and defects in RNA editing are associated with many diseases. Recent advances in sequencing have enabled the genome-wide identification of RNA editing sites in mammalian transcriptomes. Here, we demonstrate that canonical RNA editing (A-to-I and C-to-U occurs in liver, white adipose, and bone tissues of the laboratory mouse, and we show that apparent non-canonical editing (all other possible base substitutions is an artifact of current high-throughput sequencing technology. Further, we report that high-confidence canonical RNA editing sites can cause non-synonymous amino acid changes and are significantly enriched in 3' UTRs, specifically at microRNA target sites, suggesting both regulatory and functional consequences for RNA editing.

  18. TbRGG2 facilitates kinetoplastid RNA editing initiation and progression past intrinsic pause sites.

    Science.gov (United States)

    Ammerman, Michelle L; Presnyak, Vladimir; Fisk, John C; Foda, Bardees M; Read, Laurie K

    2010-11-01

    TbRGG2 is an essential kinetoplastid RNA editing accessory factor that acts specifically on pan-edited RNAs. To understand the mechanism of TbRGG2 action, we undertook an in-depth analysis of edited RNA populations in TbRGG2 knockdown cells and an in vitro examination of the biochemical activities of the protein. We demonstrate that TbRGG2 down-regulation more severely impacts editing at the 5' ends of pan-edited RNAs than at their 3' ends. The initiation of editing is reduced to some extent in TbRGG2 knockdown cells. In addition, TbRGG2 plays a post-initiation role as editing becomes stalled in TbRGG2-depleted cells, resulting in an overall decrease in the 3' to 5' progression of editing. Detailed analyses of edited RNAs from wild-type and TbRGG2-depleted cells reveal that TbRGG2 facilitates progression of editing past intrinsic pause sites that often correspond to the 3' ends of cognate guide RNAs (gRNAs). In addition, noncanonically edited junction regions are either absent or significantly shortened in TbRGG2-depleted cells, consistent with impaired gRNA transitions. Sequence analysis further suggests that TbRGG2 facilitates complete utilization of certain gRNAs. In vitro RNA annealing and in vivo RNA unwinding assays demonstrate that TbRGG2 can modulate RNA-RNA interactions. Collectively, these data are consistent with a model in which TbRGG2 facilitates initiation and 3' to 5' progression of editing through its ability to affect gRNA utilization, both during the transition between specific gRNAs and during usage of certain gRNAs. PMID:20855539

  19. RNA Editing Sites Exist in Protein-coding Genes in the Chloroplast Genome of Cycas taitungensis

    Institute of Scientific and Technical Information of China (English)

    Haiyan Chen; Likun Deng; Yuan Jiang; Ping Lu; Jianing Yu

    2011-01-01

    RNA editing is a post-transcriptional process that results in modifications of ribonucleotides at specific locations.In land plants editing can occur in both mitochondria and chloroplasts and most commonly involves C-to-U changes,especially in seed plants.Using prediction and experimental determination,we investigated RNA editing in 40 protein-coding genes from the chloroplast genome of Cycas taitungensis.A total of 85 editing sites were identified in 25 transcripts.Comparison analysis of the published editotypes of these 25 transcripts in eight species showed that RNA editing events gradually disappear during plant evolution.The editing in the first and third codon position disappeared quicker than that in the second codon position,ndh genes have the highest editing frequency while serine and proline codons were more frequently edited than the codons of other amino acids.These results imply that retained RNA editing sites have imbalanced distribution in genes and most of them may function by changing protein structure or interaction.Mitochondrion protein-coding genes have three times the editing sites compared with chloroplast genes of Cycas,most likely due to slower evolution speed.

  20. Reprogramming, Circular Reasoning and Self versus Non-self: One-Stop Shopping with RNA Editing

    Science.gov (United States)

    Savva, Yiannis A.; Rezaei, Ali; St. Laurent, Georges; Reenan, Robert A.

    2016-01-01

    Transcription of genetic information from archival DNA into RNA molecule working copies is vital for proper cellular function and is highly accurate. In turn, RNAs serve structural, enzymatic, and regulatory roles, as well as being informational templates for the ribosomal translation of proteins. Following RNA synthesis, maturing of RNA molecules occurs through various RNA processing events. One component of the collection of processes involving RNA species, broadly defined as RNA metabolism, is the RNA-editing pathway and is found in all animals. Acting specifically on RNA substrates with double-stranded character, RNA editing has been shown to regulate a plethora of genomic outputs, including gene recoding, RNA splicing, biogenesis and targeting actions of microRNAs and small interfering RNAs, and global gene expression. Recent evidence suggests that RNA modifications mediated via RNA editing influence the biogenesis of circular RNAs and safeguard against aberrant innate immune responses generated to endogenous RNA sources. These novel roles have the potential to contribute new insights into molecular mechanisms underlying pathogenesis mediated by mishandling of double-stranded RNA. Here, we discuss recent advances in the field, which highlight novel roles associated with the RNA-editing process and emphasize their importance during cellular RNA metabolism. In addition, we highlight the relevance of these newly discovered roles in the context of neurological disorders and the more general concept of innate recognition of self versus non-self. PMID:27458478

  1. Genetic algorithm learning as a robust approach to RNA editing site prediction

    Directory of Open Access Journals (Sweden)

    Gopal Shuba

    2006-03-01

    Full Text Available Abstract Background RNA editing is one of several post-transcriptional modifications that may contribute to organismal complexity in the face of limited gene complement in a genome. One form, known as C → U editing, appears to exist in a wide range of organisms, but most instances of this form of RNA editing have been discovered serendipitously. With the large amount of genomic and transcriptomic data now available, a computational analysis could provide a more rapid means of identifying novel sites of C → U RNA editing. Previous efforts have had some success but also some limitations. We present a computational method for identifying C → U RNA editing sites in genomic sequences that is both robust and generalizable. We evaluate its potential use on the best data set available for these purposes: C → U editing sites in plant mitochondrial genomes. Results Our method is derived from a machine learning approach known as a genetic algorithm. REGAL (RNA Editing site prediction by Genetic Algorithm Learning is 87% accurate when tested on three mitochondrial genomes, with an overall sensitivity of 82% and an overall specificity of 91%. REGAL's performance significantly improves on other ab initio approaches to predicting RNA editing sites in this data set. REGAL has a comparable sensitivity and higher specificity than approaches which rely on sequence homology, and it has the advantage that strong sequence conservation is not required for reliable prediction of edit sites. Conclusion Our results suggest that ab initio methods can generate robust classifiers of putative edit sites, and we highlight the value of combinatorial approaches as embodied by genetic algorithms. We present REGAL as one approach with the potential to be generalized to other organisms exhibiting C → U RNA editing.

  2. Role of tRNAPro in pretransfer editing of alanine by prolyl-tRNA synthetase

    Directory of Open Access Journals (Sweden)

    Boyarshin K. S.

    2013-09-01

    Full Text Available Aim. To characterize the process of tRNA-dependent pretransfer edi- ting of alanine by prolyl-tRNA synthetase of bacteria Enterococcus faecalis (ProRSEf. Methods. Velocity of the editing processes in vitro was determined by ATP hydrolysis by ProRSEf. Pretransfer and posttransfer editing were experimentally separated by site-directed mutagenesis. Results. tRNA-dependent pretransfer editing is characterized by three-fold larger velocity then tRNA-independent editing. Effectivity of the process depends on the presence of 2'-hydroxyle group of A76 tRNAPro. In the absence of tRNAPro selective release of alanyl-AMP occurs simultaneously with tRNA-independent pretransfer editing. Released alanyl-AMP can be re-bound and hydrolyzed. Conclusions. tRNA-dependent pretransfer editing of alanine by ProRSEf is the catalytic mechanism, mediated by 2'-hydroxyl group of A76 tRNAPro. In the absence of tRNAPro tRNA-independent pretransfer editing and selective release of alanyl-AMP occur.

  3. Improved Computational Target Site Prediction for Pentatricopeptide Repeat RNA Editing Factors

    OpenAIRE

    TAKENAKA, MIZUKI; Zehrmann, Anja; Brennicke, Axel; Graichen, Knut

    2013-01-01

    Pentatricopeptide repeat (PPR) proteins with an E domain have been identified as specific factors for C to U RNA editing in plant organelles. These PPR proteins bind to a unique sequence motif 5′ of their target editing sites. Recently, involvement of a combinatorial amino acid code in the P (normal length) and S type (short) PPR domains in sequence specific RNA binding was reported. PPR proteins involved in RNA editing, however, contain not only P and S motifs but also their long variants L ...

  4. RNA-Editing with Combined Insertion and Deletion Preserves Regularity

    Directory of Open Access Journals (Sweden)

    E.P. de Vink

    2013-01-01

    Full Text Available We consider two elementary forms of string rewriting called guided insertion/deletion and guided rewriting. The original strings are modified depending on the match with a given set of auxiliary strings, called guides. Guided insertion/deletion considers matching of a string and a guide with respect to a specific correspondence of strings. Guided rewriting considers matching of a string and a guide with respect to an equivalence relation on the alphabet. Guided insertion/deletion is inspired by {RNA}-editing, a biological process by which the original genetic information stored in DNA is modified before its final expression. The formalism here allows for simultaneous insertion and deletion of string elements. Guided rewriting, based on a letter-to-letter relation, is technically more appealing than guided insertion/deletion. We prove that guided rewriting preserves regularity: for every regular language its closure under guided rewriting is regular too. In the proof we will rely on the auxiliary notion of a slice sequence. We establish a correspondence of slice sequences and guide rewrite sequences. Because of their left-to-right nature, slice sequences are more convenient to deal with than guided rewrite sequences in the construction of the finite automata that we encounter in the proofs of regularity. Based on the result for guided rewriting we establish that guided insertion/deletion preserves regularity as well.

  5. A distant cis acting intronic element induces site-selective RNA editing

    DEFF Research Database (Denmark)

    Daniel, Chammiran; Venø, Morten Trillingsgaard; Ekdahl, Ylva;

    2012-01-01

    requirements for editing at the I/M site in the Gabra-3 transcript of the GABA(A) receptor. We identify an evolutionarily conserved intronic duplex, 150 nt downstream of the exonic hairpin where the I/M site resides, which is required for its editing. This is the first time a distant RNA structure has been...

  6. A-to-I RNA editing: A new mechanism of genomic information modification

    Institute of Scientific and Technical Information of China (English)

    2003-01-01

    A-to-I RNA editing, the important event of gene modification, which takes place at post-transcriptional level, was firstly reported in 1991. The molecular mechanism of A-to-I RNA editing involves site-selective deamination of adenosine to inosine in pre-mRNA, which leads to altering translation codons and splicing in nuclear transcripts, thereby functionally distinct proteins can be produced from a single gene. The mammalian editing enzymes ADARs (adenosine deaminases acting on RNA) are widely expressed in brain and other tissues, however, up to date their substrates are mainly found in the central nervous system. It has recently been noticed that imperfect editing of these RNA substrates play critical roles in corresponding diseases, indicating that A-to-I RNA editing may be quite important in physiological or pathophysiological processes. Finding more new substrates of ADARs, especially in peripheral tissues, and performing functional research on new genes will be helpful to elucidate the biological significance of A-to-I RNA editing.

  7. Splice-mediated insertion of an Alu sequence inactivates ornithine δ-aminotransferase: A role for Alu elements in human mutation

    International Nuclear Information System (INIS)

    In studies of mutations causing deficiency of ornithine δ-aminotransferase the authors found an allele whose mature mRNA has a 142-nucleotide insertion at the junction of sequences from exons 3 and 4. The insert derives from an Alu element in ornithine δ-aminotransferase intron 3 oriented in the direction opposite to transcription (an antisense Alu). A guanine → cytosine transversion creates a donor splice site in this Alu, activating a cryptic acceptor splice site at its 5' end and causing splice-mediated insertion of an Alu fragment into the mature ornithine-δ-aminotransferase mRNA. The authors note that the complement of the Alu consensus sequence has at least two cryptic acceptor sites and several potential donor sequences and predict that similar mutations will be found in other genes

  8. Boosting CRISPR/Cas9 multiplex editing capability with the endogenous tRNA-processing system

    OpenAIRE

    Xie, Kabin; Minkenberg, Bastian; Yang, Yinong

    2015-01-01

    The clustered regularly interspaced short palindromic repeat (CRISPR)/CRISPR-associated protein 9 nuclease (Cas9) system has recently emerged as an efficient and versatile tool for genome editing in various organisms. However, its targeting capability and multiplex editing efficiency are often limited by the guide RNA (gRNA)-expressing device. This study demonstrates a general strategy and platform for precise processing and efficient production of numerous gRNAs in vivo from a synthetic poly...

  9. Genome-Wide Characterization of RNA Editing in Chicken Embryos Reveals Common Features among Vertebrates

    Science.gov (United States)

    Frésard, Laure; Leroux, Sophie; Roux, Pierre-François; Klopp, Christophe; Fabre, Stéphane; Esquerré, Diane; Dehais, Patrice; Djari, Anis; Gourichon, David

    2015-01-01

    RNA editing results in a post-transcriptional nucleotide change in the RNA sequence that creates an alternative nucleotide not present in the DNA sequence. This leads to a diversification of transcription products with potential functional consequences. Two nucleotide substitutions are mainly described in animals, from adenosine to inosine (A-to-I) and from cytidine to uridine (C-to-U). This phenomenon is described in more details in mammals, notably since the availability of next generation sequencing technologies allowing whole genome screening of RNA-DNA differences. The number of studies recording RNA editing in other vertebrates like chicken is still limited. We chose to use high throughput sequencing technologies to search for RNA editing in chicken, and to extend the knowledge of its conservation among vertebrates. We performed sequencing of RNA and DNA from 8 embryos. Being aware of common pitfalls inherent to sequence analyses that lead to false positive discovery, we stringently filtered our datasets and found fewer than 40 reliable candidates. Conservation of particular sites of RNA editing was attested by the presence of 3 edited sites previously detected in mammals. We then characterized editing levels for selected candidates in several tissues and at different time points, from 4.5 days of embryonic development to adults, and observed a clear tissue-specificity and a gradual increase of editing level with time. By characterizing the RNA editing landscape in chicken, our results highlight the extent of evolutionary conservation of this phenomenon within vertebrates, attest to its tissue and stage specificity and provide support of the absence of non A-to-I events from the chicken transcriptome. PMID:26024316

  10. Loss of matK RNA editing in seed plant chloroplasts

    Directory of Open Access Journals (Sweden)

    Maier Uwe G

    2009-08-01

    Full Text Available Abstract Background RNA editing in chloroplasts of angiosperms proceeds by C-to-U conversions at specific sites. Nuclear-encoded factors are required for the recognition of cis-elements located immediately upstream of editing sites. The ensemble of editing sites in a chloroplast genome differs widely between species, and editing sites are thought to evolve rapidly. However, large-scale analyses of the evolution of individual editing sites have not yet been undertaken. Results Here, we analyzed the evolution of two chloroplast editing sites, matK-2 and matK-3, for which DNA sequences from thousands of angiosperm species are available. Both sites are found in most major taxa, including deep-branching families such as the nymphaeaceae. However, 36 isolated taxa scattered across the entire tree lack a C at one of the two matK editing sites. Tests of several exemplary species from this in silico analysis of matK processing unexpectedly revealed that one of the two sites remain unedited in almost half of all species examined. A comparison of sequences between editors and non-editors showed that specific nucleotides co-evolve with the C at the matK editing sites, suggesting that these nucleotides are critical for editing-site recognition. Conclusion (i Both matK editing sites were present in the common ancestor of all angiosperms and have been independently lost multiple times during angiosperm evolution. (ii The editing activities corresponding to matK-2 and matK-3 are unstable. (iii A small number of third-codon positions in the vicinity of editing sites are selectively constrained independent of the presence of the editing site, most likely because of interacting RNA-binding proteins.

  11. Editing of HIV-1 RNA by the double-stranded RNA deaminase ADAR1 stimulates viral infection

    Science.gov (United States)

    Doria, Margherita; Neri, Francesca; Gallo, Angela; Farace, Maria Giulia; Michienzi, Alessandro

    2009-01-01

    Adenosine deaminases that act on dsRNA (ADARs) are enzymes that target double-stranded regions of RNA converting adenosines into inosines (A-to-I editing) thus contributing to genome complexity and fine regulation of gene expression. It has been described that a member of the ADAR family, ADAR1, can target viruses and affect their replication process. Here we report evidence showing that ADAR1 stimulates human immuno deficiency virus type 1 (HIV-1) replication by using both editing-dependent and editing-independent mechanisms. We show that over-expression of ADAR1 in HIV-1 producer cells increases viral protein accumulation in an editing-independent manner. Moreover, HIV-1 virions generated in the presence of over-expressed ADAR1 but not an editing-inactive ADAR1 mutant are released more efficiently and display enhanced infectivity, as demonstrated by challenge assays performed with T cell lines and primary CD4+ T lymphocytes. Finally, we report that ADAR1 associates with HIV-1 RNAs and edits adenosines in the 5′ untranslated region (UTR) and the Rev and Tat coding sequence. Overall these results suggest that HIV-1 has evolved mechanisms to take advantage of specific RNA editing activity of the host cell and disclose a stimulatory function of ADAR1 in the spread of HIV-1. PMID:19651874

  12. Multiplex single-base extension typing to identify nuclear genes required for RNA editing in plant organelles

    OpenAIRE

    Takenaka, Mizuki; Brennicke, Axel

    2008-01-01

    We developed a multiplex single-base extension single-nucleotide polymorphism-typing procedure for screening large numbers of plants for mutations in mitochondrial RNA editing. The high sensitivity of the approach detects changes in the RNA editing status generated in total cellular cDNA from pooled RNA preparations of up to 50 green plants. The method has been employed to tag several nuclear encoded genes required for RNA editing at specific sites in mitochondria of Arabidopsis thaliana. Thi...

  13. A distant cis acting intronic element induces site-selective RNA editing.

    Science.gov (United States)

    Daniel, Chammiran; Venø, Morten T; Ekdahl, Ylva; Kjems, Jørgen; Öhman, Marie

    2012-10-01

    Transcripts have been found to be site selectively edited from adenosine-to-inosine (A-to-I) in the mammalian brain, mostly in genes involved in neurotransmission. While A-to-I editing occurs at double-stranded structures, other structural requirements are largely unknown. We have investigated the requirements for editing at the I/M site in the Gabra-3 transcript of the GABA(A) receptor. We identify an evolutionarily conserved intronic duplex, 150 nt downstream of the exonic hairpin where the I/M site resides, which is required for its editing. This is the first time a distant RNA structure has been shown to be important for A-to-I editing. We demonstrate that the element also can induce editing in related but normally not edited RNA sequences. In human, thousands of genes are edited in duplexes formed by inverted repeats in non-coding regions. It is likely that numerous such duplexes can induce editing of coding regions throughout the transcriptome. PMID:22848101

  14. Glycine receptors caught between genome and proteome - functional implications of RNA editing and splicing

    Directory of Open Access Journals (Sweden)

    Pascal Legendre

    2009-11-01

    Full Text Available Information processing in the brain requires a delicate balance between excitation and inhibition. Glycine receptors (GlyR are involved in inhibitory mechanisms mainly at a synaptic level, but potential novel roles for these receptors recently emerged due to the discovery of posttranscriptional processing. GLR transcripts are edited through enzymatic modification of a single nucleotide leading to amino acid substitution within the neurotransmitter binding domain. RNA editing produces gain-of-function receptors well suited for generation and maintenance of tonic inhibition of neuronal excitability. As neuronal activity deprivation in early stages of development or in epileptic tissue is detrimental to neurons and because RNA editing of GlyR is up-regulated in temporal lobe epilepsy patients with a severe course of disease a pathophysiological role of these receptors emerges. This review contains a state-of-the-art discussion of (pathophysiological implications of GlyR RNA editing.

  15. Distinct role of Arabidopsis mitochondrial P-type pentatricopeptide repeat protein-modulating editing protein, PPME, in nad1 RNA editing.

    Science.gov (United States)

    Leu, Kuan-Chieh; Hsieh, Ming-Hsiun; Wang, Huei-Jing; Hsieh, Hsu-Liang; Jauh, Guang-Yuh

    2016-06-01

    The mitochondrion is an important power generator in most eukaryotic cells. To preserve its function, many essential nuclear-encoded factors play specific roles in mitochondrial RNA metabolic processes, including RNA editing. RNA editing consists of post-transcriptional deamination, which alters specific nucleotides in transcripts to mediate gene expression. In plant cells, many pentatricopeptide repeat proteins (PPRs) participate in diverse organellar RNA metabolic processes, but only PLS-type PPRs are involved in RNA editing. Here, we report a P-type PPR protein from Arabidopsis thaliana, P-type PPR-Modulating Editing (PPME), which has a distinct role in mitochondrial nad1 RNA editing via RNA binding activity. In the homozygous ppme mutant, cytosine (C)-to-uracil (U) conversions at both the nad1-898 and 937 sites were abolished, disrupting Arg(300)-to-Trp(300) and Pro(313)-to-Ser(313) amino acid changes in the mitochondrial NAD1 protein. NAD1 is a critical component of mitochondrial respiration complex I; its activity is severely reduced in the homozygous ppme mutant, resulting in significantly altered growth and development. Both abolished RNA editing and defective complex I activity were completely rescued by CaMV 35S promoter- and PPME native promoter-driven PPME genomic fragments tagged with GFP in a homozygous ppme background. Our experimental results demonstrate a distinct role of a P-type PPR protein, PPME, in RNA editing in plant organelles. PMID:27149614

  16. A computational screen for site selective A-to-I editing detects novel sites in neuron specific Hu proteins

    Directory of Open Access Journals (Sweden)

    Furey Terrence S

    2010-01-01

    Full Text Available Abstract Background Several bioinformatic approaches have previously been used to find novel sites of ADAR mediated A-to-I RNA editing in human. These studies have discovered thousands of genes that are hyper-edited in their non-coding intronic regions, especially in alu retrotransposable elements, but very few substrates that are site-selectively edited in coding regions. Known RNA edited substrates suggest, however, that site selective A-to-I editing is particularly important for normal brain development in mammals. Results We have compiled a screen that enables the identification of new sites of site-selective editing, primarily in coding sequences. To avoid hyper-edited repeat regions, we applied our screen to the alu-free mouse genome. Focusing on the mouse also facilitated better experimental verification. To identify candidate sites of RNA editing, we first performed an explorative screen based on RNA structure and genomic sequence conservation. We further evaluated the results of the explorative screen by determining which transcripts were enriched for A-G mismatches between the genomic template and the expressed sequence since the editing product, inosine (I, is read as guanosine (G by the translational machinery. For expressed sequences, we only considered coding regions to focus entirely on re-coding events. Lastly, we refined the results from the explorative screen using a novel scoring scheme based on characteristics for known A-to-I edited sites. The extent of editing in the final candidate genes was verified using total RNA from mouse brain and 454 sequencing. Conclusions Using this method, we identified and confirmed efficient editing at one site in the Gabra3 gene. Editing was also verified at several other novel sites within candidates predicted to be edited. Five of these sites are situated in genes coding for the neuron-specific RNA binding proteins HuB and HuD.

  17. Cas9 gRNA engineering for genome editing, activation and repression

    OpenAIRE

    Kiani, Samira; Chavez, Alejandro; Tuttle, Marcelle; Hall, Richard N; Chari, Raj; Ter-Ovanesyan, Dmitry; Qian, Jason; Pruitt, Benjamin W.; Beal, Jacob; Vora, Suhani; Buchthal, Joanna; Kowal, Emma J K; Ebrahimkhani, Mohammad R.; James J Collins; Weiss, Ron

    2015-01-01

    We demonstrate that by altering the length of Cas9-associated guide RNA(gRNA) we were able to control Cas9 nuclease activity and simultaneously perform genome editing and transcriptional regulation with a single Cas9 protein. We exploited these principles to engineer mammalian synthetic circuits with combined transcriptional regulation and kill functions governed by a single multifunctional Cas9 protein.

  18. Editing of HIV-1 RNA by the double-stranded RNA deaminase ADAR1 stimulates viral infection

    OpenAIRE

    Doria, Margherita; Neri, Francesca; Gallo, Angela; Farace, Maria Giulia; Michienzi, Alessandro

    2009-01-01

    Adenosine deaminases that act on dsRNA (ADARs) are enzymes that target double-stranded regions of RNA converting adenosines into inosines (A-to-I editing) thus contributing to genome complexity and fine regulation of gene expression. It has been described that a member of the ADAR family, ADAR1, can target viruses and affect their replication process. Here we report evidence showing that ADAR1 stimulates human immuno deficiency virus type 1 (HIV-1) replication by using both editing-dependent ...

  19. A Novel Computational Strategy to Identify A-to-I RNA Editing Sites by RNA-Seq Data: De Novo Detection in Human Spinal Cord Tissue

    Science.gov (United States)

    Picardi, Ernesto; Gallo, Angela; Galeano, Federica; Tomaselli, Sara; Pesole, Graziano

    2012-01-01

    RNA editing is a post-transcriptional process occurring in a wide range of organisms. In human brain, the A-to-I RNA editing, in which individual adenosine (A) bases in pre-mRNA are modified to yield inosine (I), is the most frequent event. Modulating gene expression, RNA editing is essential for cellular homeostasis. Indeed, its deregulation has been linked to several neurological and neurodegenerative diseases. To date, many RNA editing sites have been identified by next generation sequencing technologies employing massive transcriptome sequencing together with whole genome or exome sequencing. While genome and transcriptome reads are not always available for single individuals, RNA-Seq data are widespread through public databases and represent a relevant source of yet unexplored RNA editing sites. In this context, we propose a simple computational strategy to identify genomic positions enriched in novel hypothetical RNA editing events by means of a new two-steps mapping procedure requiring only RNA-Seq data and no a priori knowledge of RNA editing characteristics and genomic reads. We assessed the suitability of our procedure by confirming A-to-I candidates using conventional Sanger sequencing and performing RNA-Seq as well as whole exome sequencing of human spinal cord tissue from a single individual. PMID:22957051

  20. The expression of apoB mRNA editing factors is not the sole determinant for the induction of editing in differentiating Caco-2 cells

    International Nuclear Information System (INIS)

    Apolipoprotein B mRNA is edited at cytidine 6666 in the enterocytes lining the small intestine of all mammals; converting a CAA codon to a UAA stop codon. The conversion is ∼80% efficient in this tissue and leads to the expression of the truncated protein, ApoB48, essential for secretion of dietary lipid as chylomicrons. Caco-2 cell raft cultures have been used as an in vitro model for the induction of editing activity during human small intestinal cell differentiation. This induction of apoB mRNA editing has been ascribed to the expression of APOBEC-1. In agreement our data demonstrated differentiation-dependent induction of expression of the editing enzyme APOBEC-1 and in addition we show alternative splicing of the essential auxiliary factor ACF. However, transfection of these editing factors in undifferentiated proliferating Caco-2 cells was not sufficient to induce robust apoB mRNA editing activity. Only differentiation of Caco-2 cells could induce more physiological like levels of apoB mRNA editing. The data suggested that additional regulatory mechanism(s) were induced by differentiation that controlled the functional activity of editing factors.

  1. A potential role for NF1 mRNA editing in the pathogenesis of NF1 tumors

    Energy Technology Data Exchange (ETDEWEB)

    Cappione, A.J.; French, B.L.; Skuse, G.R. [Univ. of Rochester School of Medicine and Dentistry, NY (United States)

    1997-02-01

    Neurofibromatosis type I (NF1) is a common disorder that predisposes to neoplasia in tissues derived from the embryonic neural crest. The NF1 gene encodes a tumor suppressor that most likely acts through the interaction of its GTPase-activating protein (GAP)-related domain (GRD) with the product of the ras protooncogene. We have previously identified a site in the NF1 mRNA, within the first half of the NF1 GRD, which undergoes base-modification editing. Editing at that site changes a C to a U, thereby introducing an in-frame stop codon. NF1 RNA editing has been detected in all cell types studied, to date. In order to investigate the role played by editing in NF1 tumorigenesis, we analyzed RNA from 19 NF1 and 4 non-NF1 tumors. We observed varying levels of NF1 mRNA editing in different tumors, with a higher range of editing levels in more malignant tumors (e.g., neurofibrosarcomas) compared to benign tumors (cutaneous neurofibromas). Plexiform neurofibromas have an intermediate range of levels of NF1 mRNA editing. We also compared tumor and nontumor tissues from several NF1 individuals, to determine the extent of variability present in the constitutional levels of NF1 mRNA editing and to determine whether higher levels are present in tumors. The constitutional levels of NF1 mRNA editing varied slightly but were consistent with the levels observed in non-NF1 individuals. In every case, there was a greater level of NF1 mRNA editing in the tumor than in the nontumor tissue from the same patient. These results suggest that inappropriately high levels of NF1 mRNA editing does play a role in NF1 tumorigenesis and that editing may result in the functional equivalent of biallelic inactivation of the NF1 tumor suppressor. 24 refs., 4 figs., 2 tabs.

  2. The Genomic Landscape and Clinical Relevance of A-to-I RNA Editing in Human Cancers | Office of Cancer Genomics

    Science.gov (United States)

    Adenosine-to-inosine (A-to-I) RNA editing is a widespread post-transcriptional mechanism, but its genomic landscape and clinical relevance in cancer have not been investigated systematically. We characterized the global A-to-I RNA editing profiles of 6,236 patient samples of 17 cancer types from The Cancer Genome Atlas and revealed a striking diversity of altered RNA-editing patterns in tumors relative to normal tissues. We identified an appreciable number of clinically relevant editing events, many of which are in noncoding regions.

  3. Impact of Alu repeats on the evolution of human p53 binding sites

    Directory of Open Access Journals (Sweden)

    Sirotin Michael V

    2011-01-01

    Full Text Available Abstract Background The p53 tumor suppressor protein is involved in a complicated regulatory network, mediating expression of ~1000 human genes. Recent studies have shown that many p53 in vivo binding sites (BSs reside in transposable repeats. The relationship between these BSs and functional p53 response elements (REs remains unknown, however. We sought to understand whether the p53 REs also reside in transposable elements and particularly in the most-abundant Alu repeats. Results We have analyzed ~160 functional p53 REs identified so far and found that 24 of them occur in repeats. More than half of these repeat-associated REs reside in Alu elements. In addition, using a position weight matrix approach, we found ~400,000 potential p53 BSs in Alu elements genome-wide. Importantly, these putative BSs are located in the same regions of Alu repeats as the functional p53 REs - namely, in the vicinity of Boxes A/A' and B of the internal RNA polymerase III promoter. Earlier nucleosome-mapping experiments showed that the Boxes A/A' and B have a different chromatin environment, which is critical for the binding of p53 to DNA. Here, we compare the Alu-residing p53 sites with the corresponding Alu consensus sequences and conclude that the p53 sites likely evolved through two different mechanisms - the sites overlapping with the Boxes A/A' were generated by CG → TG mutations; the other sites apparently pre-existed in the progenitors of several Alu subfamilies, such as AluJo and AluSq. The binding affinity of p53 to the Alu-residing sites generally correlates with the age of Alu subfamilies, so that the strongest sites are embedded in the 'relatively young' Alu repeats. Conclusions The primate-specific Alu repeats play an important role in shaping the p53 regulatory network in the context of chromatin. One of the selective factors responsible for the frequent occurrence of Alu repeats in introns may be related to the p53-mediated regulation of Alu

  4. The ADAR RNA editing enzyme controls neuronal excitability in Drosophila melanogaster.

    Science.gov (United States)

    Li, Xianghua; Overton, Ian M; Baines, Richard A; Keegan, Liam P; O'Connell, Mary A

    2014-01-01

    RNA editing by deamination of specific adenosine bases to inosines during pre-mRNA processing generates edited isoforms of proteins. Recoding RNA editing is more widespread in Drosophila than in vertebrates. Editing levels rise strongly at metamorphosis, and Adar(5G1) null mutant flies lack editing events in hundreds of CNS transcripts; mutant flies have reduced viability, severely defective locomotion and age-dependent neurodegeneration. On the other hand, overexpressing an adult dADAR isoform with high enzymatic activity ubiquitously during larval and pupal stages is lethal. Advantage was taken of this to screen for genetic modifiers; Adar overexpression lethality is rescued by reduced dosage of the Rdl (Resistant to dieldrin), gene encoding a subunit of inhibitory GABA receptors. Reduced dosage of the Gad1 gene encoding the GABA synthetase also rescues Adar overexpression lethality. Drosophila Adar(5G1) mutant phenotypes are ameliorated by feeding GABA modulators. We demonstrate that neuronal excitability is linked to dADAR expression levels in individual neurons; Adar-overexpressing larval motor neurons show reduced excitability whereas Adar(5G1) null mutant or targeted Adar knockdown motor neurons exhibit increased excitability. GABA inhibitory signalling is impaired in human epileptic and autistic conditions, and vertebrate ADARs may have a relevant evolutionarily conserved control over neuronal excitability. PMID:24137011

  5. New Insights into the Biological Role of Mammalian ADARs; the RNA Editing Proteins

    Directory of Open Access Journals (Sweden)

    Niamh Mannion

    2015-09-01

    Full Text Available The ADAR proteins deaminate adenosine to inosine in double-stranded RNA which is one of the most abundant modifications present in mammalian RNA. Inosine can have a profound effect on the RNAs that are edited, not only changing the base-pairing properties, but can also result in recoding, as inosine behaves as if it were guanosine. In mammals there are three ADAR proteins and two ADAR-related proteins (ADAD expressed. All have a very similar modular structure; however, both their expression and biological function differ significantly. Only two of the ADAR proteins have enzymatic activity. However, both ADAR and ADAD proteins possess the ability to bind double-strand RNA. Mutations in ADARs have been associated with many diseases ranging from cancer, innate immunity to neurological disorders. Here, we will discuss in detail the domain structure of mammalian ADARs, the effects of RNA editing, and the role of ADARs in human diseases.

  6. C-->U editing of apolipoprotein B mRNA in marsupials: identification and characterisation of APOBEC-1 from the American opossum Monodelphus domestica.

    OpenAIRE

    Fujino, T; Navaratnam, N; Jarmuz, A; von Haeseler, A; Scott, J

    1999-01-01

    The C->U editing of RNA is widely found in plant and animal species. In mammals it is a discrete process confined to the editing of apolipoprotein B (apoB) mRNA in eutherians and the editing of the mitochondrial tRNA for glycine in marsupials. Here we have identified and characterised apoB mRNA editing in the American opossum Monodelphus domestica. The apoB mRNA editing site is highly conserved in the opossum and undergoes complete editing in the small intestine, but not in the liver or other...

  7. Targeted Genome Editing of Sweet Orange Using Cas9/sgRNA

    OpenAIRE

    Hongge Jia; Nian Wang

    2014-01-01

    Genetic modification, including plant breeding, has been widely used to improve crop yield and quality, as well as to increase disease resistance. Targeted genome engineering is expected to contribute significantly to future varietal improvement, and genome editing technologies using zinc finger nucleases (ZFNs), transcription activator-like effector nucleases (TALENs), and clustered regularly interspaced short palindromic repeat (CRISPR)/Cas9/single guide RNA (sgRNA) have already been succes...

  8. Epigenome Editing of Potato by Grafting Using Transgenic Tobacco as siRNA Donor.

    Science.gov (United States)

    Kasai, Atsushi; Bai, Songling; Hojo, Hatsune; Harada, Takeo

    2016-01-01

    In plants, it is possible to induce heritable transcriptional gene silencing (TGS) via RNA-directed DNA methylation (RdDM) using artificially synthesized small RNA (siRNA) homologous to the 5'-flanking region of the target gene. As the siRNA signal with a specific RNA determinant moves through plasmodesmata and sieve elements, we attempted to induce TGS of a transgene and an endogenous gene of potato (Solanum tuberosum) rootstock by grafting using siRNA produced in a tobacco (Nicotiana benthamiana) scion. Our results provide evidence that this system can induce TGS of target genes in tubers formed on potato rootstock. The TGS is maintained in the progeny tubers lacking the transported siRNAs. Our findings reveal that epigenome editing using mobile RNA has the potential to allow breeding of artificial sport cultivars in vegetative propagation crops. PMID:27564864

  9. Rat prostatic steroid binding protein: characterisation of the Alu element upstream of the C3 genes.

    OpenAIRE

    Hurst, H C; Parker, M G

    1984-01-01

    We have characterised an Alu-like repetitive element found about 400 bp upstream of the gene encoding the C3 component of rat prostatic steroid binding protein and suggest, from comparisons with other published sequences, that it is an example of a third class of rodent Alu-equivalent sequences. Members of this class are 80-90 bp long, share greater than 90% sequence homology, and contain sequences resembling the RNA polymerase III bipartite promoter. The Alu type III element within the C3 ge...

  10. Dimeric CRISPR RNA-guided FokI nucleases for highly specific genome editing

    OpenAIRE

    Tsai, Shengdar Q.; Wyvekens, Nicolas; Khayter, Cyd; Foden, Jennifer A.; Thapar, Vishal; Reyon, Deepak; Goodwin, Mathew J.; Aryee, Martin J; Joung, J. Keith

    2014-01-01

    Monomeric CRISPR-Cas9 nucleases are widely used for targeted genome editing but can induce unwanted off-target mutations with high frequencies. Here we describe dimeric RNA-guided FokI Nucleases (RFNs) that recognize extended sequences and can edit endogenous genes with high efficiencies in human cells. The cleavage activity of an RFN depends strictly on the binding of two guide RNAs (gRNAs) to DNA with a defined spacing and orientation and therefore show improved specificities relative to wi...

  11. Dramatic Enhancement of Genome Editing by CRISPR/Cas9 Through Improved Guide RNA Design

    OpenAIRE

    Farboud, B; Meyer, BJ

    2015-01-01

    Success with genome editing by the RNA-programmed nuclease Cas9 has been limited by the inability to predict effective guide RNAs and DNA target sites. Not all guide RNAs have been successful, and even those that were, varied widely in their efficacy. Here we describe and validate a strategy for Caenorhabditis elegans that reliably achieved a high frequency of genome editing for all targets tested in vivo. The key innovation was to design guide RNAs with a GG motif at the 3′ end of their targ...

  12. Thiolation Controls Cytoplasmic tRNA Stability and Acts as a Negative Determinant for tRNA Editing in Mitochondria

    Czech Academy of Sciences Publication Activity Database

    Wohlgamuth-Benedum, J. M.; RUBIO, M. A. T.; Paris, Zdeněk; Long, Shaojun; Poliak, Pavel; Lukeš, Julius; Alfonzo, J. D.

    2009-01-01

    Roč. 284, č. 36 (2009), s. 23947-23953. ISSN 0021-9258 R&D Projects: GA ČR GA204/06/1558; GA MŠk LC07032; GA MŠk 2B06129 Institutional research plan: CEZ:AV0Z60220518 Keywords : T. brucei * tRNA * 2-thiolation * Fe-S cluster * editing Subject RIV: EB - Genetics ; Molecular Biology Impact factor: 5.328, year: 2009

  13. Redefining the structural motifs that determine RNA binding and RNA editing by pentatricopeptide repeat proteins in land plants.

    Science.gov (United States)

    Cheng, Shifeng; Gutmann, Bernard; Zhong, Xiao; Ye, Yongtao; Fisher, Mark F; Bai, Fengqi; Castleden, Ian; Song, Yue; Song, Bo; Huang, Jiaying; Liu, Xin; Xu, Xun; Lim, Boon L; Bond, Charles S; Yiu, Siu-Ming; Small, Ian

    2016-02-01

    The pentatricopeptide repeat (PPR) proteins form one of the largest protein families in land plants. They are characterised by tandem 30-40 amino acid motifs that form an extended binding surface capable of sequence-specific recognition of RNA strands. Almost all of them are post-translationally targeted to plastids and mitochondria, where they play important roles in post-transcriptional processes including splicing, RNA editing and the initiation of translation. A code describing how PPR proteins recognise their RNA targets promises to accelerate research on these proteins, but making use of this code requires accurate definition and annotation of all of the various nucleotide-binding motifs in each protein. We have used a structural modelling approach to define 10 different variants of the PPR motif found in plant proteins, in addition to the putative deaminase motif that is found at the C-terminus of many RNA-editing factors. We show that the super-helical RNA-binding surface of RNA-editing factors is potentially longer than previously recognised. We used the redefined motifs to develop accurate and consistent annotations of PPR sequences from 109 genomes. We report a high error rate in PPR gene models in many public plant proteomes, due to gene fusions and insertions of spurious introns. These consistently annotated datasets across a wide range of species are valuable resources for future comparative genomics studies, and an essential pre-requisite for accurate large-scale computational predictions of PPR targets. We have created a web portal (http://www.plantppr.com) that provides open access to these resources for the community. PMID:26764122

  14. Regulation of gene expression in neuronal tissue by RNA interference and editing

    DEFF Research Database (Denmark)

    Venø, Morten Trillingsgaard

    No tissue in the mammalian organism is more complex than the brain. This complexity is in part the result of precise timing and interplay of a large number mechanisms modulating gene expression post-transcriptionally. Fine-tuning mechanisms such as A-to-I editing of RNA transcripts and regulation...... mediated by microRNAs are crucial for the correct function of the mammalian brain. We are addressing A-to-I editing and regulation by microRNAs with spatio-temporal resolution in the embryonic porcine brain by Solexa sequencing of microRNAs and 454 sequencing of edited neuronal messenger RNAs, resulting in......RNAs, causing these transgenic mice to be less prone to cocaine addictive behavior. Another study demonstrated that abolishing the expression of histone methylases, GLP and G9a, increases the expression level of a large number of miRNAs. A possible feed-back mechanism is suggested, since a subset of these mi...

  15. Small RNA and A-to-I Editing in Autism Spectrum Disorders

    Science.gov (United States)

    Eran, Alal

    One in every 88 children is diagnosed with Autism Spectrum Disorders (ASDs), a set of neurodevelopmental conditions characterized by social impairments, communication deficits, and repetitive behavior. ASDs have a substantial genetic component, but the specific cause of most cases remains unknown. Understanding gene-environment interactions underlying ASD is essential for improving early diagnosis and identifying critical targets for intervention and prevention. Towards this goal, we surveyed adenosine-to-inosine (A-to-I) RNA editing in autistic brains. A-to-I editing is an epigenetic mechanism that fine-tunes synaptic function in response to environmental stimuli, shown to modulate complex behavior in animals. We used ultradeep sequencing to quantify A-to-I receding of candidate synaptic genes in postmortem cerebella from individuals with ASD and neurotypical controls. We found unexpectedly wide distributions of human A-to-I editing levels, whose extremes were consistently populated by individuals with ASD. We correlated A-to-I editing with isoform usage, identified clusters of correlated sites, and examined differential editing patterns. Importantly, we found that individuals with ASD commonly use a dysfunctional form of the editing enzyme ADARB1. We next profiled small RNAs thought to regulate A-to-I editing, which originate from one of the most commonly altered loci in ASD, 15q11. Deep targeted sequencing of SNORD115 and SNORD116 transcripts enabled their high-resolution detection in human brains, and revealed a strong gender bias underlying their expression. The consistent 2-fold upregulation of 15q11 small RNAs in male vs. female cerebella could be important in delineating the role of this locus in ASD, a male dominant disorder. Overall, these studies provide an accurate population-level view of small RNA and A-to-I editing in human cerebella, and suggest that A-to-I editing of synaptic genes may be informative for assessing the epigenetic risk for autism

  16. Dramatic enhancement of genome editing by CRISPR/Cas9 through improved guide RNA design.

    Science.gov (United States)

    Farboud, Behnom; Meyer, Barbara J

    2015-04-01

    Success with genome editing by the RNA-programmed nuclease Cas9 has been limited by the inability to predict effective guide RNAs and DNA target sites. Not all guide RNAs have been successful, and even those that were, varied widely in their efficacy. Here we describe and validate a strategy for Caenorhabditis elegans that reliably achieved a high frequency of genome editing for all targets tested in vivo. The key innovation was to design guide RNAs with a GG motif at the 3' end of their target-specific sequences. All guides designed using this simple principle induced a high frequency of targeted mutagenesis via nonhomologous end joining (NHEJ) and a high frequency of precise DNA integration from exogenous DNA templates via homology-directed repair (HDR). Related guide RNAs having the GG motif shifted by only three nucleotides showed severely reduced or no genome editing. We also combined the 3' GG guide improvement with a co-CRISPR/co-conversion approach. For this co-conversion scheme, animals were only screened for genome editing at designated targets if they exhibited a dominant phenotype caused by Cas9-dependent editing of an unrelated target. Combining the two strategies further enhanced the ease of mutant recovery, thereby providing a powerful means to obtain desired genetic changes in an otherwise unaltered genome. PMID:25695951

  17. Evidence for co-evolution between human microRNAs and Alu-repeats.

    Directory of Open Access Journals (Sweden)

    Stefan Lehnert

    Full Text Available This paper connects Alu repeats, the most abundant repetitive elements in the human genome and microRNAs, small RNAs that alter gene expression at the post-transcriptional level. Base-pair complementarity could be demonstrated between the seed sequence of a subset of human microRNAs and Alu repeats that are integrated parallel (sense in mRNAs. The most common target site coincides with the evolutionary most conserved part of Alu. A primate-specific gene cluster on chromosome 19 encodes the majority of miRNAs that target the most conserved sense Alu site. The individual miRNA genes within this cluster are flanked by an Alu-LINE signature, which has been duplicated with the clustered miRNA genes. Gene duplication events in this locus are supported by comparing repeat length variations of the LINE elements within the cluster with those in the rest of the chromosome. Thus, a dual relationship exists between an evolutionary young miRNA cluster and their Alu targets that may have evolved in the same time window. One hypothesis for this dual relationship is that these miRNAs could protect against too high rates of duplicative transposition, which would destroy the genome.

  18. CRISPR–Cas9-mediated genome editing and guide RNA design

    OpenAIRE

    Wiles, Michael V.; Qin, Wenning; Cheng, Albert W; Wang, Haoyi

    2015-01-01

    CRISPR and CRISPR-associated (Cas) proteins, which in nature comprise the RNA-based adaptive immune system in bacteria and archaea, have emerged as particularly powerful genome editing tools owing to their unrivaled ease of use and ability to modify genomes across mammalian model systems. As such, the CRISPR–Cas9 system holds promise as a “system of choice” for functional mammalian genetic studies across biological disciplines. Here we briefly review this fast moving field, introduce the CRIS...

  19. Dimeric CRISPR RNA-guided FokI nucleases for highly specific genome editing.

    Science.gov (United States)

    Tsai, Shengdar Q; Wyvekens, Nicolas; Khayter, Cyd; Foden, Jennifer A; Thapar, Vishal; Reyon, Deepak; Goodwin, Mathew J; Aryee, Martin J; Joung, J Keith

    2014-06-01

    Monomeric CRISPR-Cas9 nucleases are widely used for targeted genome editing but can induce unwanted off-target mutations with high frequencies. Here we describe dimeric RNA-guided FokI nucleases (RFNs) that can recognize extended sequences and edit endogenous genes with high efficiencies in human cells. RFN cleavage activity depends strictly on the binding of two guide RNAs (gRNAs) to DNA with a defined spacing and orientation substantially reducing the likelihood that a suitable target site will occur more than once in the genome and therefore improving specificities relative to wild-type Cas9 monomers. RFNs guided by a single gRNA generally induce lower levels of unwanted mutations than matched monomeric Cas9 nickases. In addition, we describe a simple method for expressing multiple gRNAs bearing any 5' end nucleotide, which gives dimeric RFNs a broad targeting range. RFNs combine the ease of RNA-based targeting with the specificity enhancement inherent to dimerization and are likely to be useful in applications that require highly precise genome editing. PMID:24770325

  20. Targeted genome editing of sweet orange using Cas9/sgRNA.

    Directory of Open Access Journals (Sweden)

    Hongge Jia

    Full Text Available Genetic modification, including plant breeding, has been widely used to improve crop yield and quality, as well as to increase disease resistance. Targeted genome engineering is expected to contribute significantly to future varietal improvement, and genome editing technologies using zinc finger nucleases (ZFNs, transcription activator-like effector nucleases (TALENs, and clustered regularly interspaced short palindromic repeat (CRISPR/Cas9/single guide RNA (sgRNA have already been successfully used to genetically modify plants. However, to date, there has been no reported use of any of the current genome editing approaches in sweet orange, an important fruit crop. In this study, we first developed a novel tool, Xcc-facilitated agroinfiltration, for enhancing transient protein expression in sweet orange leaves. We then successfully employed Xcc-facilitated agroinfiltration to deliver Cas9, along with a synthetic sgRNA targeting the CsPDS gene, into sweet orange. DNA sequencing confirmed that the CsPDS gene was mutated at the target site in treated sweet orange leaves. The mutation rate using the Cas9/sgRNA system was approximately 3.2 to 3.9%. Off-target mutagenesis was not detected for CsPDS-related DNA sequences in our study. This is the first report of targeted genome modification in citrus using the Cas9/sgRNA system-a system that holds significant promise for the study of citrus gene function and for targeted genetic modification.

  1. ADAR2 affects mRNA coding sequence edits with only modest effects on gene expression or splicing in vivo.

    Science.gov (United States)

    Dillman, Allissa A; Cookson, Mark R; Galter, Dagmar

    2016-01-01

    Adenosine deaminases bind double stranded RNA and convert adenosine to inosine. Editing creates multiple isoforms of neurotransmitter receptors, such as with Gria2. Adar2 KO mice die of seizures shortly after birth, but if the Gria2 Q/R editing site is mutated to mimic the edited version then the animals are viable. We performed RNA-Seq on frontal cortices of Adar2(-/-) Gria2(R/R) mice and littermates. We found 56 editing sites with significantly diminished editing levels in Adar2 deficient animals with the majority in coding regions. Only two genes and 3 exons showed statistically significant differences in expression levels. This work illustrates that ADAR2 is important in site-specific changes of protein coding sequences but has relatively modest effects on gene expression and splicing in the adult mouse frontal cortex. PMID:26669816

  2. Modulation of Aminoacylation and Editing Properties of Leucyl-tRNA Synthetase by a Conserved Structural Module.

    Science.gov (United States)

    Yan, Wei; Ye, Qing; Tan, Min; Chen, Xi; Eriani, Gilbert; Wang, En-Duo

    2015-05-01

    A conserved structural module following the KMSKS catalytic loop exhibits α-α-β-α topology in class Ia and Ib aminoacyl-tRNA synthetases. However, the function of this domain has received little attention. Here, we describe the effect this module has on the aminoacylation and editing capacities of leucyl-tRNA synthetases (LeuRSs) by characterizing the key residues from various species. Mutation of highly conserved basic residues on the third α-helix of this domain impairs the affinity of LeuRS for the anticodon stem of tRNA(Leu), which decreases both aminoacylation and editing activities. Two glycine residues on this α-helix contribute to flexibility, leucine activation, and editing of LeuRS from Escherichia coli (EcLeuRS). Acidic residues on the β-strand enhance the editing activity of EcLeuRS and sense the size of the tRNA(Leu) D-loop. Incorporation of these residues stimulates the tRNA-dependent editing activity of the chimeric minimalist enzyme Mycoplasma mobile LeuRS fused to the connective polypeptide 1 editing domain and leucine-specific domain from EcLeuRS. Together, these results reveal the stem contact-fold to be a functional as well as a structural linker between the catalytic site and the tRNA binding domain. Sequence comparison of the EcLeuRS stem contact-fold domain with editing-deficient enzymes suggests that key residues of this module have evolved an adaptive strategy to follow the editing functions of LeuRS. PMID:25817995

  3. RNA interference analyses suggest a transcript-specific regulatory role for mitochondrial RNA-binding proteins MRP1 and MRP2 in RNA editing and other RNA processing in Trypanosoma brucei

    Czech Academy of Sciences Publication Activity Database

    Horáková, Eva; Van Den Burg, J.; Zíková, Alena; Ernst, N. L.; Stuart, K.; Benne, R.; Lukeš, Julius

    2005-01-01

    Roč. 280, č. 4 (2005), s. 2429-2438. ISSN 0021-9258 R&D Projects: GA AV ČR IAA6022903 Institutional research plan: CEZ:AV0Z60220518 Keywords : Trypanosoma brucei * RNA editing * interference RNA Subject RIV: EB - Genetics ; Molecular Biology Impact factor: 5.854, year: 2005

  4. Editing for an AMPA receptor subunit RNA in prefrontal cortex and striatum in Alzheimer's disease, Huntington's disease and schizophrenia

    Science.gov (United States)

    Akbarian, S.; Smith, M. A.; Jones, E. G.; Bloom, F. E. (Principal Investigator)

    1995-01-01

    Animal studies and cell culture experiments demonstrated that posttranscriptional editing of the transcript of the GluR-2 gene, resulting in substitution of an arginine for glutamine in the second transmembrane region (TM II) of the expressed protein, is associated with a reduction in Ca2+ permeability of the receptor channel. Thus, disturbances in GluR-2 RNA editing with alteration of intracellular Ca2+ homeostasis could lead to neuronal dysfunction and even neuronal degeneration. The present study determined the proportions of edited and unedited GluR-2 RNA in the prefrontal cortex of brains from patients with Alzheimer's disease, in the striatum of brains from patients with Huntington's disease, and in the same areas of brains from age-matched schizophrenics and controls, by using reverse transcriptase-polymerase chain reaction, restriction endonuclease digestion, gel electrophoresis and scintillation radiometry. In the prefrontal cortex of controls, 99.9% were edited; in the prefrontal cortex both of schizophrenics and of Alzheimer's patients approximately 1.0% of all GluR-2 RNA molecules were unedited and 99% were edited. In the striatum of controls and of schizophrenics, approximately 0.5% of GluR-2 RNA molecules were unedited and 99.5% were edited; in the striatum of Huntington's patients nearly 5.0% of GluR-2 RNA was unedited. In the prefrontal white matter of controls, approximately 7.0% of GluR-2 RNA was unedited. In the normal human prefrontal cortex and striatum, the large majority of GluR-2 RNA molecules contains a CGG codon for arginine in the TMII coding region; this implies that the corresponding AMPA receptors have a low Ca2+ permeability, as previously demonstrated for the rat brain. The process of GluR-2 RNA editing is compromised in a region-specific manner in schizophrenia, in Alzheimer's disease and Huntington's Chorea although in each of these disorders there is still a large excess of edited GluR-2 RNA molecules. Disturbances of GluR-2 RNA

  5. Analysis of the Resistance Mechanism of a Benzoxaborole Inhibitor Reveals Insight into the Leucyl-tRNA Synthetase Editing Mechanism.

    Science.gov (United States)

    Zhao, Hanchao; Palencia, Andres; Seiradake, Elena; Ghaemi, Zhaleh; Cusack, Stephen; Luthey-Schulten, Zaida; Martinis, Susan

    2015-10-16

    A new class of antimicrobial benzoxaborole compounds was identified as a potent inhibitor of leucyl-tRNA synthetase (LeuRS) and therefore of protein synthesis. In a novel mechanism, AN2690 (5-fluoro-1,3-dihydro-1-hydroxy-2,1-benzoxaborole) blocks fungal cytoplasmic LeuRS by covalently trapping tRNA(Leu) in the editing site of the enzyme's CP1 domain. However, some resistant mutation sites are located outside of the CP1 hydrolytic editing active site. Thus, their mode of action that undermines drug inhibition was not understood. A combination of X-ray crystallography, molecular dynamics, metadynamics, biochemical experiments, and mutational analysis of a distal benzoxaborole-resistant mutant uncovered a eukaryote-specific tyrosine "switch" that is critical to tRNA-dependent post-transfer editing. The tyrosine "switch" has three states that shift between interactions with a lysine and the 3'-hydroxyl of the tRNA terminus, to inhibit or promote post-transfer editing. The oxaborole's mechanism of action capitalizes upon one of these editing active site states. This tunable editing mechanism in eukaryotic and archaeal LeuRSs is proposed to facilitate precise quality control of aminoacylation fidelity. These mechanistic distinctions could also be capitalized upon for development of the benzoxaboroles as a broad spectrum antibacterial. PMID:26172575

  6. 4SALE – A tool for synchronous RNA sequence and secondary structure alignment and editing

    Directory of Open Access Journals (Sweden)

    Schultz Jörg

    2006-11-01

    Full Text Available Abstract Background In sequence analysis the multiple alignment builds the fundament of all proceeding analyses. Errors in an alignment could strongly influence all succeeding analyses and therefore could lead to wrong predictions. Hand-crafted and hand-improved alignments are necessary and meanwhile good common practice. For RNA sequences often the primary sequence as well as a secondary structure consensus is well known, e.g., the cloverleaf structure of the t-RNA. Recently, some alignment editors are proposed that are able to include and model both kinds of information. However, with the advent of a large amount of reliable RNA sequences together with their solved secondary structures (available from e.g. the ITS2 Database, we are faced with the problem to handle sequences and their associated secondary structures synchronously. Results 4SALE fills this gap. The application allows a fast sequence and synchronous secondary structure alignment for large data sets and for the first time synchronous manual editing of aligned sequences and their secondary structures. This study describes an algorithm for the synchronous alignment of sequences and their associated secondary structures as well as the main features of 4SALE used for further analyses and editing. 4SALE builds an optimal and unique starting point for every RNA sequence and structure analysis. Conclusion 4SALE, which provides an user-friendly and intuitive interface, is a comprehensive toolbox for RNA analysis based on sequence and secondary structure information. The program connects sequence and structure databases like the ITS2 Database to phylogeny programs as for example the CBCAnalyzer. 4SALE is written in JAVA and therefore platform independent. The software is freely available and distributed from the website at http://4sale.bioapps.biozentrum.uni-wuerzburg.de

  7. Alu retrotransposons promote differentiation of human carcinoma cells through the aryl hydrocarbon receptor.

    Science.gov (United States)

    Morales-Hernández, Antonio; González-Rico, Francisco J; Román, Angel C; Rico-Leo, Eva; Alvarez-Barrientos, Alberto; Sánchez, Laura; Macia, Ángela; Heras, Sara R; García-Pérez, José L; Merino, Jaime M; Fernández-Salguero, Pedro M

    2016-06-01

    Cell differentiation is a central process in development and in cancer growth and dissemination. OCT4 (POU5F1) and NANOG are essential for cell stemness and pluripotency; yet, the mechanisms that regulate their expression remain largely unknown. Repetitive elements account for almost half of the Human Genome; still, their role in gene regulation is poorly understood. Here, we show that the dioxin receptor (AHR) leads to differentiation of human carcinoma cells through the transcriptional upregulation of Alu retrotransposons, whose RNA transcripts can repress pluripotency genes. Despite the genome-wide presence of Alu elements, we provide evidences that those located at the NANOG and OCT4 promoters bind AHR, are transcribed by RNA polymerase-III and repress NANOG and OCT4 in differentiated cells. OCT4 and NANOG repression likely involves processing of Alu-derived transcripts through the miRNA machinery involving the Microprocessor and RISC. Consistently, stable AHR knockdown led to basal undifferentiation, impaired Alus transcription and blockade of OCT4 and NANOG repression. We suggest that transcripts produced from AHR-regulated Alu retrotransposons may control the expression of stemness genes OCT4 and NANOG during differentiation of carcinoma cells. The control of discrete Alu elements by specific transcription factors may have a dynamic role in genome regulation under physiological and diseased conditions. PMID:26883630

  8. Regulation of Na+/K+ ATPase transport velocity by RNA editing.

    Directory of Open Access Journals (Sweden)

    Claudia Colina

    Full Text Available Because firing properties and metabolic rates vary widely, neurons require different transport rates from their Na(+/K(+ pumps in order to maintain ion homeostasis. In this study we show that Na(+/K(+ pump activity is tightly regulated by a novel process, RNA editing. Three codons within the squid Na(+/K(+ ATPase gene can be recoded at the RNA level, and the efficiency of conversion for each varies dramatically, and independently, between tissues. At one site, a highly conserved isoleucine in the seventh transmembrane span can be converted to a valine, a change that shifts the pump's intrinsic voltage dependence. Mechanistically, the removal of a single methyl group specifically targets the process of Na(+ release to the extracellular solution, causing a higher turnover rate at the resting membrane potential.

  9. Is plant mitochondrial RNA editing a source of phylogenetic incongruence? An answer from in silico and in vivo data sets

    Directory of Open Access Journals (Sweden)

    Quagliariello Carla

    2008-03-01

    Full Text Available Abstract Background In plant mitochondria, the post-transcriptional RNA editing process converts C to U at a number of specific sites of the mRNA sequence and usually restores phylogenetically conserved codons and the encoded amino acid residues. Sites undergoing RNA editing evolve at a higher rate than sites not modified by the process. As a result, editing sites strongly affect the evolution of plant mitochondrial genomes, representing an important source of sequence variability and potentially informative characters. To date no clear and convincing evidence has established whether or not editing sites really affect the topology of reconstructed phylogenetic trees. For this reason, we investigated here the effect of RNA editing on the tree building process of twenty different plant mitochondrial gene sequences and by means of computer simulations. Results Based on our simulation study we suggest that the editing ‘noise’ in tree topology inference is mainly manifested at the cDNA level. In particular, editing sites tend to confuse tree topologies when artificial genomic and cDNA sequences are generated shorter than 500 bp and with an editing percentage higher than 5.0%. Similar results have been also obtained with genuine plant mitochondrial genes. In this latter instance, indeed, the topology incongruence increases when the editing percentage goes up from about 3.0 to 14.0%. However, when the average gene length is higher than 1,000 bp (rps3, matR and atp1 no differences in the comparison between inferred genomic and cDNA topologies could be detected. Conclusions Our findings by the here reported in silico and in vivo computer simulation system seem to strongly suggest that editing sites contribute in the generation of misleading phylogenetic trees if the analyzed mitochondrial gene sequence is highly edited (higher than 3.0% and reduced in length (shorter than 500 bp. In the current lack of direct experimental evidence the results

  10. RED: A Java-MySQL Software for Identifying and Visualizing RNA Editing Sites Using Rule-Based and Statistical Filters.

    Directory of Open Access Journals (Sweden)

    Yongmei Sun

    Full Text Available RNA editing is one of the post- or co-transcriptional processes that can lead to amino acid substitutions in protein sequences, alternative pre-mRNA splicing, and changes in gene expression levels. Although several methods have been suggested to identify RNA editing sites, there remains challenges to be addressed in distinguishing true RNA editing sites from its counterparts on genome and technical artifacts. In addition, there lacks a software framework to identify and visualize potential RNA editing sites. Here, we presented a software - 'RED' (RNA Editing sites Detector - for the identification of RNA editing sites by integrating multiple rule-based and statistical filters. The potential RNA editing sites can be visualized at the genome and the site levels by graphical user interface (GUI. To improve performance, we used MySQL database management system (DBMS for high-throughput data storage and query. We demonstrated the validity and utility of RED by identifying the presence and absence of C→U RNA-editing sites experimentally validated, in comparison with REDItools, a command line tool to perform high-throughput investigation of RNA editing. In an analysis of a sample data-set with 28 experimentally validated C→U RNA editing sites, RED had sensitivity and specificity of 0.64 and 0.5. In comparison, REDItools had a better sensitivity (0.75 but similar specificity (0.5. RED is an easy-to-use, platform-independent Java-based software, and can be applied to RNA-seq data without or with DNA sequencing data. The package is freely available under the GPLv3 license at http://github.com/REDetector/RED or https://sourceforge.net/projects/redetector.

  11. RED: A Java-MySQL Software for Identifying and Visualizing RNA Editing Sites Using Rule-Based and Statistical Filters

    Science.gov (United States)

    Sun, Yongmei; Li, Xing; Wu, Di; Pan, Qi; Ji, Yuefeng; Ren, Hong; Ding, Keyue

    2016-01-01

    RNA editing is one of the post- or co-transcriptional processes that can lead to amino acid substitutions in protein sequences, alternative pre-mRNA splicing, and changes in gene expression levels. Although several methods have been suggested to identify RNA editing sites, there remains challenges to be addressed in distinguishing true RNA editing sites from its counterparts on genome and technical artifacts. In addition, there lacks a software framework to identify and visualize potential RNA editing sites. Here, we presented a software − ‘RED’ (RNA Editing sites Detector) − for the identification of RNA editing sites by integrating multiple rule-based and statistical filters. The potential RNA editing sites can be visualized at the genome and the site levels by graphical user interface (GUI). To improve performance, we used MySQL database management system (DBMS) for high-throughput data storage and query. We demonstrated the validity and utility of RED by identifying the presence and absence of C→U RNA-editing sites experimentally validated, in comparison with REDItools, a command line tool to perform high-throughput investigation of RNA editing. In an analysis of a sample data-set with 28 experimentally validated C→U RNA editing sites, RED had sensitivity and specificity of 0.64 and 0.5. In comparison, REDItools had a better sensitivity (0.75) but similar specificity (0.5). RED is an easy-to-use, platform-independent Java-based software, and can be applied to RNA-seq data without or with DNA sequencing data. The package is freely available under the GPLv3 license at http://github.com/REDetector/RED or https://sourceforge.net/projects/redetector. PMID:26930599

  12. The association of Alu repeats with the generation of potential AU-rich elements (ARE at 3' untranslated regions.

    Directory of Open Access Journals (Sweden)

    Bhak Jonghwa

    2004-12-01

    Full Text Available Abstract Background A significant portion (about 8% in the human genome of mammalian mRNA sequences contains AU (Adenine and Uracil rich elements or AREs at their 3' untranslated regions (UTR. These mRNA sequences are usually stable. However, an increasing number of observations have been made of unstable species, possibly depending on certain elements such as Alu repeats. ARE motifs are repeats of the tetramer AUUU and a monomer A at the end of the repeats ((AUUUnA. The importance of AREs in biology is that they make certain mRNA unstable. Proto-oncogene, such as c-fos, c-myc, and c-jun in humans, are associated with AREs. Although it has been known that the increased number of ARE motifs caused the decrease of the half-life of mRNA containing ARE repeats, the exact mechanism is as of yet unknown. We analyzed the occurrences of AREs and Alu and propose a possible mechanism for how human mRNA could acquire and keep AREs at its 3' UTR originating from Alu repeats. Results Interspersed in the human genome, Alu repeats occupy 5% of the 3' UTR of mRNA sequences. Alu has poly-adenine (poly-A regions at its end, which lead to poly-thymine (poly-T regions at the end of its complementary Alu. It has been found that AREs are present at the poly-T regions. From the 3' UTR of the NCBI's reference mRNA sequence database, we found nearly 40% (38.5% of ARE (Class I were associated with Alu sequences (Table 1 within one mismatch allowance in ARE sequences. Other ARE classes had statistically significant associations as well. This is far from a random occurrence given their limited quantity. At each ARE class, random distribution was simulated 1,000 times, and it was shown that there is a special relationship between ARE patterns and the Alu repeats. Table 1 Defined ARE classes. (Symbol marks are used in this study instead of full sequences. Symbol ARE sequence Class I (AUUU5A AUUUAUUUAUUUAUUUAUUUA Class II (AUUU4A AUUUAUUUAUUUAUUUA Class III U(AUUU3AU

  13. Adenosine-to-inosine RNA editing affects trafficking of the gamma-aminobutyric acid type A (GABA(A)) receptor.

    Science.gov (United States)

    Daniel, Chammiran; Wahlstedt, Helene; Ohlson, Johan; Björk, Petra; Ohman, Marie

    2011-01-21

    Recoding by adenosine-to-inosine RNA editing plays an important role in diversifying proteins involved in neurotransmission. We have previously shown that the Gabra-3 transcript, coding for the α3 subunit of the GABA(A) receptor is edited in mouse, causing an isoleucine to methionine (I/M) change. Here we show that this editing event is evolutionarily conserved from human to chicken. Analyzing recombinant GABA(A) receptor subunits expressed in HEK293 cells, our results suggest that editing at the I/M site in α3 has functional consequences on receptor expression. We demonstrate that I/M editing reduces the cell surface and the total number of α3 subunits. The reduction in cell surface levels is independent of the subunit combination as it is observed for α3 in combination with either the β2 or the β3 subunit. Further, an amino acid substitution at the corresponding I/M site in the α1 subunit has a similar effect on cell surface presentation, indicating the importance of this site for receptor trafficking. We show that the I/M editing during brain development is inversely related to the α3 protein abundance. Our results suggest that editing controls trafficking of α3-containing receptors and may therefore facilitate the switch of subunit compositions during development as well as the subcellular distribution of α subunits in the adult brain. PMID:21030585

  14. Adenosine-to-Inosine RNA Editing Affects Trafficking of the γ-Aminobutyric Acid Type A (GABAA) Receptor*

    Science.gov (United States)

    Daniel, Chammiran; Wahlstedt, Helene; Ohlson, Johan; Björk, Petra; Öhman, Marie

    2011-01-01

    Recoding by adenosine-to-inosine RNA editing plays an important role in diversifying proteins involved in neurotransmission. We have previously shown that the Gabra-3 transcript, coding for the α3 subunit of the GABAA receptor is edited in mouse, causing an isoleucine to methionine (I/M) change. Here we show that this editing event is evolutionarily conserved from human to chicken. Analyzing recombinant GABAA receptor subunits expressed in HEK293 cells, our results suggest that editing at the I/M site in α3 has functional consequences on receptor expression. We demonstrate that I/M editing reduces the cell surface and the total number of α3 subunits. The reduction in cell surface levels is independent of the subunit combination as it is observed for α3 in combination with either the β2 or the β3 subunit. Further, an amino acid substitution at the corresponding I/M site in the α1 subunit has a similar effect on cell surface presentation, indicating the importance of this site for receptor trafficking. We show that the I/M editing during brain development is inversely related to the α3 protein abundance. Our results suggest that editing controls trafficking of α3-containing receptors and may therefore facilitate the switch of subunit compositions during development as well as the subcellular distribution of α subunits in the adult brain. PMID:21030585

  15. Efficient and transgene-free genome editing in wheat through transient expression of CRISPR/Cas9 DNA or RNA

    Science.gov (United States)

    Zhang, Yi; Liang, Zhen; Zong, Yuan; Wang, Yanpeng; Liu, Jinxing; Chen, Kunling; Qiu, Jin-Long; Gao, Caixia

    2016-01-01

    Editing plant genomes is technically challenging in hard-to-transform plants and usually involves transgenic intermediates, which causes regulatory concerns. Here we report two simple and efficient genome-editing methods in which plants are regenerated from callus cells transiently expressing CRISPR/Cas9 introduced as DNA or RNA. This transient expression-based genome-editing system is highly efficient and specific for producing transgene-free and homozygous wheat mutants in the T0 generation. We demonstrate our protocol to edit genes in hexaploid bread wheat and tetraploid durum wheat, and show that we are able to generate mutants with no detectable transgenes. Our methods may be applicable to other plant species, thus offering the potential to accelerate basic and applied plant genome-engineering research. PMID:27558837

  16. High-efficiency targeted editing of large viral genomes by RNA-guided nucleases.

    Directory of Open Access Journals (Sweden)

    Yanwei Bi

    2014-05-01

    Full Text Available A facile and efficient method for the precise editing of large viral genomes is required for the selection of attenuated vaccine strains and the construction of gene therapy vectors. The type II prokaryotic CRISPR-Cas (clustered regularly interspaced short palindromic repeats (CRISPR-associated (Cas RNA-guided nuclease system can be introduced into host cells during viral replication. The CRISPR-Cas9 system robustly stimulates targeted double-stranded breaks in the genomes of DNA viruses, where the non-homologous end joining (NHEJ and homology-directed repair (HDR pathways can be exploited to introduce site-specific indels or insert heterologous genes with high frequency. Furthermore, CRISPR-Cas9 can specifically inhibit the replication of the original virus, thereby significantly increasing the abundance of the recombinant virus among progeny virus. As a result, purified recombinant virus can be obtained with only a single round of selection. In this study, we used recombinant adenovirus and type I herpes simplex virus as examples to demonstrate that the CRISPR-Cas9 system is a valuable tool for editing the genomes of large DNA viruses.

  17. A survey of PPR proteins identifies DYW domains like those of land plant RNA editing factors in diverse eukaryotes

    OpenAIRE

    Schallenberg-Rüdinger, Mareike; Lenz, Henning; Polsakiewicz, Monika; Gott, Jonatha M.; Knoop, Volker

    2013-01-01

    The pentatricopeptide repeat modules of PPR proteins are key to their sequence-specific binding to RNAs. Gene families encoding PPR proteins are greatly expanded in land plants where hundreds of them participate in RNA maturation, mainly in mitochondria and chloroplasts. Many plant PPR proteins contain additional carboxyterminal domains and have been identified as essential factors for specific events of C-to-U RNA editing, which is abundant in the two endosymbiotic plant organelles. Among th...

  18. Cytidine deaminase motifs within the DYW domain of two pentatricopeptide repeat-containing proteins are required for site-specific chloroplast RNA editing.

    Science.gov (United States)

    Wagoner, Jessica A; Sun, Tao; Lin, Lin; Hanson, Maureen R

    2015-01-30

    In angiosperm organelles, cytidines are converted to uridines by a deamination reaction in the process termed RNA editing. The C targets of editing are recognized by members of the pentatricopeptide repeat (PPR) protein family. Although other members of the editosome have begun to be identified, the enzyme that catalyzes the C-U conversion is still unknown. The DYW motif at the C terminus of many PPR editing factors contains residues conserved with known cytidine deaminase active sites; however, some PPR editing factors lack a DYW motif. Furthermore, in many PPR-DYW editing factors, the truncation of the DYW motif does not affect editing efficiency, so the role of the DYW motif in RNA editing is unclear. Here, a chloroplast PPR-DYW editing factor, quintuple editing factor 1 (QED1), was shown to affect five different plastid editing sites, the greatest number of chloroplast C targets known to be affected by a single PPR protein. Loss of editing at the five sites resulted in stunted growth and accumulation of apparent photodamage. Adding a C-terminal protein tag to QED1 was found to severely inhibit editing function. QED1 and RARE1, another plastid PPR-DYW editing factor, were discovered to require their DYW motifs for efficient editing. To identify specific residues critical for editing, conserved deaminase residues in each PPR protein were mutagenized. The mutant PPR proteins, when expressed in qed1 or rare1 mutant protoplasts, could not complement the editing defect. Therefore, the DYW motif, and specifically, the deaminase residues, of QED1 and RARE1 are required for editing efficiency. PMID:25512379

  19. A survey of PPR proteins identifies DYW domains like those of land plant RNA editing factors in diverse eukaryotes.

    Science.gov (United States)

    Schallenberg-Rüdinger, Mareike; Lenz, Henning; Polsakiewicz, Monika; Gott, Jonatha M; Knoop, Volker

    2013-01-01

    The pentatricopeptide repeat modules of PPR proteins are key to their sequence-specific binding to RNAs. Gene families encoding PPR proteins are greatly expanded in land plants where hundreds of them participate in RNA maturation, mainly in mitochondria and chloroplasts. Many plant PPR proteins contain additional carboxyterminal domains and have been identified as essential factors for specific events of C-to-U RNA editing, which is abundant in the two endosymbiotic plant organelles. Among those carboxyterminal domain additions to plant PPR proteins, the so-called DYW domain is particularly interesting given its similarity to cytidine deaminases. The frequency of organelle C-to-U RNA editing and the diversity of DYW-type PPR proteins correlate well in plants and both were recently identified outside of land plants, in the protist Naegleria gruberi. Here we present a systematic survey of PPR protein genes and report on the identification of additional DYW-type PPR proteins in the protists Acanthamoeba castellanii, Malawimonas jakobiformis, and Physarum polycephalum. Moreover, DYW domains were also found in basal branches of multi-cellular lineages outside of land plants, including the alga Nitella flexilis and the rotifers Adineta ricciae and Philodina roseola. Intriguingly, the well-characterized and curious patterns of mitochondrial RNA editing in the slime mold Physarum also include examples of C-to-U changes. Finally, we identify candidate sites for mitochondrial RNA editing in Malawimonas, further supporting a link between DYW-type PPR proteins and C-to-U editing, which may have remained hitherto unnoticed in additional eukaryote lineages. PMID:23899506

  20. The essential role of AMPA receptor GluR2 subunit RNA editing in the normal and diseased brain

    Directory of Open Access Journals (Sweden)

    Amanda Lorraine Wright

    2012-04-01

    Full Text Available AMPA receptors are comprised of different combinations of GluR1-GluR4 (also known as GluA1-GluA4 and GluR-A to GluR-D subunits. The GluR2 subunit is subject to Q/R site RNA editing by the ADAR2 enzyme, which converts a codon for glutamine (Q, present in the GluR2 gene, to a codon for arginine (R found in the mRNA. AMPA receptors are calcium (Ca2+-permeable if they contain the unedited GluR2(Q subunit or if they lack the GluR2 subunit. While most AMPA receptors in the brain contain the edited GluR2(R subunit and are therefore Ca2+-impermeable, recent evidence suggests that Ca2+-permeable GluR2-lacking AMPA receptors are important in synaptic plasticity and learning. However, the presence of Ca2+-permeable AMPA receptors containing unedited GluR2 leads to excitotoxic cell loss. Recent studies have indicated that RNA editing of GluR2 is deregulated in diseases, such as amyotrophic lateral sclerosis (ALS, as well in acute neurodegenerative conditions, such as ischemia. More recently, studies have investigated the regulation of RNA editing and possible causes for its deregulation during disease. In this review, we will explore the role of GluR2 RNA editing in the healthy and diseased brain and outline new insights into the mechanisms that control this process.

  1. Demonstration of mRNA editing and localization of guide RNA genes in kinetoplast-mitochondria of the plant trypanosomatid Phytomonas serpens.

    Science.gov (United States)

    Maslov, D A; Hollar, L; Haghighat, P; Nawathean, P

    1998-06-01

    Maxicircle molecules of kDNA in several isolates of Phytomonas were detected by hybridization with the 12S rRNA gene probe from Leishmania tarentolae. The estimated size of maxicircles is isolate-specific and varies from 27 to 36 kb. Fully edited and polyadenylated mRNA for kinetoplast-encoded ribosomal protein S12 (RPS12) was found in the steady-state kinetoplast RNA isolated from Phytomonas serpens strain 1G. Two minicircles (1.45 kb) from this strain were also sequenced. Each minicircle contains two 120 bp conserved regions positioned 180 degrees apart, a region enriched with G and T bases and a variable region. One minicircle encodes a gRNA for the first block of editing of RPSl2 mRNA, and the other encodes a gRNA with unknown function. A gRNA gene for the second block of RPSl2 was found on a minicircle sequenced previously. On each minicircle, a gRNA gene is located in the variable region in a similar position and orientation with respect to the conserved regions. PMID:9662707

  2. Region-specific alterations of A-to-I RNA editing of serotonin 2c receptor in the cortex of suicides with major depression.

    Science.gov (United States)

    Weissmann, D; van der Laan, S; Underwood, M D; Salvetat, N; Cavarec, L; Vincent, L; Molina, F; Mann, J J; Arango, V; Pujol, J F

    2016-01-01

    Brain region-specific abnormalities in serotonergic transmission appear to underlie suicidal behavior. Alterations of RNA editing on the serotonin receptor 2C (HTR2C) pre-mRNA in the brain of suicides produce transcripts that attenuate 5-HT2CR signaling by impairing intracellular G-protein coupling and subsequent intracellular signal transduction. In brain, the distribution of RNA-editing enzymes catalyzing deamination (A-to-I modification) shows regional variation, including within the cerebral cortex. We tested the hypothesis that altered pre-mRNA 5-HT2CR receptor editing in suicide is region-specific. To this end, we investigated the complete 5-HT2CR mRNA-editing profile in two architectonically distinct cortical areas involved in mood regulation and decision-making in a clinically well-characterized cohort of age- and sex-matched non-psychiatric drug-free controls and depressed suicides. By using an original biochemical detection method, that is, capillary electrophoresis single-stranded conformational polymorphism (CE-SSCP), we corroborated the 5-HT2CR mRNA-editing profile previously described in the dorsolateral prefrontal cortex (Brodmann area 9 (BA9)). Editing of 5-HT2CR mRNA displayed clear regional difference when comparing dorsolateral prefrontal cortex (BA9) and anterior cingulate cortex (BA24). Compared with non-psychiatric control individuals, alterations of editing levels of 5-HT2CR mRNA were detected in both cortical areas of depressed suicides. A marked increase in editing on 5-HT2CR was especially observed in the anterior cingulate cortex in suicides, implicating this cortical area in suicide risk. The results suggest that region-specific changes in RNA editing of 5-HT2CR mRNA and deficient receptor function likely contribute to the etiology of major depressive disorder or suicide. PMID:27576167

  3. Chemical modification of nucleotide bases and mRNA editing depend on hexamer or nucleoprotein phase in Sendai virus nucleocapsids.

    OpenAIRE

    Iseni, Frédéric; Baudin, Florence; Garcin, Dominique; Marq, Jean-Baptiste; Ruigrok, Rob W. H.; Kolakofsky, Daniel

    2002-01-01

    The minus-strand genome of Sendai virus is an assembly of the nucleocapsid protein (N) and RNA, in which each N subunit is associated with precisely 6 nt. Only genomes that are a multiple of 6 nt long replicate efficiently or are found naturally, and their replication promoters contain sequence elements with hexamer repeats. Paramyxoviruses that are governed by this hexamer rule also edit their P gene mRNA during its synthesis, by G insertions, via a controlled form of viral RNA polymerase "s...

  4. Striking differences in RNA editing requirements to express the rps4 gene in magnolia and sunflower mitochondria.

    Science.gov (United States)

    Regina, Teresa M R; Lopez, Loredana; Picardi, Ernesto; Quagliariello, Carla

    2002-03-01

    The ribosomal protein S4 gene (rps4) has been identified as a single copy sequence in the mitochondrial genomes of two distant higher plants, Magnolia and Helianthus. Sequence analysis revealed that the rps4 genes present in the magnolia and sunflower mitochondrial genomes encode S4 polypeptides of 352 and 331 amino acids, respectively, longer than their counterparts in liverwort and bacteria. Expression of the rps4 genes in the investigated higher plant mitochondria was confirmed by Western blot analysis. In Helianthus, one of two short nucleotide insertions at the 3'-end introduces in the coding region a premature termination codon. Northern hybridizations and reverse transcription-polymerase chain reaction analysis demonstrated that the monocistronic RNA transcripts generated from the rps4 locus in Magnolia and Helianthus mitochondria are modified by RNA editing at 28 and 13 positions, respectively. Although evolutionarily conserved, RNA editing requirements of the rps4 appear more extensive in Magnolia than in Helianthus and in the other higher plants so far investigated. Furthermore, our analysis also suggests that selection of editing sites is RNA sequence-specific in a duplicated sequence context. PMID:11943458

  5. Alterations of RNA Editing for the Mitochondrial ATP9 Gene in a New orf220-type Cytoplasmic Male-sterile Line of Stem Mustard (Brassica juncea var. tumida)

    Institute of Scientific and Technical Information of China (English)

    Jing-Hua Yang; Ming-Fang Zhang; Jing-Quan Yu

    2007-01-01

    RNA editing for the mitochondrial ATP9 gene of encoding regions has been observed in both cytoplasmic malesterile and maintainer lines of stem mustard, where its editing capacity varied spatially and temporally in the cytoplasmic male sterility (CMS) line. There were four RNA editing sites for the mitochondrial ATP9 gene according to its normal editing sites in mustard, of which three sites occurred as C-to-U changes and one as a U-to-C change.As a result, the hydrophobicity of deduced ATP9 protein was reduced due to the conversions at its 17th, 45th and 64th positions. Meanwhile, the conservation of deduced ATP9 protein was enhanced by changes at the 56th position.Loss of a specific editing site for ATP9 was observed in juvenile roots, senile roots, senile leaves and floret buds of the CMS line. Comparatively, complete RNA editing for ATP9 gene was retained in juvenile roots, juvenile leaves and floret buds of its maintainer line; however, the loss of a specific editing site for ATP9 gene occurred at senile roots and senile leaves in its maintainer line. These observations allow us to produce a hypothesis that the dysfunction of a specific mitochondrisl gene arising from RNA editing could probably be a factor triggering CMS and organ senescence through unknown cross-talk pathways during development.

  6. The assembly of F1FO-ATP synthase is disrupted upon interference of RNA editing in Trypanosoma brucei

    Czech Academy of Sciences Publication Activity Database

    Hashimi, Hassan; Benkovičová, V.; Čermáková, P.; Lai, De Hua; Horváth, A.; Lukeš, Julius

    2010-01-01

    Roč. 40, č. 1 (2010), s. 45-54. ISSN 0020-7519 R&D Projects: GA ČR GA204/06/1558; GA AV ČR IAA500960705 Institutional research plan: CEZ:AV0Z60220518 Keywords : RNA editing * ATP synthase * mitochondrion * Trypanosoma * respiratory complex * membrane potential Subject RIV: EB - Genetics ; Molecular Biology Impact factor: 3.822, year: 2010

  7. Crystallization and X-ray diffraction analysis of the Trp/amber editing site of hepatitis delta virus (+)RNA: a case of rational design

    Energy Technology Data Exchange (ETDEWEB)

    MacElrevey, Celeste; Wedekind, Joseph E., E-mail: joseph-wedekind@urmc.rochester.edu [Department of Biochemistry and Biophysics, University of Rochester School of Medicine and Dentistry, Rochester, New York 14642 (United States)

    2005-12-01

    Well diffracting decamer crystals of the hepatitis delta virus RNA-editing site were prepared, but exhibited merohedral twinning and base averaging owing to duplex symmetry. A longer asymmetric construct that includes additional flanking RNA sequences has been crystallized that does not appear to exhibit these defects. RNA editing by mammalian ADAR1 (Adenosine Deaminase Acting on RNA) is required for the life cycle of the hepatitis delta virus (HDV). Editing extends the single viral open reading frame to yield two protein products of alternate length. ADARs are believed to recognize double-stranded RNA substrates via a ‘structure-based’ readout mechanism. Crystals of 10-mer duplexes representing the HDV RNA-editing site diffracted to 1.35 Å resolution, but suffered from merohedral twinning and averaging of the base registry. Expansion of the construct to include two flanking 3 × 1 internal loops yielded crystals in the primitive tetragonal space group P4{sub 1}2{sub 1}2 or P4{sub 3}2{sub 1}2. X-ray diffraction data were collected to 2.8 Å resolution, revealing a unit cell with parameters a = 62.5, c = 63.5 Å. The crystallization and X-ray analysis of multiple forms of the HDV RNA-editing substrate, encounters with common RNA crystal-growth defects and a strategy to overcome these problems are reported.

  8. Crystallization and X-ray diffraction analysis of the Trp/amber editing site of hepatitis delta virus (+)RNA: a case of rational design

    International Nuclear Information System (INIS)

    Well diffracting decamer crystals of the hepatitis delta virus RNA-editing site were prepared, but exhibited merohedral twinning and base averaging owing to duplex symmetry. A longer asymmetric construct that includes additional flanking RNA sequences has been crystallized that does not appear to exhibit these defects. RNA editing by mammalian ADAR1 (Adenosine Deaminase Acting on RNA) is required for the life cycle of the hepatitis delta virus (HDV). Editing extends the single viral open reading frame to yield two protein products of alternate length. ADARs are believed to recognize double-stranded RNA substrates via a ‘structure-based’ readout mechanism. Crystals of 10-mer duplexes representing the HDV RNA-editing site diffracted to 1.35 Å resolution, but suffered from merohedral twinning and averaging of the base registry. Expansion of the construct to include two flanking 3 × 1 internal loops yielded crystals in the primitive tetragonal space group P41212 or P43212. X-ray diffraction data were collected to 2.8 Å resolution, revealing a unit cell with parameters a = 62.5, c = 63.5 Å. The crystallization and X-ray analysis of multiple forms of the HDV RNA-editing substrate, encounters with common RNA crystal-growth defects and a strategy to overcome these problems are reported

  9. Altered mRNA editing and expression of ionotropic glutamate receptors after kainic acid exposure in cyclooxygenase-2 deficient mice.

    Directory of Open Access Journals (Sweden)

    Luca Caracciolo

    Full Text Available Kainic acid (KA binds to the AMPA/KA receptors and induces seizures that result in inflammation, oxidative damage and neuronal death. We previously showed that cyclooxygenase-2 deficient (COX-2(-/- mice are more vulnerable to KA-induced excitotoxicity. Here, we investigated whether the increased susceptibility of COX-2(-/- mice to KA is associated with altered mRNA expression and editing of glutamate receptors. The expression of AMPA GluR2, GluR3 and KA GluR6 was increased in vehicle-injected COX-2(-/- mice compared to wild type (WT mice in hippocampus and cortex, whereas gene expression of NMDA receptors was decreased. KA treatment decreased the expression of AMPA, KA and NMDA receptors in the hippocampus, with a significant effect in COX-2(-/- mice. Furthermore, we analyzed RNA editing levels and found that the level of GluR3 R/G editing site was selectively increased in the hippocampus and decreased in the cortex in COX-2(-/- compared with WT mice. After KA, GluR4 R/G editing site, flip form, was increased in the hippocampus of COX-2(-/- mice. Treatment of WT mice with the COX-2 inhibitor celecoxib for two weeks decreased the expression of AMPA/KA and NMDAR subunits after KA, as observed in COX-2(-/- mice. After KA exposure, COX-2(-/- mice showed increased mRNA expression of markers of inflammation and oxidative stress, such as cytokines (TNF-α, IL-1β and IL-6, inducible nitric oxide synthase (iNOS, microglia (CD11b and astrocyte (GFAP. Thus, COX-2 gene deletion can exacerbate the inflammatory response to KA. We suggest that COX-2 plays a role in attenuating glutamate excitotoxicity by modulating RNA editing of AMPA/KA and mRNA expression of all ionotropic glutamate receptor subunits and, in turn, neuronal excitability. These changes may contribute to the increased vulnerability of COX-2(-/- mice to KA. The overstimulation of glutamate receptors as a consequence of COX-2 gene deletion suggests a functional coupling between COX-2 and the

  10. Studio dell'espressione dei retrotrasposoni umani Alu e LINE-1 e della sua possibile modulazione in risposta a fattori ambientali

    OpenAIRE

    Conti, Anastasia

    2015-01-01

    Gli elementi trasponibili costituiscono circa il 45% del genoma umano e solo la classe dei retrotrasposoni non-LTR risulta attiva oggigiorno dal punto di vista della retrotrasposizione. Fanno parte di questa classe, gli elementi LINE-1 ed Alu, i quali si inseriscono in un locus genomico diverso da quello di origine, duplicandosi attraverso la generazione di un intermedio ad RNA, aumentando così notevolmente il loro numero di copie genomiche (circa 1 milione per gli elementi Alu e 500 mila per...

  11. Regulation of oxytocin receptor gene expression in sheep: tissue specificity, multiple transcripts and mRNA editing.

    Science.gov (United States)

    Feng, H C; Bhave, M; Fairclough, R J

    2000-09-01

    The increase in uterine oxytocin receptor concentrations over the late luteal phase of the oestrous cycle in sheep is thought to play an important role in the regulation of the duration of the cycle by facilitating the effect of oxytocin on uterine prostaglandin release. Experiments indicated that oxytocin receptor mRNA expression in the endometrium was high at oestrus compared with at days 2, 7 and 12 of the oestrous cycle. The amount of oxytocin receptor mRNA expression in the pituitary gland did not show any significant differences during the oestrous cycle. Oxytocin receptor cDNA was obtained and characterized from ovine uterine endometrium on day 15 of the oestrous cycle, using RT-PCR techniques, to study the mechanisms underlying the resolution of oxytocin receptor expression. The cDNA sequence for the oxytocin receptor gene in sheep was found to be similar to that described previously, except for a difference of seven nucleotides. These nucleotide differences resulted in changes in four of the deduced amino acids in the oxytocin receptor sequence. The heterogeneity of the different sized oxytocin receptor transcripts in sheep is due, at least in part, to the alternative use of polyadenylation sites. Northern hybridization confirmed that the oxytocin receptor gene is expressed in ovine corpus luteum. The investigations on oxytocin receptor gene expression indicate that the patten of oxytocin receptor gene expression in sheep is not only tissue-specific, but also highly function-related. Evidence was obtained of mRNA editing in both the coding and the 3'-untranslated (3'UTR) regions of oxytocin receptor gene transcripts in ovine endometrium; this was the first demonstration of this phenomenon for oxytocin receptor mRNA. The present results indicate that the observed differences in oxytocin receptor mRNA sequences for the different oxytocin receptor populations in endometrium are due to mRNA editing. mRNA editing of oxytocin receptor transcripts may be

  12. Changing blue fluorescent protein to green fluorescent protein using chemical RNA editing as a novel strategy in genetic restoration.

    Science.gov (United States)

    Vu, Luyen T; Nguyen, Thanh T K; Alam, Shafiul; Sakamoto, Takashi; Fujimoto, Kenzo; Suzuki, Hitoshi; Tsukahara, Toshifumi

    2015-11-01

    Using the transition from cytosine of BFP (blue fluorescent protein) gene to uridine of GFP (green fluorescent protein) gene at position 199 as a model, we successfully controlled photochemical RNA editing to effect site-directed deamination of cytidine (C) to uridine (U). Oligodeoxynucleotides (ODNs) containing 5'-carboxyvinyl-2'-deoxyuridine ((CV) U) were used for reversible photoligation, and single-stranded 100-nt BFP DNA and in vitro-transcribed full-length BFP mRNA were the targets. Photo-cross-linking with the responsive ODNs was performed using UV (366 nm) irradiation, which was followed by heat treatment, and the cross-linked nucleotide was cleaved through photosplitting (UV, 312 nm). The products were analyzed using restriction fragment length polymorphism (RFLP) and fluorescence measurements. Western blotting and fluorescence-analysis results revealed that in vitro-translated proteins were synthesized from mRNAs after site-directed RNA editing. We detected substantial amounts of the target-base-substituted fragment using RFLP and observed highly reproducible spectra of the transition-GFP signal using fluorescence spectroscopy, which indicated protein stability. ODNc restored approximately 10% of the C-to-U transition. Thus, we successfully used non-enzymatic site-directed deamination for genetic restoration in vitro. In the near future, in vivo studies that include cultured cells and model animals will be conducted to treat genetic disorders. PMID:26031895

  13. A population study of the minicircles in Trypanosoma cruzi: predicting guide RNAs in the absence of empirical RNA editing

    Directory of Open Access Journals (Sweden)

    Westenberger Scott J

    2007-05-01

    Full Text Available Abstract Background The structurally complex network of minicircles and maxicircles comprising the mitochondrial DNA of kinetoplastids mirrors the complexity of the RNA editing process that is required for faithful expression of encrypted maxicircle genes. Although a few of the guide RNAs that direct this editing process have been discovered on maxicircles, guide RNAs are mostly found on the minicircles. The nuclear and maxicircle genomes have been sequenced and assembled for Trypanosoma cruzi, the causative agent of Chagas disease, however the complement of 1.4-kb minicircles, carrying four guide RNA genes per molecule in this parasite, has been less thoroughly characterised. Results Fifty-four CL Brener and 53 Esmeraldo strain minicircle sequence reads were extracted from T. cruzi whole genome shotgun sequencing data. With these sequences and all published T. cruzi minicircle sequences, 108 unique guide RNAs from all known T. cruzi minicircle sequences and two guide RNAs from the CL Brener maxicircle were predicted using a local alignment algorithm and mapped onto predicted or experimentally determined sequences of edited maxicircle open reading frames. For half of the sequences no statistically significant guide RNA could be assigned. Likely positions of these unidentified gRNAs in T. cruzi minicircle sequences are estimated using a simple Hidden Markov Model. With the local alignment predictions as a standard, the HMM had an ~85% chance of correctly identifying at least 20 nucleotides of guide RNA from a given minicircle sequence. Inter-minicircle recombination was documented. Variable regions contain species-specific areas of distinct nucleotide preference. Two maxicircle guide RNA genes were found. Conclusion The identification of new minicircle sequences and the further characterization of all published minicircles are presented, including the first observation of recombination between minicircles. Extrapolation suggests a level of 4

  14. The DYW Subgroup PPR Protein MEF35 Targets RNA Editing Sites in the Mitochondrial rpl16, nad4 and cob mRNAs in Arabidopsis thaliana.

    Directory of Open Access Journals (Sweden)

    Nadja Brehme

    Full Text Available RNA editing in plant mitochondria and plastids alters specific nucleotides from cytidine (C to uridine (U mostly in mRNAs. A number of PLS-class PPR proteins have been characterized as RNA recognition factors for specific RNA editing sites, all containing a C-terminal extension, the E domain, and some an additional DYW domain, named after the characteristic C-terminal amino acid triplet of this domain. Presently the recognition factors for more than 300 mitochondrial editing sites are still unidentified. In order to characterize these missing factors, the recently proposed computational prediction tool could be of use to assign target RNA editing sites to PPR proteins of yet unknown function. Using this target prediction approach we identified the nuclear gene MEF35 (Mitochondrial Editing Factor 35 to be required for RNA editing at three sites in mitochondria of Arabidopsis thaliana. The MEF35 protein contains eleven PPR repeats and E and DYW extensions at the C-terminus. Two T-DNA insertion mutants, one inserted just upstream and the other inside the reading frame encoding the DYW domain, show loss of editing at a site in each of the mRNAs for protein 16 in the large ribosomal subunit (site rpl16-209, for cytochrome b (cob-286 and for subunit 4 of complex I (nad4-1373, respectively. Editing is restored upon introduction of the wild type MEF35 gene in the reading frame mutant. The MEF35 protein interacts in Y2H assays with the mitochondrial MORF1 and MORF8 proteins, mutation of the latter also influences editing at two of the three MEF35 target sites. Homozygous mutant plants develop indistinguishably from wild type plants, although the RPL16 and COB/CYTB proteins are essential and the amino acids encoded after the editing events are conserved in most plant species. These results demonstrate the feasibility of the computational target prediction to screen for target RNA editing sites of E domain containing PLS-class PPR proteins.

  15. Alu repeats: A source for the genesis of primate microsatellites

    Energy Technology Data Exchange (ETDEWEB)

    Arcot, S.S.; Batzer, M.A. [Lawrence Livermore National Lab., CA (United States); Wang, Zhenyuan [Marshfield Medical Research Foundation, WI (United States)] [and others

    1995-09-01

    As a result of their abundance, relatively uniform distribution, and high degree of polymorphism, microsatellites and minisatellites have become valuable tools in genetic mapping, forensic identity testing, and population studies. In recent years, a number of microsatellite repeats have been found to be associated with Alu interspersed repeated DNA elements. The association of an Alu element with a microsatellite repeat could result from the integration of an Alu element within a preexisting microsatellite repeat. Alternatively, Alu elements could have a direct role in the origin of microsatellite repeats. Errors introduced during reverse transcription of the primary transcript derived from an Alu {open_quotes}master{close_quote} gene or the accumulation of random mutations in the middle A-rich regions and oligo(dA)-rich tails of Alu elements after insertion and subsequent expansion and contraction of these sequences could result in the genesis of a microsatellite repeat. We have tested these hypotheses by a direct evolutionary comparison of the sequences of some recent Alu elements that are found only in humans and are absent from nonhuman primates, as well as some older Alu elements that are present at orthologous positions in a number of nonhuman primates. The origin of {open_quotes}young{close_quotes} Alu insertions, absence of sequences that resemble microsatellite repeats at the orthologous loci in chimpanzees, and the gradual expansion of microsatellite repeats in some old Alu repeats at orthologous positions within the genomes of a number of nonhuman primates suggest that Alu elements are a source for the genesis of primate microsatellite repeats. 48 refs., 5 figs., 3 tabs.

  16. Research Progress on Relationship of PPR Proteins with RNA Editing%PPR蛋白参与RNA编辑机制的研究进展

    Institute of Scientific and Technical Information of China (English)

    何鹏; 陈海燕; 俞嘉宁

    2013-01-01

    RNA editing is a post-transcription phenomenon of gene processing and modification at specific nucleotide sites,which including nucleotide insertion/deletion or nucleotide conversion.RNA editing mainly occurs at genomic of mitochondria and chloroplast in higher plants.It has many important biology functions,but the mechanism of this phenomenon is still unclear.During recently years,the PPR proteins,might be as trans-acting factor of RNA editing,has been put forward and become a research focus in molecular biology.In this review,we introduced PPR proteins,RNA editing and possibility interaction pattern between PPR proteins and RNA editing.%RNA编辑是一种发生在转录后核苷酸特异位点的加工修饰现象,包括核苷酸的插入、删除和改变.高等植物中RNA编辑主要发生在线粒体与叶绿体中,具有重要的生物学功能,其机制仍在探索中.而PPR蛋白作为RNA编辑的反式作用因子,成为近几年来分子生物学的研究热点.该文就PPR蛋白、RNA编辑及PPR蛋白参与RNA编辑的机制等进行了综述.

  17. Elucidation of the RNA Recognition Code for Pentatricopeptide Repeat Proteins Involved in Organelle RNA Editing in Plants

    OpenAIRE

    Yagi, Yusuke; Hayashi, Shimpei; Kobayashi, Keiko; Hirayama, Takashi; Nakamura, Takahiro

    2013-01-01

    Pentatricopeptide repeat (PPR) proteins are eukaryotic RNA-binding proteins that are commonly found in plants. Organelle transcript processing and stability are mediated by PPR proteins in a gene-specific manner through recognition by tandem arrays of degenerate 35-amino-acid repeating units, the PPR motifs. However, the sequence-specific RNA recognition mechanism of the PPR protein remains largely unknown. Here, we show the principle underlying RNA recognition for PPR proteins involved in RN...

  18. Multifunctional G-rich and RRM-containing domains of TbRGG2 perform separate yet essential functions in trypanosome RNA editing.

    Science.gov (United States)

    Foda, Bardees M; Downey, Kurtis M; Fisk, John C; Read, Laurie K

    2012-09-01

    Efficient editing of Trypanosoma brucei mitochondrial RNAs involves the actions of multiple accessory factors. T. brucei RGG2 (TbRGG2) is an essential protein crucial for initiation and 3'-to-5' progression of editing. TbRGG2 comprises an N-terminal G-rich region containing GWG and RG repeats and a C-terminal RNA recognition motif (RRM)-containing domain. Here, we perform in vitro and in vivo separation-of-function studies to interrogate the mechanism of TbRGG2 action in RNA editing. TbRGG2 preferentially binds preedited mRNA in vitro with high affinity attributable to its G-rich region. RNA-annealing and -melting activities are separable, carried out primarily by the G-rich and RRM domains, respectively. In vivo, the G-rich domain partially complements TbRGG2 knockdown, but the RRM domain is also required. Notably, TbRGG2's RNA-melting activity is dispensable for RNA editing in vivo. Interactions between TbRGG2 and MRB1 complex proteins are mediated by both G-rich and RRM-containing domains, depending on the binding partner. Overall, our results are consistent with a model in which the high-affinity RNA binding and RNA-annealing activities of the G-rich domain are essential for RNA editing in vivo. The RRM domain may have key functions involving interactions with the MRB1 complex and/or regulation of the activities of the G-rich domain. PMID:22798390

  19. Effective Alu repeat based RT-Qpcr normalization in cancer cell perturbation experiments.

    Directory of Open Access Journals (Sweden)

    Ali Rihani

    Full Text Available BACKGROUND: Measuring messenger RNA (mRNA levels using the reverse transcription quantitative polymerase chain reaction (RT-qPCR is common practice in many laboratories. A specific set of mRNAs as internal control reference genes is considered as the preferred strategy to normalize RT-qPCR data. Proper selection of reference genes is a critical issue, especially in cancer cells that are subjected to different in vitro manipulations. These manipulations may result in dramatic alterations in gene expression levels, even of assumed reference genes. In this study, we evaluated the expression levels of 11 commonly used reference genes as internal controls for normalization of 19 experiments that include neuroblastoma, T-ALL, melanoma, breast cancer, non small cell lung cancer (NSCL, acute myeloid leukemia (AML, prostate cancer, colorectal cancer, and cervical cancer cell lines subjected to various perturbations. RESULTS: The geNorm algorithm in the software package qbase+ was used to rank the candidate reference genes according to their expression stability. We observed that the stability of most of the candidate reference genes varies greatly in perturbation experiments. Expressed Alu repeats show relatively stable expression regardless of experimental condition. These Alu repeats are ranked among the best reference assays in all perturbation experiments and display acceptable average expression stability values (M<0.5. CONCLUSIONS: We propose the use of Alu repeats as a reference assay when performing cancer cell perturbation experiments.

  20. Alu repeats as markers for forensic DNA analyses

    Energy Technology Data Exchange (ETDEWEB)

    Batzer, M.A.; Alegria-Hartman, M. [Lawrence Livermore National Lab., CA (United States); Kass, D.H. [Louisiana State Univ., New Orleans, LA (United States)] [and others

    1994-01-01

    The Human-Specific (HS) subfamily of Alu sequences is comprised of a group of 500 nearly identical members which are almost exclusively restricted to the human genome. Individual subfamily members share an average of 98.9% nucleotide identity with the HS subfamily consensus sequence, and have an average age of 2.8 million years. We have developed a Polymerase Chain Reaction (PCR) based assay using primers complementary to the 5 inch and 3 inch unique flanking DNA sequences from each HS Alu that allow the locus to be assayed for the presence or absence of the Alu repeat. The dimorphic HS Alu sequences probably inserted in the human genome after the radiation of modem humans (within the last 200,000-one million years) and represent a unique source of information for human population genetics and forensic DNA analyses. These sites can be developed into Dimorphic Alu Sequence Tagged Sites (DASTS) for the Human Genome Project. HS Alu family member insertions differ from other types of polymorphism (e.g. Variable Number of Tandem Repeat [VNTR] or Restriction Fragment Length Polymorphism [RFLP]) in that polymorphisms due to Alu insertions arise as a result of a unique event which has occurred only one time in the human population and spread through the population from that point. Therefore, individuals that share HS Alu repeats inherited these elements from a common ancestor. Most VNTR and RFLP polymorphisms may arise multiple times in parallel within a population.

  1. Diverse splicing patterns of exonized Alu elements in human tissues.

    Directory of Open Access Journals (Sweden)

    Lan Lin

    Full Text Available Exonization of Alu elements is a major mechanism for birth of new exons in primate genomes. Prior analyses of expressed sequence tags show that almost all Alu-derived exons are alternatively spliced, and the vast majority of these exons have low transcript inclusion levels. In this work, we provide genomic and experimental evidence for diverse splicing patterns of exonized Alu elements in human tissues. Using Exon array data of 330 Alu-derived exons in 11 human tissues and detailed RT-PCR analyses of 38 exons, we show that some Alu-derived exons are constitutively spliced in a broad range of human tissues, and some display strong tissue-specific switch in their transcript inclusion levels. Most of such exons are derived from ancient Alu elements in the genome. In SEPN1, mutations of which are linked to a form of congenital muscular dystrophy, the muscle-specific inclusion of an Alu-derived exon may be important for regulating SEPN1 activity in muscle. Realtime qPCR analysis of this SEPN1 exon in macaque and chimpanzee tissues indicates human-specific increase in its transcript inclusion level and muscle specificity after the divergence of humans and chimpanzees. Our results imply that some Alu exonization events may have acquired adaptive benefits during the evolution of primate transcriptomes.

  2. The mRNA expression of apolipoprotein B mRNA-editing enzyme-catalytic polypeptide-like 3G in peripheral blood mononuclear cells in patients with chronic hepatitis C and its regulation by interferon-α

    Institute of Scientific and Technical Information of China (English)

    蔡卫平

    2013-01-01

    Objective To study the mRNA expression of apolipoprotein B mRNA-editing enzyme-catalytic polypeptide-like 3G(APOBEC3G) in the peripheral blood mononuclear cells(PBMC) in patients with chronic hepatitis C(CHC) and its regulation by exogenous interferon-α

  3. The conserved protein kinase-A target motif in synapsin of Drosophila is effectively modified by pre-mRNA editing

    Directory of Open Access Journals (Sweden)

    Buchner Erich

    2006-11-01

    Full Text Available Abstract Background Synapsins are abundant synaptic vesicle associated phosphoproteins that are involved in the fine regulation of neurotransmitter release. The Drosophila member of this protein family contains three conserved domains (A, C, and E and is expressed in most or all synaptic terminals. Similar to mouse mutants, synapsin knock-out flies show no obvious structural defects but are disturbed in complex behaviour, notably learning and memory. Results We demonstrate that the N-terminal phosphorylation consensus motif RRxS that is conserved in all synapsins investigated so far, is modified in Drosophila by pre-mRNA editing. In mammals this motif represents the target site P1 of protein kinase A (PKA and calcium/calmodulin dependent protein kinase I/IV. The result of this editing, by which RRFS is modified to RGFS, can be observed in cDNAs of larvae and adults and in both isolated heads and bodies. It is also seen in several newly collected wild-type strains and thus does not represent an adaptation to laboratory culture conditions. A likely editing site complementary sequence is found in a downstream intron indicating that the synapsin pre-mRNA can form a double-stranded RNA structure that is required for editing by the adenosine deaminase acting on RNA (ADAR enzyme. A deletion in the Drosophila Adar gene generated by transposon remobilization prevents this modification, proving that the ADAR enzyme is responsible for the pre-mRNA editing described here. We also provide evidence for a likely function of synapsin editing in Drosophila. The N-terminal synapsin undeca-peptide containing the genomic motif (RRFS represents an excellent substrate for in-vitro phosphorylation by bovine PKA while the edited peptide (RGFS is not significantly phosphorylated. Thus pre-mRNA editing by ADAR could modulate the function of ubiquitously expressed synapsin in a cell-specific manner during development and adulthood. Conclusion Similar to several other

  4. Alu repeats as markers for human population genetics

    Energy Technology Data Exchange (ETDEWEB)

    Batzer, M.A.; Alegria-Hartman, M. [Lawrence Livermore National Lab., CA (United States); Bazan, H. [Louisiana State Univ., New Orleans, LA (United States). Medical Center] [and others

    1993-09-01

    The Human-Specific (HS) subfamily of Alu sequences is comprised of a group of 500 nearly identical members which are almost exclusively restricted to the human genome. Individual subfamily members share an average of 97.9% nucleotide identity with each other and an average of 98.9% nucleotide identity with the HS subfamily consensus sequence. HS Alu family members are thought to be derived from a single source ``master`` gene, and have an average age of 2.8 million years. We have developed a Polymerase Chain Reaction (PCR) based assay using primers complementary to the 5 in. and 3 in. unique flanking DNA sequences from each HS Alu that allows the locus to be assayed for the presence or absence of an Alu repeat. Individual HS Alu sequences were found to be either monomorphic or dimorphic for the presence or absence of each repeat. The monomorphic HS Alu family members inserted in the human genome after the human/great ape divergence (which is thought to have occurred 4--6 million years ago), but before the radiation of modem man. The dimorphic HS Alu sequences inserted in the human genome after the radiation of modem man (within the last 200,000-one million years) and represent a unique source of information for human population genetics and forensic DNA analyses. These sites can be developed into Dimorphic Alu Sequence Tagged Sites (DASTS) for the Human Genome Project as well. HS Alu family member insertion dimorphism differs from other types of polymorphism (e.g. Variable Number of Tandem Repeat [VNTR] or Restriction Fragment Length Polymorphism [RFLP]) because individuals share HS Alu family member insertions based upon identity by descent from a common ancestor as a result of a single event which occurred one time within the human population. The VNTR and RFLP polymorphisms may arise multiple times within a population and are identical by state only.

  5. Alu-Alu Recombination Underlying the First Large Genomic Deletion in GlcNAc-Phosphotransferase Alpha/Beta (GNPTAB) Gene in a MLII Alpha/Beta Patient

    DEFF Research Database (Denmark)

    Coutinho, F; da Silva Santos, L; Lacerda, L;

    2012-01-01

    -Alu unequal homologous recombination. RT-PCR methods were used to further evaluate the consequences of the alteration for the processing of the mutant pre mRNA GNPTAB, revealing the production of three abnormal transcripts: one without exon 19 (p.Lys1146_Trp1201del); another with an additional loss of exon 20...... been reported. Here we present the first case of a large homozygous intragenic GNPTAB gene deletion (c.3435-386_3602 + 343del897) encompassing exon 19, identified in a ML II α/β patient. Long-range PCR and sequencing methodologies were used to refine the characterization of this rearrangement, leading...

  6. Functional and structural insights revealed by molecular dynamics simulations of an essential RNA editing ligase in Trypanosoma brucei.

    Directory of Open Access Journals (Sweden)

    Rommie E Amaro

    Full Text Available RNA editing ligase 1 (TbREL1 is required for the survival of both the insect and bloodstream forms of Trypanosoma brucei, the parasite responsible for the devastating tropical disease African sleeping sickness. The type of RNA editing that TbREL1 is involved in is unique to the trypanosomes, and no close human homolog is known to exist. In addition, the high-resolution crystal structure revealed several unique features of the active site, making this enzyme a promising target for structure-based drug design. In this work, two 20 ns atomistic molecular dynamics (MD simulations are employed to investigate the dynamics of TbREL1, both with and without the ATP substrate present. The flexibility of the active site, dynamics of conserved residues and crystallized water molecules, and the interactions between TbREL1 and the ATP substrate are investigated and discussed in the context of TbREL1's function. Differences in local and global motion upon ATP binding suggest that two peripheral loops, unique to the trypanosomes, may be involved in interdomain signaling events. Notably, a significant structural rearrangement of the enzyme's active site occurs during the apo simulations, opening an additional cavity adjacent to the ATP binding site that could be exploited in the development of effective inhibitors directed against this protozoan parasite. Finally, ensemble averaged electrostatics calculations over the MD simulations reveal a novel putative RNA binding site, a discovery that has previously eluded scientists. Ultimately, we use the insights gained through the MD simulations to make several predictions and recommendations, which we anticipate will help direct future experimental studies and structure-based drug discovery efforts against this vital enzyme.

  7. APOBEC3A is implicated in a novel class of G-to-A mRNA editing in WT1 transcripts.

    Science.gov (United States)

    Niavarani, Ahmadreza; Currie, Erin; Reyal, Yasmin; Anjos-Afonso, Fernando; Horswell, Stuart; Griessinger, Emmanuel; Luis Sardina, Jose; Bonnet, Dominique

    2015-01-01

    Classic deamination mRNA changes, including cytidine to uridine (C-to-U) and adenosine to inosine (A-to-I), are important exceptions to the central dogma and lead to significant alterations in gene transcripts and products. Although there are a few reports of non-classic mRNA alterations, as yet there is no molecular explanation for these alternative changes. Wilms Tumor 1 (WT1) mutations and variants are implicated in several diseases, including Wilms tumor and acute myeloid leukemia (AML). We observed two alternative G-to-A changes, namely c.1303G>A and c.1586G>A in cDNA clones and found them to be recurrent in a series of 21 umbilical cord blood mononuclear cell (CBMC) samples studied. Two less conserved U-to-C changes were also observed. These alternative changes were found to be significantly higher in non-progenitor as compared to progenitor CBMCs, while they were found to be absent in a series of AML samples studied, indicating they are targeted, cell type-specific mRNA editing modifications. Since APOBEC/ADAR family members are implicated in RNA/DNA editing, we screened them by RNA-interference (RNAi) for WT1-mRNA changes and observed near complete reversal of WT1 c.1303G>A alteration upon APOBEC3A (A3A) knockdown. The role of A3A in mediating this change was confirmed by A3A overexpression in Fujioka cells, which led to a significant increase in WT1 c.1303G>A mRNA editing. Non-progenitor CBMCs showed correspondingly higher levels of A3A-mRNA and protein as compared to the progenitor ones. To our knowledge, this is the first report of mRNA modifying activity for an APOBEC3 protein and implicates A3A in a novel G-to-A form of editing. These findings open the way to further investigations into the mechanisms of other potential mRNA changes, which will help to redefine the RNA editing paradigm in both health and disease. PMID:25807502

  8. The Development of a Viral Mediated CRISPR/Cas9 System with Doxycycline Dependent gRNA Expression for Inducible In vitro and In vivo Genome Editing.

    Science.gov (United States)

    de Solis, Christopher A; Ho, Anthony; Holehonnur, Roopashri; Ploski, Jonathan E

    2016-01-01

    The RNA-guided Cas9 nuclease, from the type II prokaryotic Clustered Regularly Interspersed Short Palindromic Repeats (CRISPR) adaptive immune system, has been adapted and utilized by scientists to edit the genomes of eukaryotic cells. Here, we report the development of a viral mediated CRISPR/Cas9 system that can be rendered inducible utilizing doxycycline (Dox) and can be delivered to cells in vitro and in vivo utilizing adeno-associated virus (AAV). Specifically, we developed an inducible gRNA (gRNAi) AAV vector that is designed to express the gRNA from a H1/TO promoter. This AAV vector is also designed to express the Tet repressor (TetR) to regulate the expression of the gRNAi in a Dox dependent manner. We show that H1/TO promoters of varying length and a U6/TO promoter can edit DNA with similar efficiency in vitro, in a Dox dependent manner. We also demonstrate that our inducible gRNAi vector can be used to edit the genomes of neurons in vivo within the mouse brain in a Dox dependent manner. Genome editing can be induced in vivo with this system by supplying animals Dox containing food for as little as 1 day. This system might be cross compatible with many existing S. pyogenes Cas9 systems (i.e., Cas9 mouse, CRISPRi, etc.), and therefore it likely can be used to render these systems inducible as well. PMID:27587996

  9. Gene Loss and Error-Prone RNA Editing in the Mitochondrion of Perkinsela, an Endosymbiotic Kinetoplastid

    Czech Academy of Sciences Publication Activity Database

    David, Vojtěch; Flegontov, P.; Gerasimov, E.; Tanifuji, G.; Hashimi, Hassan; Logacheva, M.D.; Maruyama, S.; Onodera, N. T.; Gray, M.W.; Archibald, J.M.; Lukeš, Julius

    2015-01-01

    Roč. 6, č. 6 (2015), e01498-15. ISSN 2150-7511 R&D Projects: GA MŠk LH12104; GA ČR GAP305/12/2261 EU Projects: European Commission(XE) 316304; European Commission(XE) 289007 Institutional support: RVO:60077344 Keywords : kinetoplastid * protist * RNA-seq Subject RIV: EB - Genetics ; Molecular Biology Impact factor: 6.786, year: 2014

  10. Dimeric CRISPR RNA-Guided FokI-dCas9 Nucleases Directed by Truncated gRNAs for Highly Specific Genome Editing.

    Science.gov (United States)

    Wyvekens, Nicolas; Topkar, Ved V; Khayter, Cyd; Joung, J Keith; Tsai, Shengdar Q

    2015-07-01

    Monomeric clustered regularly interspaced short palindromic repeats (CRISPR)-CRISPR associated 9 (Cas9) nucleases have been widely adopted for simple and robust targeted genome editing but also have the potential to induce high-frequency off-target mutations. In principle, two orthogonal strategies for reducing off-target cleavage, truncated guide RNAs (tru-gRNAs) and dimerization-dependent RNA-guided FokI-dCas9 nucleases (RFNs), could be combined as tru-RFNs to further improve genome editing specificity. Here we identify a robust tru-RFN architecture that shows high activity in human cancer cell lines and embryonic stem cells. Additionally, we demonstrate that tru-gRNAs reduce the undesirable mutagenic effects of monomeric FokI-dCas9. Tru-RFNs combine the advantages of two orthogonal strategies for improving the specificity of CRISPR-Cas nucleases and therefore provide a highly specific platform for performing genome editing. PMID:26068112

  11. Activity-regulated RNA editing in select neuronal subfields in hippocampus

    Czech Academy of Sciences Publication Activity Database

    Balík, Aleš; Penn, A.C.; Nemoda, Z.; Greger, I. H.

    2013-01-01

    Roč. 41, č. 2 (2013), s. 1124-1134. ISSN 0305-1048 R&D Projects: GA ČR(CZ) GBP304/12/G069 Grant ostatní: Rada Programu interní podpory projektů mezinárodní spolupráce AV ČR(CZ) M200110971; Medical Research Council(GB) U105174197 Institutional support: RVO:67985823 Keywords : hippocampus * RNA * adenosine Subject RIV: ED - Physiology Impact factor: 8.808, year: 2013

  12. Antiviral Chemistry & Chemotherapy's current antiviral agents FactFile 2008 (2nd edition): RNA viruses.

    Science.gov (United States)

    De Clercq, Erik; Field, Hugh J

    2008-01-01

    Among the RNA viruses, other than the retroviruses (that is, HIV), which are dealt with separately in the current FactFile, the most important targets for the development of antiviral agents at the moment are the orthomyxoviruses (that is, influenza), the hepaciviruses (that is, hepatitis C virus [HCV]) and, to a lesser extent, the picornaviruses. Although the uncoating inhibitors amantadine and rimantadine were the first known inhibitors of influenza A, the neuraminidase inhibitors oseltamivir, zanamivir and peramivir have now become the prime antiviral drugs for the treatment of influenza A and B virus infections. For HCV infections, standard treatment consists of the combination of pegylated interferon-alpha with ribavirin, but several other antivirals targeted at specific viral functions such as the HCV protease and/ or polymerase may be expected to soon take an important share of this important market. Still untapped is the potential of a variety of uncoating inhibitors, as well as protease and/or polymerase inhibitors against the wide spectrum of picornaviruses. While ribavirin has been available for 35 years as a broad-spectrum anti-RNA virus agent, relatively new and unexplored is favipiravir (T-705) accredited with activity against influenza as well as flaviviruses, bunyaviruses and arenaviruses. PMID:18727441

  13. AEF1/MPR25 is implicated in RNA editing of plastid atpF and mitochondrial nad5, and also promotes atpF splicing in Arabidopsis and rice.

    Science.gov (United States)

    Yap, Aaron; Kindgren, Peter; Colas des Francs-Small, Catherine; Kazama, Tomohiko; Tanz, Sandra K; Toriyama, Kinya; Small, Ian

    2015-03-01

    RNA editing is an essential mechanism that modifies target cytidines to uridine in both mitochondrial and plastid mRNA. Target sites are recognized by pentatricopeptide repeat (PPR) proteins. Using bioinformatics predictions based on the code describing sequence recognition by PPR proteins, we have identified an Arabidopsis editing factor required for editing of atpF in plastids. A loss-of-function mutation in ATPF EDITING FACTOR 1 (AEF1, AT3G22150) results in severe variegation, presumably due to decreased plastid ATP synthase levels. Loss of editing at the atpF site is coupled with a large decrease in splicing of the atpF transcript, even though the editing site is within an exon and 53 nucleotides distant from the splice site. The rice orthologue of AEF1, MPR25, has been reported to be required for editing of a site in mitochondrial nad5 transcripts, and we confirm that editing of the same site is affected in the Arabidopsis aef1 mutant. We also show that splicing of chloroplast atpF transcripts is affected in the rice mpr25 mutant. AEF1 is thus highly unusual for an RNA editing specificity factor in that it has functions in both organelles. PMID:25585673

  14. A knowledgebase of the human Alu repetitive elements.

    Science.gov (United States)

    Mallona, Izaskun; Jordà, Mireia; Peinado, Miguel A

    2016-04-01

    Alu elements are the most abundant retrotransposons in the human genome with more than one million copies. Alu repeats have been reported to participate in multiple processes related with genome regulation and compartmentalization. Moreover, they have been involved in the facilitation of pathological mutations in many diseases, including cancer. The contribution of Alus and other repeats in genomic regulation is often overlooked because their study poses technical and analytical challenges hardly attainable with conventional strategies. Here we propose the integration of ontology-based semantic methods to query a knowledgebase for the human Alus. The knowledgebase for the human Alus leverages Sequence (SO) and Gene Ontologies (GO) and is devoted to address functional and genetic information in the genomic context of the Alus. For each Alu element, the closest gene and transcript are stored, as well their functional annotation according to GO, the state of the chromatin and the transcription factors binding sites inside the Alu. The model uses Web Ontology Language (OWL) and Semantic Web Rule Language (SWRL). As a case of use and to illustrate the utility of the tool, we have evaluated the epigenetic states of Alu repeats associated with gene promoters according to their transcriptional activity. The ontology is easily extendable, offering a scaffold for the inclusion of new experimental data. The RDF/XML formalization is freely available at http://aluontology.sourceforge.net/. PMID:26827622

  15. Analysis of Intron Sequences Reveals Hallmarks of Circular RNA Biogenesis in Animals

    Directory of Open Access Journals (Sweden)

    Andranik Ivanov

    2015-01-01

    Full Text Available Circular RNAs (circRNAs are a large class of animal RNAs. To investigate possible circRNA functions, it is important to understand circRNA biogenesis. Besides human ALU repeats, sequence features that promote exon circularization are largely unknown. We experimentally identified circRNAs in C. elegans. Reverse complementary sequences between introns bracketing circRNAs were significantly enriched in comparison to linear controls. By scoring the presence of reverse complementary sequences in human introns, we predicted and experimentally validated circRNAs. We show that introns bracketing circRNAs are highly enriched in RNA editing or hyperediting events. Knockdown of the double-strand RNA-editing enzyme ADAR1 significantly and specifically upregulated circRNA expression. Together, our data support a model of animal circRNA biogenesis in which competing RNA-RNA interactions of introns form larger structures that promote circularization of embedded exons, whereas ADAR1 antagonizes circRNA expression by melting stems within these interactions.

  16. Transcriptional Slippage and RNA Editing Increase the Diversity of Transcripts in Chloroplasts: Insight from Deep Sequencing of Vigna radiata Genome and Transcriptome.

    Directory of Open Access Journals (Sweden)

    Ching-Ping Lin

    Full Text Available We performed deep sequencing of the nuclear and organellar genomes of three mungbean genotypes: Vigna radiata ssp. sublobata TC1966, V. radiata var. radiata NM92 and the recombinant inbred line RIL59 derived from a cross between TC1966 and NM92. Moreover, we performed deep sequencing of the RIL59 transcriptome to investigate transcript variability. The mungbean chloroplast genome has a quadripartite structure including a pair of inverted repeats separated by two single copy regions. A total of 213 simple sequence repeats were identified in the chloroplast genomes of NM92 and RIL59; 78 single nucleotide variants and nine indels were discovered in comparing the chloroplast genomes of TC1966 and NM92. Analysis of the mungbean chloroplast transcriptome revealed mRNAs that were affected by transcriptional slippage and RNA editing. Transcriptional slippage frequency was positively correlated with the length of simple sequence repeats of the mungbean chloroplast genome (R2=0.9911. In total, 41 C-to-U editing sites were found in 23 chloroplast genes and in one intergenic spacer. No editing site that swapped U to C was found. A combination of bioinformatics and experimental methods revealed that the plastid-encoded RNA polymerase-transcribed genes psbF and ndhA are affected by transcriptional slippage in mungbean and in main lineages of land plants, including three dicots (Glycine max, Brassica rapa, and Nicotiana tabacum, two monocots (Oryza sativa and Zea mays, two gymnosperms (Pinus taeda and Ginkgo biloba and one moss (Physcomitrella patens. Transcript analysis of the rps2 gene showed that transcriptional slippage could affect transcripts at single sequence repeat regions with poly-A runs. It showed that transcriptional slippage together with incomplete RNA editing may cause sequence diversity of transcripts in chloroplasts of land plants.

  17. Highly efficient homology-driven genome editing in human T cells by combining zinc-finger nuclease mRNA and AAV6 donor delivery.

    Science.gov (United States)

    Wang, Jianbin; DeClercq, Joshua J; Hayward, Samuel B; Li, Patrick Wai-Lun; Shivak, David A; Gregory, Philip D; Lee, Gary; Holmes, Michael C

    2016-02-18

    The adoptive transfer of engineered T cells for the treatment of cancer, autoimmunity, and infectious disease is a rapidly growing field that has shown great promise in recent clinical trials. Nuclease-driven genome editing provides a method in which to precisely target genetic changes to further enhance T cell function in vivo. We describe the development of a highly efficient method to genome edit both primary human CD8 and CD4 T cells by homology-directed repair at a pre-defined site of the genome. Two different homology donor templates were evaluated, representing both minor gene editing events (restriction site insertion) to mimic gene correction, or the more significant insertion of a larger gene cassette. By combining zinc finger nuclease mRNA delivery with AAV6 delivery of a homologous donor we could gene correct 41% of CCR5 or 55% of PPP1R12C (AAVS1) alleles in CD8(+) T cells and gene targeting of a GFP transgene cassette in >40% of CD8(+) and CD4(+) T cells at both the CCR5 and AAVS1 safe harbor locus, potentially providing a robust genome editing tool for T cell-based immunotherapy. PMID:26527725

  18. Identification of human-specific AluS elements through comparative genomics.

    Science.gov (United States)

    Lee, Jae; Kim, Yun-Ji; Mun, Seyoung; Kim, Heui-Soo; Han, Kyudong

    2015-01-25

    Mobile elements are responsible for ~45% of the human genome. Among them is the Alu element, accounting for 10% of the human genome (>1.1million copies). Several studies of Alu elements have reported that they are frequently involved in human genetic diseases and genomic rearrangements. In this study, we investigated the AluS subfamily, which is a relatively old Alu subfamily and has the highest copy number in primate genomes. Previously, a set of 263 human-specific AluS insertions was identified in the human genome. To validate these, we compared each of the human-specific AluS loci with its pre-insertion site in other primate genomes, including chimpanzee, gorilla, and orangutan. We obtained 24 putative human-specific AluS candidates via the in silico analysis and manual inspection, and then tried to verify them using PCR amplification and DNA sequencing. Through the PCR product sequencing, we were able to detect two instances of near-parallel Alu insertions in nearby sites that led to computational false negatives. Finally, we computationally and experimentally verified 23 human-specific AluS elements. We reported three alternative Alu insertion events, which are accompanied by filler DNA and/or Alu retrotransposition mediated-deletion. Bisulfite sequencing was carried out to examine DNA methylation levels of human-specific AluS elements. The results showed that fixed AluS elements are hypermethylated compared with polymorphic elements, indicating a possible relation between DNA methylation and Alu fixation in the human genome. PMID:25447892

  19. Methylation status of individual CpG sites within Alu elements in the human genome and Alu hypomethylation in gastric carcinomas

    International Nuclear Information System (INIS)

    Alu methylation is correlated with the overall level of DNA methylation and recombination activity of the genome. However, the maintenance and methylation status of each CpG site within Alu elements (Alu) and its methylation status have not well characterized. This information is useful for understanding natural status of Alu in the genome and helpful for developing an optimal assay to quantify Alu hypomethylation. Bisulfite clone sequencing was carried out in 14 human gastric samples initially. A Cac8I COBRA-DHPLC assay was developed to detect methylated-Alu proportion in cell lines and 48 paired gastric carcinomas and 55 gastritis samples. DHPLC data were statistically interpreted using SPSS version 16.0. From the results of 427 Alu bisulfite clone sequences, we found that only 27.2% of CpG sites within Alu elements were preserved (4.6 of 17 analyzed CpGs, A ~ Q) and that 86.6% of remaining-CpGs were methylated. Deamination was the main reason for low preservation of methylation targets. A high correlation coefficient of methylation was observed between Alu clones and CpG site J (0.963), A (0.950), H (0.946), D (0.945). Comethylation of the sites H and J were used as an indicator of the proportion of methylated-Alu in a Cac8I COBRA-DHPLC assay. Validation studies showed that hypermethylation or hypomethylation of Alu elements in human cell lines could be detected sensitively by the assay after treatment with 5-aza-dC and M.SssI, respectively. The proportion of methylated-Alu copies in gastric carcinomas (3.01%) was significantly lower than that in the corresponding normal samples (3.19%) and gastritis biopsies (3.23%). Most Alu CpG sites are deaminated in the genome. 27% of Alu CpG sites represented in our amplification products. 87% of the remaining CpG sites are methylated. Alu hypomethylation in primary gastric carcinomas could be detected with the Cac8I COBRA-DHPLC assay quantitatively

  20. Design Approach for Fault Recoverable ALU with Improved Fault Tolerance

    OpenAIRE

    Ankit K V; S Murali Narasimham; Viajya Prakash A M

    2015-01-01

    A new design for fault tolerant and fault recoverable ALU System has been proposed in this paper. Reliability is one of the most critical factors that have to be considered during the designing phase of any IC. In critical applications like Medical equipment & Military applications this reliability factor plays a very critical role in determining the acceptance of product. Insertion of special modules in the main design for reliability enhancement will give considerable amount of ...

  1. Marketingový mix firmy ALU KOLA CB

    OpenAIRE

    Urban, Karel

    2011-01-01

    This bachelor thesis is focused on a marketing mix practical application in my own company ALU KOLA CB. My company sells alloy wheels and tyres for personal cars. In a literary review are introduced and explained terms marketing, marketing mix and its parts - product, price, place and promotion. In a practical part of this thesis are these terms applied on my company. The end of this part contains results and improvement suggestions.

  2. Recognition of tRNALeu by Aquifex aeolicus leucyl-tRNA synthetase during the aminoacylation and editing steps

    OpenAIRE

    Yao, Peng; Zhu, Bin; Jaeger, Sophie; Eriani, Gilbert; Wang, En-Duo

    2008-01-01

    Recognition of tRNA by the cognate aminoacyl-tRNA synthetase during translation is crucial to ensure the correct expression of the genetic code. To understand tRNALeu recognition sets and their evolution, the recognition of tRNALeu by the leucyl-tRNA synthetase (LeuRS) from the primitive hyperthermophilic bacterium Aquifex aeolicus was studied by RNA probing and mutagenesis. The results show that the base A73; the core structure of tRNA formed by the tertiary interactions U8–A14, G18–U55 and ...

  3. Analysis of western lowland gorilla (Gorilla gorilla gorilla) specific Alu repeats

    OpenAIRE

    McLain, Adam T.; Carman, Glenn W; Fullerton, Mitchell L; Beckstrom, Thomas O; Gensler, William; Meyer, Thomas J.; Faulk, Christopher; Batzer, Mark A.

    2013-01-01

    Background Research into great ape genomes has revealed widely divergent activity levels over time for Alu elements. However, the diversity of this mobile element family in the genome of the western lowland gorilla has previously been uncharacterized. Alu elements are primate-specific short interspersed elements that have been used as phylogenetic and population genetic markers for more than two decades. Alu elements are present at high copy number in the genomes of all primates surveyed thus...

  4. Cloning, expression, purification, crystallization and preliminary X-ray crystallographic analyses of threonyl-tRNA synthetase editing domain from Aeropyrum pernix

    International Nuclear Information System (INIS)

    The editing domain of threonyl-tRNA synthetase from the archaeon Aeropyrum pernix has been overexpressed, purified and crystallized. The crystal diffracted to a resolution of 1.66 Å. The proofreading function of aminoacyl-tRNA synthetases is crucial in maintaining the fidelity of protein synthesis. Most archaeal threonyl-tRNA synthetases (ThrRSs) possess a unique proofreading domain unrelated to their eukaryotic/bacterial counterpart. The crystal structure of this domain from the archaeon Pyrococcus abysii in complex with its cognate and noncognate substrate analogues had given insights into its catalytic and discriminatory mechanisms. To probe further into the mechanistic and evolutionary aspects of this domain, work has been extended to another archaeon Aeropyrum pernix. The organism possesses two proteins corresponding to threonyl-tRNA synthetase, i.e. ThrRS1 and ThrRS2, encoded by two different genes, thrS1 and thrS2, respectively. ThrRS1 is responsible for aminoacylation and ThrRS2 for proofreading activity. Here the purification, crystallization and preliminary X-ray crystallographic investigation of the N-terminal proofreading domain of ThrRS2 from A. pernix is reported. The crystals belong to either the P41212 or P43212 space group and consist of one monomer per asymmetric unit

  5. Design, Analysis, Implementation and Synthesis of 16 bit Reversible ALU by using Xilinx 12.2

    Directory of Open Access Journals (Sweden)

    S.Anusha

    2014-04-01

    Full Text Available In the modern world, Arithmetic Logic Unit (ALU is one of the most crucial components of any system and is used in many appliances like calculators, cell phones, and computers and so on. An arithmetic logic unit is a multi-functional circuit that conditionally performs one of several possible functions on two operands A and B depending on control inputs. This paper proposes the design of programmable reversible logic gate structures, targeted for the ALU implementation and their use in the realization of an efficient reversible ALU. Reversible or information-lossless circuits have applications in digital signal processing, communication, computer graphics and cryptography. This ALU consists of thirteen operations, 5 arithmetic, 4 logical operations and 4 shifting operations. All the modules are being designed using the basic reversible gates. Using reversible logic gates instead of traditional logic AND/OR gates, a reversible ALU whose function is the same as traditional ALU is constructed. Comparing with the number of input bits and the discarded bits of the traditional ALU, the reversible ALU significantly reduce the use and loss of information bits. The proposed reversible 16-bit ALU reduces the information bits use and loss by reusing the logic information bits logically and realizes the goal of lowering power consumption of logic circuits. Programmable reversible logic gates are realized in Verilog by using XILINX 12.2. Key words:

  6. Gene editing by co-transformation of TALEN and chimeric RNA/DNA oligonucleotides on the rice OsEPSPS gene and the inheritance of mutations.

    Directory of Open Access Journals (Sweden)

    Mugui Wang

    Full Text Available Although several site-specific nucleases (SSNs, such as zinc-finger nucleases (ZFNs, transcription activator-like effector nucleases (TALENs, and the clustered regularly interspaced short palindromic repeat (CRISPR/Cas, have emerged as powerful tools for targeted gene editing in many organisms, to date, gene targeting (GT in plants remains a formidable challenge. In the present study, we attempted to substitute a single base in situ on the rice OsEPSPS gene by co-transformation of TALEN with chimeric RNA/DNA oligonucleotides (COs, including different strand composition such as RNA/DNA (C1 or DNA/RNA (C2 but contained the same target base to be substituted. In contrast to zero GT event obtained by the co-transformation of TALEN with homologous recombination plasmid (HRP, we obtained one mutant showing target base substitution although accompanied by undesired deletion of 12 bases downstream the target site from the co-transformation of TALEN and C1. In addition to this typical event, we also obtained 16 mutants with different length of base deletions around the target site among 105 calli lines derived from transformation of TALEN alone (4/19 as well as co-transformation of TELAN with either HRP (5/30 or C1 (2/25 or C2 (5/31. Further analysis demonstrated that the homozygous gene-edited mutants without foreign gene insertion could be obtained in one generation. The induced mutations in transgenic generation were also capable to pass to the next generation stably. However, the genotypes of mutants did not segregate normally in T1 population, probably due to lethal mutations. Phenotypic assessments in T1 generation showed that the heterozygous plants with either one or three bases deletion on target sequence, called d1 and d3, were more sensitive to glyphosate and the heterozygous d1 plants had significantly lower seed-setting rate than wild-type.

  7. 基于转录组测序数据识别黑猩猩RNA编辑位点%Identification of RNA Editing Sites in Chimpanzee by Transcriptome-wide Sequencing Data

    Institute of Scientific and Technical Information of China (English)

    王端青; 何涛; 汪莉; 王玉民; 邵卫东

    2012-01-01

    RNA editing is a widespread post-transcriptional modification mechanism that alters genetic information at the RNA level by nucleotide insertions, deletions or substitutions, which can contribute to the diversification of the transcriptome and proteome. Although tens of thousands of A-to-I RNA editing events have been found in humans, there is limited knowledge of RNA editing in other nonhuman primates. For exploring the mechanism as well as potential functions of the RNA editing events in chimpanzee, we identified RNA editing sites based on chimpanzee RNA-Seq data here. By aligning between RNA-Seq data and chimpanzee genome sequences with TopHat software, all RNA-DNA mismatch sites were regarded as a candidate set. Low quality sites were filtered out by using both genome and transcriptome sequencing quality scores. The other filters containing uncertainty of sequencing at 3'-terminial positions, read coverage, SNP sites and estimated editing level were also applied on the candidate set. Statistical tests based on the Binomial distribution and Bonferroni multiple testing correction were performed on each candidate site to remove random errors between genome and transcriptome. Then, we detected tissue- and sex-specific RNA editing sites using bioinformatics approaches based on the Fisher's exact test and the Bonferroni multiple testing correction. The Two Sample Logo software was used to analyze the feature of the sequences surrounding the RNA editing site. A total of 8 334 RNA editing sites were identified in chimpanzee transcriptome and all 12 possible categories of discordances were observed. The top four distributions were A-to-G, U-to-C, G-to-A and C-to-U editing sites, which contained 1 995, 1 452, 1 293 and 1 101 sites, respectively. Forty-one editing sites alter amino acid residues, one of them creates a new stop codon which may shorten the KRT31 protein and affect its activity. Three editing sites damage the binding of microRNA potentially. Six hundred and

  8. Synthetic RNA Polymerase III Promoters Facilitate High-Efficiency CRISPR-Cas9-Mediated Genome Editing in Yarrowia lipolytica.

    Science.gov (United States)

    Schwartz, Cory M; Hussain, Murtaza Shabbir; Blenner, Mark; Wheeldon, Ian

    2016-04-15

    The oleaginous yeast Yarrowia lipolytica is a valuable microbial host for chemical production because it has a high capacity to synthesize, modify, and store intracellular lipids; however, rapid strain development has been hampered by the limited availability of genome engineering tools. We address this limitation by adapting the CRISPR-Cas9 system from Streptococcus pyogenes for markerless gene disruption and integration in Y. lipolytica. Single gene disruption efficiencies of 92% and higher were achieved when single guide RNAs (sgRNA) were transcribed with synthetic hybrid promoters that combine native RNA polymerase III (Pol III) promoters with tRNA. The Pol III-tRNA hybrid promoters exploit endogenous tRNA processing to produce mature sgRNA for Cas9 targeting. The highest efficiencies were achieved with a SCR1'-tRNA(Gly) promoter and Y. lipolytica codon-optimized Cas9 expressed from a UAS1B8-TEF promoter. Cotransformation of the Cas9 and sgRNA expressing plasmid with a homologous recombination donor plasmid resulted in markerless homologous recombination efficiency of over 64%. Homologous recombination was observed in 100% of transformants when nonhomologous end joining was disrupted. The end result of these studies was the development of pCRISPRyl, a modular tool for markerless gene disruption and integration in Y. lipolytica. PMID:26714206

  9. A comparison of 100 human genes using an alu element-based instability model.

    Directory of Open Access Journals (Sweden)

    George W Cook

    Full Text Available The human retrotransposon with the highest copy number is the Alu element. The human genome contains over one million Alu elements that collectively account for over ten percent of our DNA. Full-length Alu elements are randomly distributed throughout the genome in both forward and reverse orientations. However, full-length widely spaced Alu pairs having two Alus in the same (direct orientation are statistically more prevalent than Alu pairs having two Alus in the opposite (inverted orientation. The cause of this phenomenon is unknown. It has been hypothesized that this imbalance is the consequence of anomalous inverted Alu pair interactions. One proposed mechanism suggests that inverted Alu pairs can ectopically interact, exposing both ends of each Alu element making up the pair to a potential double-strand break, or "hit". This hypothesized "two-hit" (two double-strand breaks potential per Alu element was used to develop a model for comparing the relative instabilities of human genes. The model incorporates both 1 the two-hit double-strand break potential of Alu elements and 2 the probability of exon-damaging deletions extending from these double-strand breaks. This model was used to compare the relative instabilities of 50 deletion-prone cancer genes and 50 randomly selected genes from the human genome. The output of the Alu element-based genomic instability model developed here is shown to coincide with the observed instability of deletion-prone cancer genes. The 50 cancer genes are collectively estimated to be 58% more unstable than the randomly chosen genes using this model. Seven of the deletion-prone cancer genes, ATM, BRCA1, FANCA, FANCD2, MSH2, NCOR1 and PBRM1, were among the most unstable 10% of the 100 genes analyzed. This algorithm may lay the foundation for comparing genetic risks posed by structural variations that are unique to specific individuals, families and people groups.

  10. CRISPR/CAS9-Mediated Genome Editing of miRNA-155 Inhibits Proinflammatory Cytokine Production by RAW264.7 Cells

    Directory of Open Access Journals (Sweden)

    Weixia Jing

    2015-01-01

    Full Text Available MicroRNA 155 (miR-155 is a key proinflammatory regulator in clinical and experimental rheumatoid arthritis (RA. Here we generated a miR-155 genome knockout (GKO RAW264.7 macrophage cell line using the clustered regulatory interspaced short palindromic repeat (CRISPR/CRISPR-associated protein 9 (CAS9 technology. While upregulating the Src homology-2 domain-containing inositol 5-phosphatase 1 (SHIP1, the miR-155 GKO line is severely impaired in producing proinflammatory cytokines but slightly increased in osteoclastogenesis upon treatment with receptor activator of nuclear factor-κB ligand (RANKL. Taken together, our results suggest that genome editing of miR-155 holds the potential as a therapeutic strategy in RA.

  11. Orangutan Alu quiescence reveals possible source element: support for ancient backseat drivers

    Directory of Open Access Journals (Sweden)

    Walker Jerilyn A

    2012-04-01

    Full Text Available Abstract Background Sequence analysis of the orangutan genome revealed that recent proliferative activity of Alu elements has been uncharacteristically quiescent in the Pongo (orangutan lineage, compared with all previously studied primate genomes. With relatively few young polymorphic insertions, the genomic landscape of the orangutan seemed like the ideal place to search for a driver, or source element, of Alu retrotransposition. Results Here we report the identification of a nearly pristine insertion possessing all the known putative hallmarks of a retrotranspositionally competent Alu element. It is located in an intronic sequence of the DGKB gene on chromosome 7 and is highly conserved in Hominidae (the great apes, but absent from Hylobatidae (gibbon and siamang. We provide evidence for the evolution of a lineage-specific subfamily of this shared Alu insertion in orangutans and possibly the lineage leading to humans. In the orangutan genome, this insertion contains three orangutan-specific diagnostic mutations which are characteristic of the youngest polymorphic Alu subfamily, AluYe5b5_Pongo. In the Homininae lineage (human, chimpanzee and gorilla, this insertion has acquired three different mutations which are also found in a single human-specific Alu insertion. Conclusions This seemingly stealth-like amplification, ongoing at a very low rate over millions of years of evolution, suggests that this shared insertion may represent an ancient backseat driver of Alu element expansion.

  12. Alu Insertions and Genetic Diversity: A Preliminary Investigation by an Undergraduate Bioinformatics Class

    Science.gov (United States)

    Elwess, Nancy L.; Duprey, Stephen L.; Harney, Lindesay A.; Langman, Jessie E.; Marino, Tara C.; Martinez, Carolina; McKeon, Lauren L.; Moss, Chantel I. E.; Myrie, Sasha S.; Taylor, Luke Ryan

    2008-01-01

    "Alu"-insertion polymorphisms were used by an undergraduate Bioinformatics class to study how these insertion sites could be the basis for an investigation in human population genetics. Based on the students' investigation, both allele and genotype "Alu" frequencies were determined for African-American and Japanese populations as well as a…

  13. Genome-wide tracking of unmethylated DNA Alu repeats in normal and cancer cells

    DEFF Research Database (Denmark)

    Rodriguez, Jairo; Vives, Laura; Jordà, Mireia;

    2008-01-01

    . Normal colon epithelial cells contain in average 25 486 +/- 10 157 unmethylated Alu's per haploid genome, while in tumor cells this figure is 41 995 +/- 17 187 (P = 0.004). There is an inverse relationship in Alu families with respect to their age and methylation status: the youngest elements exhibit the...

  14. Analysis of the features and source gene composition of the AluYg6 subfamily of human retrotransposons

    Directory of Open Access Journals (Sweden)

    Brookfield John FY

    2007-07-01

    Full Text Available Abstract Background Alu elements are a family of SINE retrotransposons in primates. They are classified into subfamilies according to specific diagnostic mutations from the general Alu consensus. It is now believed that there may be several retrotranspositionally-competent source genes within an Alu subfamily. To investigate the evolution of young Alu elements it is critical to have access to complete subfamilies, which, following the release of the final human genome assembly, can now be obtained using in silico methods. Results 380 elements belonging to the young AluYg6 subfamily were identified in the human genome, a number significantly exceeding prior expectations. An AluYg6 element was also identified in the chimpanzee genome, indicating that the subfamily is older than previously estimated, and appears to have undergone a period of dormancy before its expansion. The relative contributions of back mutation and gene conversion to variation at the six diagnostic positions are examined, and cases of complete forward gene conversion events are reported. Two small subfamilies derived from AluYg6 have been identified, named AluYg6a2 and AluYg5b3, which contain 40 and 27 members, respectively. These small subfamilies are used to illustrate the ambiguity regarding Alu subfamily definition, and to assess the contribution of secondary source genes to the AluYg6 subfamily. Conclusion The number of elements in the AluYg6 subfamily greatly exceeds prior expectations, indicating that the current knowledge of young Alu subfamilies is incomplete, and that prior analyses that have been carried out using these data may have generated inaccurate results. A definition of primary and secondary source genes has been provided, and it has been shown that several source genes have contributed to the proliferation of the AluYg6 subfamily. Access to the sequence data for the complete AluYg6 subfamily will be invaluable in future computational analyses investigating

  15. Design Approach for Fault Recoverable ALU with Improved Fault Tolerance

    Directory of Open Access Journals (Sweden)

    Ankit K V

    2015-08-01

    Full Text Available A new design for fault tolerant and fault recoverable ALU System has been proposed in this paper. Reliability is one of the most critical factors that have to be considered during the designing phase of any IC. In critical applications like Medical equipment & Military applications this reliability factor plays a very critical role in determining the acceptance of product. Insertion of special modules in the main design for reliability enhancement will give considerable amount of area & power penalty. So, a novel approach to this problem is to find ways for reusing the already available components in digital system in efficient way to implement recoverable methodologies. Triple Modular Redundancy (TMR has traditionally used for protecting digital logic from the SEUs (single event upset by triplicating the critical components of the system to give fault tolerance to system. ScTMR- Scan chain-based error recovery TMR technique provides recovery for all internal faults. ScTMR uses a roll-forward approach and employs the scan chain implemented in the circuits for testability purposes to recover the system to fault-free state. The proposed design will incorporate a ScTMR controller over TMR system of ALU and will make the system fault tolerant and fault recoverable. Hence, proposed design will be more efficient & reliable to use in critical applications, than any other design present till today.

  16. Improved distance analysis in RNA using network-editing techniques for overcoming errors due to spin diffusion

    International Nuclear Information System (INIS)

    Multispin magnetization transfer, or spin diffusion, is a significant source of error in NOESY-derived distance measurements for the determination of nucleic acid solution structures. The BD-NOESY and CBD-NOESY experiments, which allow the measurement of interproton distances with greatly reduced contributions from spin diffusion, have been adapted to structural analysis in RNA oligonucleotides. The techniques are applied to a lead-dependent ribozyme (LZ2). We demonstrate the measurement of both aromatic proton-aromatic proton NOEs free of spin diffusion involving the intervening ribose moieties and aromatic proton-ribose proton NOEs free of the efficient cross-relaxation within the ribose ring. In LZ2, the accuracy and precision of the resulting distances are significantly improved. We also find that, by allowing the use of longer mixing times with greater sensitivity, the experimental attenuation of spin diffusion in RNA increases the distance range of interactions that can be analyzed. This effect permits measurement of important long-range distances in LZ2 that are not accessible with standard techniques. Thus, these techniques allow the simultaneous optimization of the number, accuracy, and precision of distance constraints used for RNA structure determinations

  17. To edit or not to edit: regulation of ADAR editing specificity and efficiency.

    Science.gov (United States)

    Deffit, Sarah N; Hundley, Heather A

    2016-01-01

    Hundreds to millions of adenosine (A)-to-inosine (I) modifications are present in eukaryotic transcriptomes and play an essential role in the creation of proteomic and phenotypic diversity. As adenosine and inosine have different base-pairing properties, the functional consequences of these modifications or 'edits' include altering coding potential, splicing, and miRNA-mediated gene silencing of transcripts. However, rather than serving as a static control of gene expression, A-to-I editing provides a means to dynamically rewire the genetic code during development and in a cell-type specific manner. Interestingly, during normal development, in specific cells, and in both neuropathological diseases and cancers, the extent of RNA editing does not directly correlate with levels of the substrate mRNA or the adenosine deaminase that act on RNA (ADAR) editing enzymes, implying that cellular factors are required for spatiotemporal regulation of A-to-I editing. The factors that affect the specificity and extent of ADAR activity have been thoroughly dissected in vitro. Yet, we still lack a complete understanding of how specific ADAR family members can selectively deaminate certain adenosines while others cannot. Additionally, in the cellular environment, ADAR specificity and editing efficiency is likely to be influenced by cellular factors, which is currently an area of intense investigation. Data from many groups have suggested two main mechanisms for controlling A-to-I editing in the cell: (1) regulating ADAR accessibility to target RNAs and (2) protein-protein interactions that directly alter ADAR enzymatic activity. Recent studies suggest cis- and trans-acting RNA elements, heterodimerization and RNA-binding proteins play important roles in regulating RNA editing levels in vivo. WIREs RNA 2016, 7:113-127. doi: 10.1002/wrna.1319. PMID:26612708

  18. Higher Alu methylation levels in catch-up growth in twenty-year-old offsprings.

    Directory of Open Access Journals (Sweden)

    Kittipan Rerkasem

    Full Text Available Alu elements and long interspersed element-1 (LINE-1 or L1 are two major human intersperse repetitive sequences. Lower Alu methylation, but not LINE-1, has been observed in blood cells of people in old age, and in menopausal women having lower bone mass and osteoporosis. Nevertheless, Alu methylation levels also vary among young individuals. Here, we explored phenotypes at birth that are associated with Alu methylation levels in young people. In 2010, 249 twenty-years-old volunteers whose mothers had participated in a study association between birth weight (BW and nutrition during pregnancy in 1990, were invited to take part in our present study. In this study, the LINE-1 and Alu methylation levels and patterns were measured in peripheral mononuclear cells and correlated with various nutritional parameters during intrauterine and postnatal period of offspring. This included the amount of maternal intake during pregnancy, the mother's weight gain during pregnancy, birth weight, birth length, and the rate of weight gain in the first year of life. Catch-up growth (CUG was defined when weight during the first year was >0.67 of the standard score, according to WHO data. No association with LINE-1 methylation was identified. The mean level of Alu methylation in the CUG group was significantly higher than those non-CUG (39.61% and 33.66 % respectively, P < 0.0001. The positive correlation between the history of CUG in the first year and higher Alu methylation indicates the role of Alu methylation, not only in aging cells, but also in the human growth process. Moreover, here is the first study that demonstrated the association between a phenotype during the newborn period and intersperse repetitive sequences methylation during young adulthood.

  19. AluY-mediated germline deletion, duplication and somatic stem cell reversion in UBE2T defines a new subtype of Fanconi anemia.

    Science.gov (United States)

    Virts, Elizabeth L; Jankowska, Anna; Mackay, Craig; Glaas, Marcel F; Wiek, Constanze; Kelich, Stephanie L; Lottmann, Nadine; Kennedy, Felicia M; Marchal, Christophe; Lehnert, Erik; Scharf, Rüdiger E; Dufour, Carlo; Lanciotti, Marina; Farruggia, Piero; Santoro, Alessandra; Savasan, Süreyya; Scheckenbach, Kathrin; Schipper, Jörg; Wagenmann, Martin; Lewis, Todd; Leffak, Michael; Farlow, Janice L; Foroud, Tatiana M; Honisch, Ellen; Niederacher, Dieter; Chakraborty, Sujata C; Vance, Gail H; Pruss, Dmitry; Timms, Kirsten M; Lanchbury, Jerry S; Alpi, Arno F; Hanenberg, Helmut

    2015-09-15

    Fanconi anemia (FA) is a rare inherited disorder clinically characterized by congenital malformations, progressive bone marrow failure and cancer susceptibility. At the cellular level, FA is associated with hypersensitivity to DNA-crosslinking genotoxins. Eight of 17 known FA genes assemble the FA E3 ligase complex, which catalyzes monoubiquitination of FANCD2 and is essential for replicative DNA crosslink repair. Here, we identify the first FA patient with biallelic germline mutations in the ubiquitin E2 conjugase UBE2T. Both mutations were aluY-mediated: a paternal deletion and maternal duplication of exons 2-6. These loss-of-function mutations in UBE2T induced a cellular phenotype similar to biallelic defects in early FA genes with the absence of FANCD2 monoubiquitination. The maternal duplication produced a mutant mRNA that could encode a functional protein but was degraded by nonsense-mediated mRNA decay. In the patient's hematopoietic stem cells, the maternal allele with the duplication of exons 2-6 spontaneously reverted to a wild-type allele by monoallelic recombination at the duplicated aluY repeat, thereby preventing bone marrow failure. Analysis of germline DNA of 814 normal individuals and 850 breast cancer patients for deletion or duplication of UBE2T exons 2-6 identified the deletion in only two controls, suggesting aluY-mediated recombinations within the UBE2T locus are rare and not associated with an increased breast cancer risk. Finally, a loss-of-function germline mutation in UBE2T was detected in a high-risk breast cancer patient with wild-type BRCA1/2. Cumulatively, we identified UBE2T as a bona fide FA gene (FANCT) that also may be a rare cancer susceptibility gene. PMID:26085575

  20. The genome editing revolution

    DEFF Research Database (Denmark)

    Stella, Stefano; Montoya, Guillermo

    2016-01-01

    In the last 10 years, we have witnessed a blooming of targeted genome editing systems and applications. The area was revolutionized by the discovery and characterization of the transcription activator-like effector proteins, which are easier to engineer to target new DNA sequences than the previo......In the last 10 years, we have witnessed a blooming of targeted genome editing systems and applications. The area was revolutionized by the discovery and characterization of the transcription activator-like effector proteins, which are easier to engineer to target new DNA sequences than...... sequence). This ribonucleoprotein complex protects bacteria from invading DNAs, and it was adapted to be used in genome editing. The CRISPR ribonucleic acid (RNA) molecule guides to the specific DNA site the Cas9 nuclease to cleave the DNA target. Two years and more than 1000 publications later, the CRISPR......-Cas system has become the main tool for genome editing in many laboratories. Currently the targeted genome editing technology has been used in many fields and may be a possible approach for human gene therapy. Furthermore, it can also be used to modifying the genomes of model organisms for studying human...

  1. Homologous recombination among three intragene Alu sequences causes an inversion-deletion resulting in the hereditary bleeding disorder glanzmann thrombasthenia

    Energy Technology Data Exchange (ETDEWEB)

    Li, L.; Bray, P.F. (Johns Hopkins Univ. Medical School, Baltimore, MD (United States))

    1993-07-01

    The crucial role of the human platelet fibrinogen receptor in maintaining normal hemostasis is best exemplified by the autosomal recessive bleeding disorder Glanzmann thrombasthenia (GT). The platelet fibrinogen receptor is a heterodimer composed of glycoproteins IIb (GPIIb) and IIIa (GPIIIa). Platelets from patients with GT have a quantitative or qualitative abnormality in GPIIb and GPIIIa and can neither bind fibrinogen nor aggregate. Very few genetic defects have been identified that cause this disorder. The authors describe a kindred with GT in which the affected individuals have a unique inversion-deletion mutation in the gene for GPIIIa. Patient platelets lacked both GPIIIa protein and mRNA. Southern blots of patient genomic DNA probed with an internal 1.0-kb GPIIIa cDNA suggested a large rearrangement of this gene but were normal when probed with small GPIIIa cDNA fragments that were outside the mutation. Cytogenetics and pulsed-field gel analysis of the GPIIIa gene were normal, making a translocation or a very large rearrangement unlikely. Additional Southern analyses suggested that the abnormality was not a small insertion. The authors constructed a patient genomic DNA library and isolated fragments containing the 5' and 3' breakpoints of the mutation. The nucleotide sequence from these genomic clones was determined and revealed that, relative to the normal gene, the mutant allele contained a 1-kb deletion immediately preceding a 15-kb inversion. The DNA breaks occurred in two inverted and one forward Alu sequence within the gene for GPIIIa and in the left, right, and left arms, respectively, of these sequences. There was a 5-bp repeat at the 3 terminus of the inversion. One copy of the repeat remained in the mutant allele breakpoint junction. The alignment and orientation of the different Alu sequences, as well as the position of the breakpoints, suggest that the inversion preceded the deletion in this complex rearrangement. 41 refs., 5 figs.

  2. A SINE in the genome of the cephalochordate amphioxus is an Alu element

    Directory of Open Access Journals (Sweden)

    2006-04-01

    Full Text Available Transposable elements of about 300 bp, termed “short interspersed nucleotide elements or SINEs are common in eukaryotes. However, Alu elements, SINEs containing restriction sites for the AluI enzyme, have been known only from primates. Here I report the first SINE found in the genome of the cephalochordate, amphioxus. It is an Alu element of 375 bp that does not share substantial identity with any genomic sequences in vertebrates. It was identified because it was located in the FoxD regulatory region in a cosmid derived from one individual, but absent from the two FoxD alleles of BACs from a second individual. However, searches of sequences of BACs and genomic traces from this second individual gave an estimate of 50-100 copies in the amphioxus genome. The finding of an Alu element in amphioxus raises the question of whether Alu elements in amphioxus and primates arose by convergent evolution or by inheritance from a common ancestor. Genome-wide analyses of transposable elements in amphioxus and other chordates such as tunicates, agnathans and cartilaginous fishes could well provide the answer.

  3. Learning Stochastic Tree Edit Distance

    OpenAIRE

    Bernard, Marc; Habrard, Amaury; Sebban, Marc

    2006-01-01

    Trees provide a suited structural representation to deal with complex tasks such as web information extraction, RNA secondary structure prediction, or conversion of tree structured documents. In this context, many applications require the calculation of similarities between tree pairs. The most studied distance is likely the tree edit distance for which improvements in terms of complexity have been achieved during the last decade. However, this classic edit distance usually uses a priori fixe...

  4. Chromosomal aberrations induced by the restriction endonucleases Alu I and Bam HI: comparison with X-rays

    International Nuclear Information System (INIS)

    Dose-effect relationships for the frequencies of polycentric chromosomes induced by the restriction endonucleases Alu I and Bam HI and by X-rays in Chinese hamster ovary (CHO) cells were analyzed and compared. 1 Gy of X-rays produce the same frequency of polycentric chromosomes as 2 units Alu I and 7.9 units Bam HI. (author)

  5. Circular RNAs are abundant, conserved, and associated with ALU repeats

    OpenAIRE

    Jeck, William R.; Sorrentino, Jessica A.; Wang, Kai; Slevin, Michael K.; Christin E Burd; Liu, Jinze; Marzluff, William F.; Sharpless, Norman E.

    2013-01-01

    Using a novel approach to identify exonic circular RNAs (ecircRNAs), the authors show widespread production of circular RNAs from a significant fraction of expressed genes in murine and human cells. They demonstrate that several of these noncoding transcripts are stable and abundant and can be targeted by siRNA. Bioinformatic analysis of the entire ecircRNA complement revealed gene features associated with RNA circularization.

  6. A young Alu subfamily amplified independently in human and African great apes lineages.

    OpenAIRE

    Zietkiewicz, E; Richer, C.; Makalowski, W; Jurka, J; Labuda, D

    1994-01-01

    A variety of Alu subfamilies amplified in primate genomes at different evolutionary time periods. Alu Sb2 belongs to a group of young subfamilies with a characteristic two-nucleotide deletion at positions 65/66. It consists of repeats having a 7-nucleotide duplication of a sequence segment involving positions 246 through 252. The presence of Sb2 inserts was examined in five genomic loci in 120 human DNA samples as well as in DNAs of higher primates. The lack of the insertional polymorphism se...

  7. Non-GMO genetically edited crop plants.

    Science.gov (United States)

    Kanchiswamy, Chidananda Nagamangala; Malnoy, Mickael; Velasco, Riccardo; Kim, Jin-Soo; Viola, Roberto

    2015-09-01

    Direct delivery of purified Cas9 protein with guide RNA into plant cells, as opposed to plasmid-mediated delivery, displays high efficiency and reduced off-target effects. Following regeneration from edited cells, the ensuing plant is also likely to bypass genetically modified organism (GMO) legislation as the genome editing complex is degraded in the recipient cells. PMID:25978870

  8. Underwater video footage, March 2014, Faga'alu Bay, Tutuila Island, American Samoa

    Data.gov (United States)

    U.S. Geological Survey, Department of the Interior — Underwater video imagery was collected in March 2014 in the nearshore waters of Faga'alu Bay on the Island of Tutuila, American Samoa, as part of the U.S....

  9. Cluster editing

    DEFF Research Database (Denmark)

    Böcker, S.; Baumbach, Jan

    2013-01-01

    The Cluster Editing problem asks to transform a graph into a disjoint union of cliques using a minimum number of edge modifications. Although the problem has been proven NP-complete several times, it has nevertheless attracted much research both from the theoretical and the applied side. The...... algorithms for biological problems. © 2013 Springer-Verlag....... problem has been the inspiration for numerous algorithms in bioinformatics, aiming at clustering entities such as genes, proteins, phenotypes, or patients. In this paper, we review exact and heuristic methods that have been proposed for the Cluster Editing problem, and also applications of these...

  10. High-throughput sequencing of partially edited trypanosome mRNAs reveals barriers to editing progression and evidence for alternative editing.

    Science.gov (United States)

    Simpson, Rachel M; Bruno, Andrew E; Bard, Jonathan E; Buck, Michael J; Read, Laurie K

    2016-05-01

    Uridine insertion/deletion RNA editing in kinetoplastids entails the addition and deletion of uridine residues throughout the length of mitochondrial transcripts to generate translatable mRNAs. This complex process requires the coordinated use of several multiprotein complexes as well as the sequential use of noncoding template RNAs called guide RNAs. The majority of steady-state mitochondrial mRNAs are partially edited and often contain regions of mis-editing, termed junctions, whose role is unclear. Here, we report a novel method for sequencing entire populations of pre-edited partially edited, and fully edited RNAs and analyzing editing characteristics across populations using a new bioinformatics tool, the Trypanosome RNA Editing Alignment Tool (TREAT). Using TREAT, we examined populations of two transcripts, RPS12 and ND7-5', in wild-typeTrypanosoma brucei We provide evidence that the majority of partially edited sequences contain junctions, that intrinsic pause sites arise during the progression of editing, and that the mechanisms that mediate pausing in the generation of canonical fully edited sequences are distinct from those that mediate the ends of junction regions. Furthermore, we identify alternatively edited sequences that constitute plausible alternative open reading frames and identify substantial variability in the 5' UTRs of both canonical and alternatively edited sequences. This work is the first to use high-throughput sequencing to examine full-length sequences of whole populations of partially edited transcripts. Our method is highly applicable to current questions in the RNA editing field, including defining mechanisms of action for editing factors and identifying potential alternatively edited sequences. PMID:26908922

  11. Adenosine to Inosine editing frequency controlled by splicing efficiency.

    Science.gov (United States)

    Licht, Konstantin; Kapoor, Utkarsh; Mayrhofer, Elisa; Jantsch, Michael F

    2016-07-27

    Alternative splicing and adenosine to inosine (A to I) RNA-editing are major factors leading to co- and post-transcriptional modification of genetic information. Both, A to I editing and splicing occur in the nucleus. As editing sites are frequently defined by exon-intron basepairing, mRNA splicing efficiency should affect editing levels. Moreover, splicing rates affect nuclear retention and will therefore also influence the exposure of pre-mRNAs to the editing-competent nuclear environment. Here, we systematically test the influence of splice rates on RNA-editing using reporter genes but also endogenous substrates. We demonstrate for the first time that the extent of editing is controlled by splicing kinetics when editing is guided by intronic elements. In contrast, editing sites that are exclusively defined by exonic structures are almost unaffected by the splicing efficiency of nearby introns. In addition, we show that editing levels in pre- and mature mRNAs do not match. This phenomenon can in part be explained by the editing state of an RNA influencing its splicing rate but also by the binding of the editing enzyme ADAR that interferes with splicing. PMID:27112566

  12. Multilinguals and Wikipedia Editing

    CERN Document Server

    Hale, Scott A

    2013-01-01

    This article analyzes one month of edits to Wikipedia in order to examine the role of users editing multiple language editions (referred to as multilingual users). Such multilingual users may serve an important function in diffusing information across different language editions of the project, and prior work has suggested this could reduce the level of self-focus bias in each edition. This study finds multilingual users are much more active than their single-edition (monolingual) counterparts. They are found in all language editions, but smaller-sized editions with fewer users have a higher percentage of multilingual users than larger-sized editions. About a quarter of multilingual users always edit the same articles in multiple languages, while just over 40% of multilingual users edit different articles in different languages. When non-English users do edit a second language edition, that edition is most frequently English. Nonetheless, several regional and linguistic cross-editing patterns are also present...

  13. Development of two highly sensitive forensic sex determination assays based on human DYZ1 and Alu repetitive DNA elements.

    Science.gov (United States)

    Fazi, Amanda; Gobeski, Brianne; Foran, David

    2014-11-01

    Sex determination is a critical component of forensic identification, the standard genetic method for which is detection of the single copy amelogenin gene that has differing homologues on the X and Y chromosomes. However, this assay may not be sensitive enough when DNA samples are minute or highly compromised, thus other strategies for sex determination are needed. In the current research, two ultrasensitive sexing assays, based on real-time PCR and pyrosequencing, were developed targeting the highly repetitive elements DYZ1 on the Y chromosome and Alu on the autosomes. The DYZ1/Alu strategy was compared to amelogenin for overall sensitivity based on high molecular weight and degraded DNA, followed by assaying the sex of 34 touch DNA samples and DNA from 30 hair shafts. The real-time DYZ1/Alu assay proved to be approximately 1500 times more sensitive than its amelogenin counterpart based on high molecular weight DNA, and even more sensitive when sexing degraded DNA. The pyrosequencing DYZ1/Alu assay correctly sexed 26 of the touch DNAs, compared to six using amelogenin. Hair shaft DNAs showed equally improved sexing results using the DYZ1/Alu assays. Overall, both DYZ1/Alu assays were far more sensitive and accurate than was the amelogenin assay, and thus show great utility for sexing poor quality and low quantity DNA evidence. PMID:25168471

  14. SSTL Based Low Power Thermal Efficient WLAN Specific 32bit ALU Design on 28nm FPGA

    DEFF Research Database (Denmark)

    Kalia, Kartik; Pandey, Bishwajeet; Das, Teerath;

    2016-01-01

    with consideration of airflow toward hit sink and different frequency on which ALU operate in network processor or any WLAN devices. We have done total power analysis of WLAN operating on different frequencies. We have considered a set of frequencies, which are based on IEEE 802.11 standards. First we...... find out most efficient IO standard. While analyzing we found out that when WLAN device shift from 343.15K to 283.15K, there is maximum thermal power reduction in SSTL135_R as compared to all considered I/O standards. When we compared same I/Os for different frequencies we observed maximum thermal...... efficiency in SSTL15 at minimum and maximum temperature as compared to all other considered I/O standards. This design has application where 32bit ALU design is considered for designing an electronic device such as WLAN. The design can be implemented on different nano chips for better efficiency depending...

  15. Genome editing comes of age.

    Science.gov (United States)

    Kim, Jin-Soo

    2016-09-01

    Genome editing harnesses programmable nucleases to cut and paste genetic information in a targeted manner in living cells and organisms. Here, I review the development of programmable nucleases, including zinc finger nucleases (ZFNs), TAL (transcription-activator-like) effector nucleases (TALENs) and CRISPR (cluster of regularly interspaced palindromic repeats)-Cas9 (CRISPR-associated protein 9) RNA-guided endonucleases (RGENs). I specifically highlight the key advances that set the foundation for the rapid and widespread implementation of CRISPR-Cas9 genome editing approaches that has revolutionized the field. PMID:27490630

  16. Development and irradiation testing of Al-U3Si2 at Chalk River Laboratories

    International Nuclear Information System (INIS)

    Mini-elements containing Al-64 wt% U3Si2 (3.15 gU/cm3), with three discrete U3Si2 particle-size distributions, have been irradiated up to 93 at% burnup in the NRU reactor. The uranium silicide (U-7.0Si) was used in the as-cast condition, and contained up to 4 wt% free uranium in the U3Si2 matrix. Post-irradiation examinations (PIE) of the high-burnup elements have been recently completed. PIE included underwater and hot-cell examinations, immersion density measurements, neutron radiography, optical and scanning electron microscopy (SEM) with wavelength dispersion X-ray (WDX) analysis, and computerized image analysis of the fission-gas bubble-size distributions. The results show that the Al-U3Si2 swelled less than Al-U3Si fuel previously irradiated under similar conditions in NRU, and no significant swelling dependence on particle-size distribution was observed. Al-U3Si2 core volume increases ranged from 4.2 to 4.7 vol%, compared to 5.8 to 6.8 vol% for Al-U3Si fuel with identical uranium loadings. SEM examinations revealed that the U3Si2 (U-7.0Si) particles contained regions with relatively ordered, very dense populations of sub-micron fission-gas bubbles. In contrast, the gas bubbles are randomly distributed within U3Si (U-3.96Si) particles, vary widely in size, and small bubbles coalesce to form larger bubbles. The capability of U3Si2 to retain fission gas in small bubbles accounts for the lower swelling. (author)

  17. In situ hybridization of bat chromosomes with human (TTAGGGn probe, after previous digestion with Alu I

    Directory of Open Access Journals (Sweden)

    Karina de Cassia Faria

    2002-01-01

    Full Text Available The purpose of this work was to verify the ability of the enzyme Alu I to cleave and/or remove satellite DNA sequences from heterochromatic regions in chromosomes of bats, by identifying the occurrence of modifications in the pattern of fluorescence in situ hybridization with telomeric DNA. The localization and fluorescence intensity of the telomeric DNA sites of the Alu-digested and undigested chromosomes of species Eumops glaucinus, Carollia perspicillata, and Platyrrhinus lineatus were analyzed. Telomeric sequences were detected at the termini of chromosomes of all three species, although, in C. perspicillata, the signals were very faint or absent in most chromosomes. This finding was interpreted as being due to a reduced number of copies of the telomeric repeat, resulting from extensive telomeric association and/or rearrangements undergone by the chromosomes of Carollia. Fluorescent signals were also observed in centromeric and pericentromeric regions in several two-arm chromosomes of E. glaucinus and C. perspicillata. In E. glaucinus and P. lineatus, some interstitial and terminal telomeric sites were observed to be in association with regions of constitutive heterochromatin and ribosomal DNA (NORs. After digestion, these telomeric sites showed a significant decrease in signal intensity, indicating that enzyme Alu I cleaves and/or removes part of the satellite DNA present in these regions. These results suggest that the telomeric sequence is a component of the heterochromatin, and that the C-band- positive regions of bat chromosomes have a different DNA composition.

  18. Structure of a putative trans-editing enzyme for prolyl-tRNA synthetase from Aeropyrum pernix K1 at 1.7 Å resolution

    International Nuclear Information System (INIS)

    The three-dimensional structure of the APE2540 protein from A. pernix K1 has been determined by the multiple anomalous dispersion method at 1.7 Å resolution. The structure includes two monomers in the asymmetric unit and shares structural similarity with the YbaK protein or cysteinyl-tRNAPro deacylase from H. influenzae. The crystal structure of APE2540, the putative trans-editing enzyme ProX from Aeropyrum pernix K1, was determined in a high-throughput manner. The crystal belongs to the monoclinic space group P21, with unit-cell parameters a = 47.4, b = 58.9, c = 53.6 Å, β = 106.8°. The structure was solved by the multiwavelength anomalous dispersion method at 1.7 Å and refined to an R factor of 16.8% (Rfree = 20.5%). The crystal structure includes two protein molecules in the asymmetric unit. Each monomer consists of eight β-strands and seven α-helices. A structure-homology search revealed similarity between the trans-editing enzyme YbaK (or cysteinyl-tRNAPro deacylase) from Haemophilus influenzae (HI1434; 22% sequence identity) and putative ProX proteins from Caulobacter crescentus (16%) and Agrobacterium tumefaciens (21%)

  19. Human nucleosomes: special role of CG dinucleotides and Alu-nucleosomes

    Directory of Open Access Journals (Sweden)

    Trifonov Edward N

    2011-05-01

    Full Text Available Abstract Background The periodical occurrence of dinucleotides with a period of 10.4 bases now is undeniably a hallmark of nucleosome positioning. Whereas many eukaryotic genomes contain visible and even strong signals for periodic distribution of dinucleotides, the human genome is rather featureless in this respect. The exact sequence features in the human genome that govern the nucleosome positioning remain largely unknown. Results When analyzing the human genome sequence with the positional autocorrelation method, we found that only the dinucleotide CG shows the 10.4 base periodicity, which is indicative of the presence of nucleosomes. There is a high occurrence of CG dinucleotides that are either 31 (10.4 × 3 or 62 (10.4 × 6 base pairs apart from one another - a sequence bias known to be characteristic of Alu-sequences. In a similar analysis with repetitive sequences removed, peaks of repeating CG motifs can be seen at positions 10, 21 and 31, the nearest integers of multiples of 10.4. Conclusions Although the CG dinucleotides are dominant, other elements of the standard nucleosome positioning pattern are present in the human genome as well. The positional autocorrelation analysis of the human genome demonstrates that the CG dinucleotide is, indeed, one visible element of the human nucleosome positioning pattern, which appears both in Alu sequences and in sequences without repeats. The dominant role that CG dinucleotides play in organizing human chromatin is to indicate the involvement of human nucleosomes in tuning the regulation of gene expression and chromatin structure, which is very likely due to cytosine-methylation/-demethylation in CG dinucleotides contained in the human nucleosomes. This is further confirmed by the positions of CG-periodical nucleosomes on Alu sequences. Alu repeats appear as monomers, dimers and trimers, harboring two to six nucleosomes in a run. Considering the exceptional role CG dinucleotides play in the

  20. Identification of homogeneously staining regions by G-banding and chromosome microdissection, and FISH marker selection using human Alu sequence primers in a scleractinian coral Coelastrea aspera Verrill, 1866 (Cnidaria).

    Science.gov (United States)

    Taguchi, Takahiro; Kubota, Satoshi; Mezaki, Takuma; Tagami, Erika; Sekida, Satoko; Nakachi, Shu; Okuda, Kazuo; Tominaga, Akira

    2016-01-01

    Karyotype analysis was performed on the scleractinian coral Coelastrea aspera Verrill, 1866, commonly found along temperate coasts in Japan (30-35°N) and in coastal waters in the Indian and Pacific oceans. G-banding of Coelastrea aspera was successfully performed, although the banding pattern was not as clear as that in mammals. The karyogram clearly revealed that this coral had a homogeneously staining region (hsr) in chromosome 11. This hsr consisted of ribosomal RNA (rRNA) related genes, which was demonstrated by fluorescence in situ hybridization (FISH) with probes generated using 28S ribosomal DNA (rDNA) primers and those generated through chromosome microdissection. In addition, we conducted silver-stained nucleolus organizer region (Ag-NOR) analysis and found Ag depositions in the interphase nuclei but not on rRNA gene loci and hsr(s) in the mitotic stage. The hsr of this coral was observed in approximately 50% of the metaphase spreads analyzed. This may explain the diversity of coral rDNA based on the molecular study of sequence analysis. Furthermore, it was discovered that human telomere and Alu repeated sequences were present in this Coelastrea aspera. Probes derived from human Alu sequences are expected to play an important role in the classification of corals. Overall, our data can be of great value in discriminating among scleractinian coral species and understanding their genetics, including chromosomal evolution. PMID:27186338

  1. High transcript abundance, RNA editing, and small RNAs in intergenic regions within the massive mitochondrial genome of the angiosperm Silene noctiflora

    Czech Academy of Sciences Publication Activity Database

    Wu, Z.; Stone, James D.; Štorchová, Helena; Sloan, D.B.

    2015-01-01

    Roč. 16, NOV 14 (2015), s. 938. ISSN 1471-2164 R&D Projects: GA MŠk(CZ) EE2.3.30.0048 Institutional support: RVO:61389030 Keywords : Antisense RNA * Junk DNA * mtDNA Subject RIV: EB - Genetics ; Molecular Biology Impact factor: 3.986, year: 2014

  2. A genomewide screen for suppressors of Alu-mediated rearrangements reveals a role for PIF1.

    Directory of Open Access Journals (Sweden)

    Karen M Chisholm

    Full Text Available Alu-mediated rearrangement of tumor suppressor genes occurs frequently during carcinogenesis. In breast cancer, this mechanism contributes to loss of the wild-type BRCA1 allele in inherited disease and to loss of heterozygosity in sporadic cancer. To identify genes required for suppression of Alu-mediated recombination we performed a genomewide screen of a collection of 4672 yeast gene deletion mutants using a direct repeat recombination assay. The primary screen and subsequent analysis identified 12 candidate genes including TSA, ELG1, and RRM3, which are known to play a significant role in maintaining genomic stability. Genetic analysis of the corresponding human homologs was performed in sporadic breast tumors and in inherited BRCA1-associated carcinomas. Sequencing of these genes in high risk breast cancer families revealed a potential role for the helicase PIF1 in cancer predisposition. PIF1 variant L319P was identified in three breast cancer families; importantly, this variant, which is predicted to be functionally damaging, was not identified in a large series of controls nor has it been reported in either dbSNP or the 1000 Genomes Project. In Schizosaccharomyces pombe, Pfh1 is required to maintain both mitochondrial and nuclear genomic integrity. Functional studies in yeast of human PIF1 L319P revealed that this variant cannot complement the essential functions of Pfh1 in either the nucleus or mitochondria. Our results provide a global view of nonessential genes involved in suppressing Alu-mediated recombination and implicate variation in PIF1 in breast cancer predisposition.

  3. RNA SURVEILLANCE– AN EMERGING ROLE FOR RNA REGULATORY NETWORKS IN AGING

    OpenAIRE

    Montano, Monty; Long, Kimberly

    2010-01-01

    In this review, we describe recent advances in the field of RNA regulatory biology and relate these advances to aging science. We introduce a new term, RNA surveillance, an RNA regulatory process that is conserved in metazoans, and describe how RNA surveillance represents molecular cross-talk between two emerging RNA regulatory systems – RNA interference and RNA editing. We discuss how RNA surveillance mechanisms influence mRNA and microRNA expression and activity during lifespan. Additionall...

  4. Novel Reversible TSG Gate and Its Application for Designing Components of Primitive Reversible/Quantum ALU

    OpenAIRE

    Thapliyal, Himanshu; Srinivas, M. B

    2006-01-01

    In recent years, reversible logic has emerged as a promising computing paradigm having application in low power CMOS, quantum computing, nanotechnology, and optical computing. The classical set of gates such as AND, OR, and EXOR are not reversible. This paper utilizes a new 4 * 4 reversible gate called TSG gate to build the components of a primitive reversible/quantum ALU. The most significant aspect of the TSG gate is that it can work singly as a reversible full adder, that is reversible ful...

  5. POMEN IN MERJENJE ORGANIZACIJSKE KULTURE. PRIMER PODJETJA ALPOS ALU D.O.O.

    OpenAIRE

    Senica, Damjana

    2013-01-01

    Diplomsko delo obravnava organizacijsko kulturo (OK), ki je splet vrednot, prepričanj, stališč in spoznanj o tem, kako organizacija deluje ali naj bi delovala. Ob teoretični obravnavi OK je bila v nalogi opravljena tudi analiza OK v podjetju Alpos Alu d.o.o., ki je zaradi splošne gospodarske krize v zapletenem položaju, zato je še posebej zanimivo za obravnavo. Z metodo OCAI avtorjev Camerona in Quinna sem prišla do ugotovitve, da v podjetju prevladuje kultura trga, zaposleni pa si žel...

  6. The potential role of Alu Y in the development of resistance to SN38 (Irinotecan) or oxaliplatin in colorectal cancer

    DEFF Research Database (Denmark)

    Lin, Xue; Stenvang, Jan; Rasmussen, Mads Heilskov;

    2015-01-01

    toxicity induced by carcinogens or drugs can reactivate Alus by altering DNA methylation. Whether or not reactivation of Alus occurs in SN38 and oxaliplatin resistance remains unknown. Results: We applied reduced representation bisulfite sequencing (RRBS) to investigate the DNA methylome in SN38 or...... oxaliplatin resistant colorectal cancer cell line models. Moreover, we extended the RRBS analysis to tumor tissue from 14 patients with colorectal cancer who either did or did not benefit from capecitabine + oxaliplatin treatment. For the clinical samples, we applied a concept of 'DNA methylation entropy' to......-especially the Alu Y subfamily. Furthermore, we identified an enrichment of Alu Y sequences that likely results from increased integration of new copies of Alu Y sequence in the drug-resistant cell lines. In the clinical samples, SOX1 and other SOX gene family members were shown to display variable DNA...

  7. An Alu element-associated hypermethylation variant of the POMC gene is associated with childhood obesity.

    Directory of Open Access Journals (Sweden)

    Peter Kuehnen

    Full Text Available The individual risk for common diseases not only depends on genetic but also on epigenetic polymorphisms. To assess the role of epigenetic variations in the individual risk for obesity, we have determined the methylation status of two CpG islands at the POMC locus in obese and normal-weight children. We found a hypermethylation variant targeting individual CpGs at the intron 2-exon 3 boundary of the POMC gene by bisulphite sequencing that was significantly associated with obesity. POMC exon 3 hypermethylation interferes with binding of the transcription enhancer P300 and reduces expression of the POMC transcript. Since intron 2 contains Alu elements that are known to influence methylation in their genomic vicinity, the exon 3 methylation variant seems to result from an Alu element-triggered default state of methylation boundary definition. Exon 3 hypermethylation in the POMC locus represents the first identified DNA methylation variant that is associated with the individual risk for obesity.

  8. The levels of edit, second edition

    Science.gov (United States)

    Vanburen, R.; Buehler, M. F.

    1980-01-01

    The editorial process is analyzed, and five levels of edit are identified. These levels represent cumulative combinations of nine types of edit: Coordination, Policy, Integrity, Screening, Copy Clarification, Format, Mechanical Style, Language, and Substantive. The levels and types of edit, although developed for specific use with external reports at the Jet Propulsion Laboratory, cover the general range of technical editing, especially as it applies to an in-house technical publications organization. Each type of edit is set forth in terms of groups of actions to be performed by editor. The edit-level concept has enhanced understanding and communication among editors, authors, and publications managers concerning the specific editorial work to be done on each manuscript. It has also proved useful as a management tool for estimating and monitoring cost.

  9. Wikipedia editing dynamics

    Science.gov (United States)

    Gandica, Y.; Carvalho, J.; Sampaio dos Aidos, F.

    2015-01-01

    A model for the probabilistic function followed in editing Wikipedia is presented and compared with simulations and real data. It is argued that the probability of editing is proportional to the editor's number of previous edits (preferential attachment), to the editor's fitness, and to an aging factor. Using these simple ingredients, it is possible to reproduce the results obtained for Wikipedia editing dynamics for a collection of single pages as well as the averaged results. Using a stochastic process framework, a recursive equation was obtained for the average of the number of edits per editor that seems to describe the editing behavior in Wikipedia.

  10. Wikipedia edition dynamics

    CERN Document Server

    Gandica, Y; Carvalho, J

    2014-01-01

    A model for the probabilistic function followed in Wikipedia edition is presented and compared with simulations and real data. It is argued that the probability to edit is proportional to the editor's number of previous editions (preferential attachment), to the editor's fitness and to an ageing factor. Using these simple ingredients, it is possible to reproduce the results obtained for Wikipedia edition dynamics for a collection of single pages as well as the averaged results. Using a stochastic process framework, a recursive equation was obtained for the average of the number of editions per editor that seems to describe the editing behaviour in Wikipedia.

  11. Edit wars in Wikipedia

    OpenAIRE

    Sumi, Róbert; Yasseri, Taha; Rung, András; Kornai, András; Kertész, János

    2011-01-01

    We present a new, efficient method for automatically detecting severe conflicts `edit wars' in Wikipedia and evaluate this method on six different language WPs. We discuss how the number of edits, reverts, the length of discussions, the burstiness of edits and reverts deviate in such pages from those following the general workflow, and argue that earlier work has significantly over-estimated the contentiousness of the Wikipedia editing process.

  12. DNA-free genome editing methods for targeted crop improvement.

    Science.gov (United States)

    Kanchiswamy, Chidananda Nagamangala

    2016-07-01

    Evolution of the next-generation clustered, regularly interspaced, short palindromic repeat/Cas9 (CRISPR/Cas9) genome editing tools, ribonucleoprotein (RNA)-guided endonuclease (RGEN) RNPs, is paving the way for developing DNA-free genetically edited crop plants. In this review, I discuss the various methods of RGEN RNPs tool delivery into plant cells and their limitations to adopt this technology to numerous crop plants. Furthermore, focus is given on the importance of developing DNA-free genome edited crop plants, including perennial crop plants. The possible regulation on the DNA-free, next-generation genome-edited crop plants is also highlighted. PMID:27100964

  13. Editing efficiency of a Drosophila gene correlates with a distant splice site selection

    OpenAIRE

    AGRAWAL, RITESH; Stormo, Gary D.

    2005-01-01

    RNA editing and alternative splicing are two processes that increase protein diversity. The relationship between the two processes is not well understood. There are a few examples of correlations between editing and alternative splicing, but these are all nearby effects. A search for alternative splicing among 16 edited genes in Drosophila reveals two novel instances of alternative splicing. In one example where alternative splicing occurs downstream of editing, a strong correlation between e...

  14. Alignments of RNA structures.

    Science.gov (United States)

    Blin, Guillaume; Denise, Alain; Dulucq, Serge; Herrbach, Claire; Touzet, Hélène

    2010-01-01

    We describe a theoretical unifying framework to express the comparison of RNA structures, which we call alignment hierarchy. This framework relies on the definition of common supersequences for arc-annotated sequences and encompasses the main existing models for RNA structure comparison based on trees and arc-annotated sequences with a variety of edit operations. It also gives rise to edit models that have not been studied yet. We provide a thorough analysis of the alignment hierarchy, including a new polynomial-time algorithm and an NP-completeness proof. The polynomial-time algorithm involves biologically relevant edit operations such as pairing or unpairing nucleotides. It has been implemented in a software, called gardenia, which is available at the Web server http://bioinfo.lifl.fr/RNA/gardenia. PMID:20431150

  15. CRISPR-assisted editing of bacterial genomes

    OpenAIRE

    Jiang, Wenyan; Bikard, David; Cox, David; Zhang, Feng; Marraffini, Luciano A.

    2013-01-01

    The targeting of nucleases to specific DNA sequences facilitates genome editing. Recent work demonstrated that the CRISPR-associated (Cas) nuclease Cas9 can be targeted to sequences in vitro simply by modifying a short7 CRISPR RNA (crRNA) guide. Here we use this CRISPR-Cas system to introduce marker-free mutations in Streptococcus pneumoniae and Escherichia coli. The approach involves re-programming Cas9 by using a crRNA complementary to a target chromosomal locus and introducing a template D...

  16. Design and Implementation of Submicron Level 10T Full Adder in ALU Using Cell Based and SOC Technology

    Directory of Open Access Journals (Sweden)

    K.Swathi

    2014-09-01

    Full Text Available As technology scales into the nanometer regime leakage current, active power, delay and area are becoming important metric for the analysis and design of complex circuits. The main concern in mobile and battery based systems are leakage current and power dissipation. A transistor resizing approach for 10 transistor single bit full adder cells is used to determine optimal sleep transistor size which reduces power dissipation and leakage current. A submicron level 10-transistor single bit full adder cell is considered to achieve low leakage current, reduced power dissipation and high speed. In this paper initially 10T full adder cell is designed with submicron technique and later this is employed to design an ALU adder unit. The modified ALU is simulated and synthesized successfully on cadence 180nm technology.

  17. Analysis of the action of the restriction endonuclease AluI using three different comet assay protocols

    International Nuclear Information System (INIS)

    Background and purpose: the comet assay offers the opportunity to measure the amount of DNA damage and the effectiveness of DNA repair in single cells. In a first part, experiments are presented comparing three different protocols of the comet assay technique with respect to the analysis of the induction of DNA damage after X-irradiation in isolated human lymphocytes and CHO cells. In a second part, the restriction enzyme AluI, an agent producing DNA double-strand breaks exclusively, was introduced into CHO cells by electroporation and the effects were analyzed using the different comet assay protocols. The experiments were carried out in order to test the assertion that comet assay techniques can measure different types of DNA damages at different pH conditions of lysis and electrophoresis. Material and methods: three different comet assay protocols were used for the analysis of DNA damage in lymphocytes and CHO cells. Results: the results clearly indicate that among the three protocols the modified comet assay technique used by the authors showed the highest sensitivity in the radiotherapy-relevant dose range between 0 and 2 Gy. All three protocols were capable of detecting an effect by AluI. This effect, however, was clearly different from radiation effects. Whereas after radiation exposure all cell nuclei show a dose-dependent increase in DNA content in the comet tail, most of the cell nuclei were unaffected by an AluI uptake. Nevertheless, there was an effect by AluI that could be detected in all three assay versions: between 5% and 15% of the nuclei showed clearly abnormal comet morphologies. Conclusion: neither the strictly alkaline nor the strictly neutral comet assay is applicable in the radiation dose range of about 2 Gy. The restriction enzyme results show that other factors than just DNA strand breaks contribute to DNA migration into the tail of the comets. (orig.)

  18. THE UTILIZATION OF FOREST RESOURCES IN PROTECTED FOREST IN ALU VILLAGE, POLEWALI MANDAR DISTRINC, PROVINCE OF WEST SULAWESI

    OpenAIRE

    Asriyanni; Asrianty

    2011-01-01

    This study aims to determine the activity of the utilization of forest resources and to know local wisdom of community in protected forest in the Alu Village areas. The results of this study are expected to be taken into consideration for policy makers in an effort to empower communities around the forests in sustainable forest management. This research is conducted with method of Rapid Rural Appraisal (RRA). Respondents of this study are communities which is leaving in around the protect...

  19. Alu Sb2 subfamily is present in all higher primates but was most succesfully amplified in humans

    Energy Technology Data Exchange (ETDEWEB)

    Richer, C.; Zietkiewicz, E.; Labuda, D. [Universite de Montreal, Que (Canada)

    1994-09-01

    Alu repeats can be classified into subfamilies which amplified in primate genomes at different evolutionary time periods. A young Alu subfamily, Sb2, with a characteristic 7-nucleotide duplication at position 256, has been described in seven human loci. An Sb2 insertion found near the HD gene was unique to two HD families, indicating that Sb2 was still retropositionally active. Here, we have shown that the Sb2 insertion in the CHOL locus was similarly rare, being absent in 120 individuals of Caucasian, Oriental and Black origin. In contrast, Sb2 inserts in five other loci were found fixed (non-polymorphic), based on measurements in the same population sample, but absent from orthologous positions in higher apes. This suggest that Sb2 repeats spread relatively early in the human lineage following divergence from other primates and that these elements may be human-specific. By quantitative PCR, we investigated the presence of Sb2 sequences in different primate DNA, using one PCR primer anchored at the 5{prime} Alu-end and the other complementary to the duplicated Sb2-specific segment. With an Sb2-containing plasmid as a standard, we estimated the number of Sb2 repeats at 1500-1800 copies per human haploid equivalent; corresponding numbers in chimpanzee and gorilla were almost two orders of magnitude lower, while the signal observed in orangutan and gibbon DNAs was consistent with the presence of a single copy. The analysis of 22 human, 11 chimpanzee and 10 gorilla sequences indicates that the Alu Sb2 dispersed independently in these three primate lineages; gorilla consensus differs from the human Sb2 sequence by one position, while all chimpanzee repeats have their linker expanded by up to eight A-residues. Should they be thus considered as separate subfamilies? It is possible that sequence modifications with respect to the human consensus are responsible for poor retroposition of Sb2 in apes.

  20. Translational arrest by a prokaryotic signal recognition particle is mediated by RNA interactions.

    Science.gov (United States)

    Beckert, Bertrand; Kedrov, Alexej; Sohmen, Daniel; Kempf, Georg; Wild, Klemens; Sinning, Irmgard; Stahlberg, Henning; Wilson, Daniel N; Beckmann, Roland

    2015-10-01

    The signal recognition particle (SRP) recognizes signal sequences of nascent polypeptides and targets ribosome-nascent chain complexes to membrane translocation sites. In eukaryotes, translating ribosomes are slowed down by the Alu domain of SRP to allow efficient targeting. In prokaryotes, however, little is known about the structure and function of Alu domain-containing SRPs. Here, we report a complete molecular model of SRP from the Gram-positive bacterium Bacillus subtilis, based on cryo-EM. The SRP comprises two subunits, 6S RNA and SRP54 or Ffh, and it facilitates elongation slowdown similarly to its eukaryotic counterpart. However, protein contacts with the small ribosomal subunit observed for the mammalian Alu domain are substituted in bacteria by RNA-RNA interactions of 6S RNA with the α-sarcin-ricin loop and helices H43 and H44 of 23S rRNA. Our findings provide a structural basis for cotranslational targeting and RNA-driven elongation arrest in prokaryotes. PMID:26344568

  1. Recent Advances in Genome Editing Using CRISPR/Cas9

    OpenAIRE

    Ding, Yuduan; Li, Hong; Chen, Ling-Ling; Xie, Kabin

    2016-01-01

    The CRISPR (clustered regularly interspaced short palindromic repeat)-Cas9 (CRISPR-associated nuclease 9) system is a versatile tool for genome engineering that uses a guide RNA (gRNA) to target Cas9 to a specific sequence. This simple RNA-guided genome-editing technology has become a revolutionary tool in biology and has many innovative applications in different fields. In this review, we briefly introduce the Cas9-mediated genome-editing method, summarize the recent advances in CRISPR/Cas9 ...

  2. Questioning the "melting pot": analysis of Alu inserts in three population samples from Uruguay.

    Science.gov (United States)

    Hidalgo, Pedro C; Mut, Patricia; Ackermann, Elizabeth; Figueiro, Gonzalo; Sans, Monica

    2014-01-01

    The way that immigrants integrate into recipient societies has been discussed for decades, mainly from the perspective of the social sciences. Uruguay, as other American countries, received diffferent waves of European immigrants, although the details of the process of assimilation, when it did occur, are unclear. In this study we used genetic markers to understand the process experienced by the Basques, one of the major migration waves that populated Uruguay, and their relation to other immigrants, as well as to Native American and African descendants. For this purpose, we analyzed the allele frequencies of 10 ALU loci (A25, ACE, APOA1, B65, D1, F13B, PV92, TPA25, HS2.43, and HS4.65) in three samples from Uruguay (two of Basque descendants, one of non-Basque descendants) from two locations: Montevideo and Trinidad. No departure from Hardy-Weinberg expectations was observed, with the exceptions of the APOA1 and D1 loci in the non-Basque descendants' samples. Our data show that the major genetic contribution in the three samples comes from Europe (78-88%), with minor African (10-15%) and Native American (0-10%) contributions. Genetic distances reveal that Basque descendants from Trinidad cluster with Europeans, whereas both Montevideo samples cluster together and are separate from other populations, showing two diffferent types of integration, related to the general characteristics of each regional population. PMID:25397699

  3. Quality text editing

    Directory of Open Access Journals (Sweden)

    Gyöngyi Bujdosó

    2009-10-01

    Full Text Available Text editing is more than the knowledge of word processing techniques. Originally typographers, printers, text editors were the ones qualified to edit texts, which were well structured, legible, easily understandable, clear, and were able to emphasize the coreof the text. Time has changed, and nowadays everyone has access to computers as well as to text editing software and most users believe that having these tools is enough to edit texts. However, text editing requires more skills. Texts appearing either in printed or inelectronic form reveal that most of the users do not realize that they are not qualified to edit and publish their works. Analyzing the ‘text-products’ of the last decade a tendency can clearly be drawn. More and more documents appear, which instead of emphasizingthe subject matter, are lost in the maze of unstructured text slices. Without further thoughts different font types, colors, sizes, strange arrangements of objects, etc. are applied. We present examples with the most common typographic and text editing errors. Our aim is to call the attention to these mistakes and persuadeusers to spend time to educate themselves in text editing. They have to realize that a well-structured text is able to strengthen the effect on the reader, thus the original message will reach the target group.

  4. RNA-DNA Differences Are Generated in Human Cells within Seconds After RNA Exits Pol II

    OpenAIRE

    Isabel X. Wang; Leighton J. Core; Hojoong Kwak; Lauren Brady; Alan Bruzel; Lee McDaniel; Allison L. Richards; Ming Wu; Christopher Grunseich; John T. Lis; Vivian G. Cheung

    2014-01-01

    RNA sequences are expected to be identical to their corresponding DNA sequences. Here, we found all 12 types of RNA-DNA sequence differences (RDDs) in nascent RNA. Our results show that RDDs begin to occur in RNA chains about 55 nucleotides from the RNA polymerase II (Pol II) active site. These RDDs occur so soon after transcription that they are incompatible with known deaminase-mediated RNA editing mechanisms. Moreover, the 55-nucleotide delay in appearance indicates they do not arise durin...

  5. CRISPR Genome Editing

    Science.gov (United States)

    A research article about a technique for gene editing known as CRISPR-Cas9. The technique has made it much easier and faster for cancer researchers to study mutations and test new therapeutic targets.

  6. Identification of an Alu-repeat-mediated deletion of OPTN upstream region in a patient with a complex ocular phenotype.

    Science.gov (United States)

    Schilter, Kala F; Reis, Linda M; Sorokina, Elena A; Semina, Elena V

    2015-11-01

    Genetic causes of ocular conditions remain largely unknown. To reveal the molecular basis for a congenital ocular phenotype associated with glaucoma we performed whole-exome sequencing (WES) and whole-genome copy number analyses of patient DNA. WES did not identify a causative variant. Copy number variation analysis identified a deletion of 10p13 in the patient and his unaffected father; the deletion breakpoint contained a single 37-bp sequence that is normally present in two distinct Alu repeats separated by ~181 kb. The deletion removed part of the upstream region of optineurin (OPTN) as well as the upstream sequence and two coding exons of coiled-coil domain containing 3 (CCDC3); analysis of the patient's second allele showed normal OPTN and CCDC3 sequences. Studies of zebrafish orthologs identified expression in the developing eye for both genes. OPTN is a known factor in dominant adult-onset glaucoma and Amyotrophic Lateral Sclerosis (ALS). The deletion eliminates 98 kb of the OPTN upstream sequence leaving only ~1 kb of the proximal promoter region. Comparison of transcriptional activation capability of the 3 kb normal and the rearranged del(10)(p13) OPTN promoter sequences demonstrated a statistically significant decrease for the deleted allele; sequence analysis of the entire deleted region identified multiple conserved elements with possible cis-regulatory activity. Additional screening of CCDC3 indicated that heterozygous loss-of-function alleles are unlikely to cause congenital ocular disease. In summary, we report the first regulatory region deletion involving OPTN, caused by Alu-mediated nonallelic homologous recombination and possibly contributing to the patient's ocular phenotype. In addition, our data indicate that Alu-mediated rearrangements of the OPTN upstream region may represent a new source of affected alleles in human conditions. Evaluation of the upstream OPTN sequences in additional ocular and ALS patients may help to determine the role

  7. Developing new levels of edit

    Energy Technology Data Exchange (ETDEWEB)

    Prono, J.K.

    1997-06-01

    Since 1985, Los Alamos National Laboratory (LANL) staff have had four levels of edit to choose from for technical reports. When a CQI survey showed that both authors and editors felt the levels were not meeting author needs, LANL set about revising them. The goals were to simplify the editing process, focus editing on improving technical clarity, and ensure value added in editing. This paper describes the revision process and product--three author-based levels of edit.

  8. Replication of cloned DNA containing the Alu family sequence during cell extract-promoting simian virus 40 DNA synthesis.

    OpenAIRE

    Ariga, H

    1984-01-01

    The replicating activity of several cloned DNAs containing putative origin sequences was examined in a cell-free extract that absolutely depends on simian virus 40 (SV40) T antigen promoting initiation of SV40 DNA replication in vitro. Of the three DNAs containing the human Alu family sequence (BLUR8), the origin of (Saccharomyces cerevisiae plasmid 2 micron DNA (pJD29), and the yeast autonomous replicating sequence (YRp7), only BLUR8 was active as a template. Replication in a reaction mixtur...

  9. International distribution and age estimation of the Portuguese BRCA2 c.156_157insAlu founder mutation

    DEFF Research Database (Denmark)

    Peixoto, Ana; Santos, Catarina; Pinheiro, Manuela;

    2011-01-01

    total of 5,443 suspected HBOC families from several countries. Whereas the c.156_157insAlu BRCA2 mutation was detected in 11 of 149 suspected HBOC families from Portugal, representing 37.9% of all deleterious mutations, in other countries it was detected only in one proband living in France and in four...... individuals requesting predictive testing living in France and in the USA, all being Portuguese immigrants. After performing an extensive haplotype study in carrier families, we estimate that this founder mutation occurred 558 +/- 215 years ago. We further demonstrate significant quantitative differences...

  10. International distribution and age estimation of the Portuguese BRCA2 c.156_157insAlu founder mutation

    DEFF Research Database (Denmark)

    Peixoto, Ana; Santos, Catarina; Pinheiro, Manuela;

    2011-01-01

    total of 5,443 suspected HBOC families from several countries. Whereas the c.156_157insAlu BRCA2 mutation was detected in 11 of 149 suspected HBOC families from Portugal, representing 37.9% of all deleterious mutations, in other countries it was detected only in one proband living in France and in four...... individuals requesting predictive testing living in France and in the USA, all being Portuguese immigrants. After performing an extensive haplotype study in carrier families, we estimate that this founder mutation occurred 558 ± 215 years ago. We further demonstrate significant quantitative differences...

  11. Editing of mouse and human immunoglobulin genes by CRISPR-Cas9 system

    OpenAIRE

    Cheong, Taek-Chin; Compagno, Mara; Chiarle, Roberto

    2016-01-01

    Applications of the CRISPR-Cas9 system to edit the genome have widely expanded to include DNA gene knock-out, deletions, chromosomal rearrangements, RNA editing and genome-wide screenings. Here we show the application of CRISPR-Cas9 technology to edit the mouse and human immunoglobulin (Ig) genes. By delivering Cas9 and guide-RNA (gRNA) with retro- or lenti-virus to IgM+ mouse B cells and hybridomas, we induce class-switch recombination (CSR) of the IgH chain to the desired subclass. Similarl...

  12. Semiautomated improvement of RNA alignments

    DEFF Research Database (Denmark)

    Andersen, Ebbe Sloth; Lind-Thomsen, Allan; Knudsen, Bjarne;

    2007-01-01

    We have developed a semiautomated RNA sequence editor (SARSE) that integrates tools for analyzing RNA alignments. The editor highlights different properties of the alignment by color, and its integrated analysis tools prevent the introduction of errors when doing alignment editing. SARSE readily...... the SARSE editor makes it a flexible tool to improve all RNA alignments with relatively little human intervention. Online documentation and software are available at (http://sarse.ku.dk)....

  13. Sequences more than 500 base pairs upstream of the human U3 small nuclear RNA gene stimulate the synthesis of U3 RNA in frog oocytes

    International Nuclear Information System (INIS)

    Small nuclear RNA (snRNA) genes contain strong promoters capable of initiating transcription once every 4 s. Studies on the human U1 snRNA gene, carried out in other laboratories, showed that sequences within 400 bp of the 5' flanking region are sufficient for maximal levels of transcription both in vivo and in frog oocytes [reviewed in Dahlberg and Lund (1988)]. The authors studied the expression of a human U3 snRNA gene by injecting 5' deletion mutants into frog oocytes. The results show that sequences more than 500 bp upstream of the U3 snRNA gene have a 2-3-fold stimulatory effect on the U3 snRNA synthesis. These results indicate that the human U3 snRNA gene is different from human U1 snRNA gene in containing regulatory elements more than 500 bp upstream. The U3 snRNA gene upstream sequences contain an AluI homologous sequence in the -1,200 region; these AluI sequences were transcribed in vitro and in frog oocytes but were not detectable in Hela cells

  14. Alu-alu recombination results in a duplication of seven exons in the lysyl hydroxylase gene in a patient with the type VI variant of Ethlers-Danlos syndrome

    Energy Technology Data Exchange (ETDEWEB)

    Pousi, B.; Hautala, T.; Heikkinen, J.; Pajunen, L.; Kivirikko, K.I.; Myllylae, R. [Univ. of Oulu (Finland)

    1994-11-01

    The type VI variant of the Ethlers-Danlos syndrome (EDS) is a recessively inherited connective-tissue disorder. The characteristic features of the variant are muscular hyptonia, kyphoscoliosis, ocular manifestations, joint hypermobility, skin fragility and hyperextensibility, and other signs of connective-tissue involvement. The biochemical defect in most but not all patients is a deficiency in lysyl hydroxylase activity. Lysyl hydroxylase is an enzyme that catalyzes the formation of hydroxylysine in collagens and other proteins with collagen-like amino acid sequences. We have recently reported an apparently homozygous large-duplication rearrangement in the gene for lysyl hydroxylase, leading to the type VI variant of EDS in two siblings. We now report an identical, apparently homozygous large duplication in an unrelated 49-year-old female originally analyzed by Sussman et al. Our simple-sequence-repeat-polymorphism analysis does not support uniparental isodisomy inheritance for either of the two duplications. Furthermore, we indicate in this study that the duplication in the lysyl hydroxylase gene is caused by an Alu-Alu recombination in both families. Cloning of the junction fragment of the duplication has allowed synthesis of appropriate primers for rapid screening for this rearrangement in other families with the type VI variant of EDS. 38 refs., 6 figs.

  15. The editing sites in transcripts of functional genes of rice mitochondria

    Institute of Scientific and Technical Information of China (English)

    2002-01-01

    RNA editing exists extensively in the higher plant mitochondria, and is a required step for forming functional proteins. There may be some relationship between RNA editing and cytoplasmic male sterility (CMS), a kind of phenomenon that is attributed to mitochondrial genome mutations. The research materials used are the gametophytic male sterility line (A), maintainer line (B) and F1 hybrid (F1) of HL-type CMS rice. cDNAs and DNAs of atp6 and coxII have been obtained from A, B and F1 by PCR and RT-PCR. Comparing sequences of cDNAs and DNAs, 18 and 15 editing sites were found respectively in the transcripts of atp6 and coxII. A, B and F1 shared the same editing sites. RNA editing improves hydrophobicity and conservation of the predicted protein as compared with other organisms.

  16. Itella logistiikan yrityskaupan yhteydessä ilmenneet päällekkäisyydet ja ongelmat Oulusta ajettavissa alue- ja runkokuljetuksissa

    OpenAIRE

    Niemelä, Hannes

    2014-01-01

    Työn aiheena on Itella Logistics Oyj:n ja VR Transtpointin yhdistymisestä aiheutuneiden ongelmien ja päällekkäisyyksien selvittäminen Oulusta ajettaviin alue- ja runkovuoroihin. Työn tavoitteena oli selvittää syntyneet ongelmat ja niihin jo tehdyt ratkaisut sekä miettiä mahdollisia ratkaisuehdotuksia selvittämättömiin ongelmiin. Työssä esitellään myös yksityiskohtaisesti Itella Logistiikan Oulusta ajettavat alue- ja runkovuorot. Työ suoritettiin teoreettisen tutkimuksen ja pääasiassa työss...

  17. Structural organization of glycophorin A and B genes: Glycophorin B gene evolved by homologous recombination at Alu repeat sequences

    International Nuclear Information System (INIS)

    Glycophorins A (GPA) and B (GPB) are two major sialoglycoproteins of the human erythrocyte membrane. Here the authors present a comparison of the genomic structures of GPA and GPB developed by analyzing DNA clones isolated from a K562 genomic library. Nucleotide sequences of exon-intron junctions and 5' and 3' flanking sequences revealed that the GPA and GPB genes consist of 7 and 5 exons, respectively, and both genes have >95% identical sequence from the 5' flanking region to the region ∼ 1 kilobase downstream from the exon encoding the transmembrane regions. In this homologous part of the genes, GPB lacks one exon due to a point mutation at the 5' splicing site of the third intron, which inactivates the 5' cleavage event of splicing and leads to ligation of the second to the fourth exon. Following these very homologous sequences, the genomic sequences for GPA and GPB diverge significantly and no homology can be detected in their 3' end sequences. The analysis of the Alu sequences and their flanking direct repeat sequences suggest that an ancestral genomic structure has been maintained in the GPA gene, whereas the GPB gene has arisen from the acquisition of 3' sequences different from those of the GPA gene by homologous recombination at the Alu repeats during or after gene duplication

  18. CRISPR-Cas9-Mediated Genome Editing in Leishmania donovani

    OpenAIRE

    Zhang, Wen-Wei; Matlashewski, Greg

    2015-01-01

    ABSTRACT The prokaryotic CRISPR (clustered regularly interspaced short palindromic repeat)-Cas9, an RNA-guided endonuclease, has been shown to mediate efficient genome editing in a wide variety of organisms. In the present study, the CRISPR-Cas9 system has been adapted to Leishmania donovani, a protozoan parasite that causes fatal human visceral leishmaniasis. We introduced the Cas9 nuclease into L. donovani and generated guide RNA (gRNA) expression vectors by using the L. donovani rRNA promo...

  19. Precision genome editing

    DEFF Research Database (Denmark)

    Steentoft, Catharina; Bennett, Eric P; Schjoldager, Katrine Ter-Borch Gram;

    2014-01-01

    of glycobiology, primarily due to their low efficiencies, with resultant failure to impose substantial phenotypic consequences upon the final glycosylation products. Here, we review novel nuclease-based precision genome editing techniques enabling efficient and stable gene editing, including gene disruption...... by introducing single or double-stranded breaks at a defined genomic sequence. We here compare and contrast the different techniques and summarize their current applications, highlighting cases from the field of glycobiology as well as pointing to future opportunities. The emerging potential of precision gene...

  20. Edit Distance for Timed Automata

    OpenAIRE

    Chatterjee, Krishnendu; Ibsen-Jensen, Rasmus; Majumdar, Rupak

    2013-01-01

    The edit distance between two (untimed) traces is the minimum cost of a sequence of edit operations (insertion, deletion, or substitution) needed to transform one trace to the other. Edit distances have been extensively studied in the untimed setting, and form the basis for approximate matching of sequences in different domains such as coding theory, parsing, and speech recognition. In this paper, we lift the study of edit distances from untimed languages to the timed setting. We define ...

  1. Simple Genome Editing of Rodent Intact Embryos by Electroporation.

    Directory of Open Access Journals (Sweden)

    Takehito Kaneko

    Full Text Available The clustered regularly interspaced short palindromic repeat (CRISPR/CRISPR-associated (Cas system is a powerful tool for genome editing in animals. Recently, new technology has been developed to genetically modify animals without using highly skilled techniques, such as pronuclear microinjection of endonucleases. Technique for animal knockout system by electroporation (TAKE method is a simple and effective technology that produces knockout rats by introducing endonuclease mRNAs into intact embryos using electroporation. Using TAKE method and CRISPR/Cas system, the present study successfully produced knockout and knock-in mice and rats. The mice and rats derived from embryos electroporated with Cas9 mRNA, gRNA and single-stranded oligodeoxynucleotide (ssODN comprised the edited targeted gene as a knockout (67% of mice and 88% of rats or knock-in (both 33%. The TAKE method could be widely used as a powerful tool to produce genetically modified animals by genome editing.

  2. Still-image frame grabs and benthic habitat interpretation of underwater video footage, March 2014, Faga`alu Bay, American Samoa

    Data.gov (United States)

    U.S. Geological Survey, Department of the Interior — Underwater video was collected in March 2014 in the nearshore waters of Faga`alu Bay on the island of Tutuila, American Samoa, as part of the U.S. Geological Survey...

  3. Effective Alu Repeat Based RT-Qpcr Normalization in Cancer Cell Perturbation Experiments

    OpenAIRE

    Ali Rihani; Tom Van Maerken; Filip Pattyn; Gert Van Peer; Anneleen Beckers; Sara De Brouwer; Candy Kumps; Evelien Mets; Joni Van der Meulen; Pieter Rondou; Carina Leonelli; Pieter Mestdagh; Frank Speleman; Jo Vandesompele

    2013-01-01

    BACKGROUND: Measuring messenger RNA (mRNA) levels using the reverse transcription quantitative polymerase chain reaction (RT-qPCR) is common practice in many laboratories. A specific set of mRNAs as internal control reference genes is considered as the preferred strategy to normalize RT-qPCR data. Proper selection of reference genes is a critical issue, especially in cancer cells that are subjected to different in vitro manipulations. These manipulations may result in dramatic alterations in ...

  4. Editing graphs for maximum effect

    Energy Technology Data Exchange (ETDEWEB)

    Murphy, P.W.; Rhiner, R.W.

    1991-01-08

    The paper contains over eighty rules for editing graphs, arranged under nine major headings in a logical sequence for editing all the graphs in a manuscript. It is excerpted from a monograph used at the Lawrence Livermore National Laboratory to train beginning technical editors in editing graphs; a corresponding Hypercard stack is also used in this training. 6 refs., 4 figs.

  5. Beginning to edit physics

    Energy Technology Data Exchange (ETDEWEB)

    Murphy, P.W.

    1995-02-01

    A physicist-turned-editor shows you the basics required for copyediting physics papers (physical quantities, symbols, units, scientific notation, the structure of mathematical expressions, the nature of graphs), and points the way to learning enough ``editorial physics`` to begin substantive editing.

  6. Nair handbook. 1995 edition

    International Nuclear Information System (INIS)

    This Handbook contains general background, administrative and technical information for those participating in the National Arrangements for Incidents involving Radioactivity (NAIR), updating and replacing the previous edition published in 1987. The overriding need for revision was brought about as a result of changes introduced by British Telecom to the telephone numbers of establishments. (UK)

  7. ISBD Consolidated Edition

    CERN Document Server

    Section, Standing Committee of the IFLA Cataloguing

    2011-01-01

    This is the new edition of the first consolidated ISBD that was published in 2007. The first years of usage have led to interesting and useful corrections and additions. Many cataloguers and practitioners worldwide will welcome this updated first class tool, which is useful and applicable for descriptions of bibliographic resources in any type of catalogue.

  8. The Craft of Editing

    DEFF Research Database (Denmark)

    Moeran, Brian

    To edit is to make a choice, or series of choices. Will I write a rough draft of this essay in longhand, or hammer it out on my computer? If the latter, what font shall I use? Times New Roman, Book Antiqua, or Garamond? Once I get started, what style shall I adopt: realistic, confessional or...

  9. Preface to Special Edition

    OpenAIRE

    Renee Nathanson

    2012-01-01

    Given that reading comprehension is at the forefront of global literacy discourse, this special edition of Per Linguam, the first number that is also published online, features a collection of articles that cover different aspects of reading comprehension and instruction, such as, strategies for comprehending texts, metacognitive awareness, the reciprocity of assessment and comprehension instruction and socio-affective factors that influence comprehension.

  10. Tokamaks (Second Edition)

    International Nuclear Information System (INIS)

    The first edition of John Wesson's book on tokamaks, published in 1987, established itself as essential reading for researchers in the field of magnetic confinement fusion: it was an excellent introduction for students to tokamak physics and also a valuable reference work for the more experienced. The second edition, published in 1997, has been completely rewritten and substantially enlarged (680 pages compared with 300). The new edition maintains the aim of providing a simple introduction to basic tokamak physics, but also includes discussion of the substantial advances in fusion research during the past decade. The new book, like its predecessor, is well written and commendable for its clarity and accuracy. In fact many of the chapters are written by a series of co-authors bringing the benefits of a wide range of expertise but, by careful editing, Wesson has maintained a uniformity of style and presentation. The chapter headings and coverage for the most part remain the same - but are expanded considerably and brought up to date. The most substantial change is that the single concluding chapter in the first edition on 'Experiments' has been replaced by three chapters: 'Tokamak experiments' which deals with some of the earlier key experiments plus a selection of recent small and medium-sized devices, 'Large experiments' which gives an excellent summary of the main results from the four large tokamaks - TFTR, JET, JT60/JT60U and DIII-D, and 'The future' which gives a very short (possibly too short in my opinion) account of reactors and ITER. This is an excellent book, which I strongly recommend should have a place - on the desk rather than in the bookshelf - of researchers in magnetic confinement fusion. (book review)

  11. Edit Distance for Pushdown Automata

    OpenAIRE

    Chatterjee, Krishnendu; Henzinger, Thomas A.; Ibsen-Jensen, Rasmus; Otop, Jan

    2015-01-01

    The edit distance between two words $w_1, w_2$ is the minimal number of word operations (letter insertions, deletions, and substitutions) necessary to transform $w_1$ to $w_2$. The edit distance generalizes to languages ${\\cal L}_1, {\\cal L}_2$, where the edit distance is the minimal number $k$ such that for every word from ${\\cal L}_1$ there exists a word in ${\\cal L}_2$ with edit distance at most $k$. We study the edit distance computation problem between pushdown automata and their subclas...

  12. Il mondo finisce a Rio: «Demônios» di Aluísio Azevedo

    Directory of Open Access Journals (Sweden)

    Giorgio De Marchis

    2013-07-01

    Full Text Available This essay analyses Demônios, a short story by the Brazilian writer Aluísio Azevedo published in 1893 in a collection bearing the same title. The text – unrelated to the naturalistic production of the author – describes a catastrophic nightmare, a carioca apocalypse reverting the evolutionary process and affecting the only two survivors, whose lives revert until they dissolve themselves in an aerial entity floating in the space. While trying to fit such an extravagant work within the production of its author, I will also explore the connection between the end of the world narrated by Azevedo as a positivist and a republican and the events involving the Brazilian republic in the early 1890s.

  13. Design and Analysis of a High Speed, Power Efficient 8 Bit ALU Based on SOI / SON MOSFET Technology

    Directory of Open Access Journals (Sweden)

    Subhramita Basak

    2014-01-01

    Full Text Available This paper shows an overall performance comparative analysis in terms of Average Power Consumption, Average Delay and Power-Delay Product for an 8 bit Arithmetic Logic Unit (ALU using bulk MOS, Silicon-on-Insulator (SOI and Silicon-on-Nothing (SON technology. The entire design is done in 32nm technology for all the three cases (Bulk, SOI & SON and then compared. The comparisons have been carried out with the help of the simulation runs on Synopsys HSpice tool, and that clearly indicates, for lower Supply Voltages (Vdd, SOI / SON technology provides a significant reduction in Average Power Consumption, Average Delay and Power-Delay Product compared to that of Bulk MOS technology.

  14. 苯乙酸抑制胰腺癌BXPC-3细胞增殖及RNA编辑酶表达的研究%Effects of phenylacetate on proliferation and RNA editing deaminase expression of human pancreatic carcinoma BXPC-3 cell lines

    Institute of Scientific and Technical Information of China (English)

    姜涛; 王岩; 任辉; 张广; 刘安安; 田宇

    2010-01-01

    Objective To evaluate the differentiation-inducing effect of phenylacetate on pancreatic carcinoma cells and underlying mechanism of RNA editing deaminase in cell proliferation and differentiation. Methods The effect of phenylacetate on cell proliferation and cell cycle were investigated in cultured pancreatic carcinoma BXPC-3 cell lines by methylthiazoletetrazolium( MTT) assay, and flow cytometry. The effect of phenylacetate on the expression of RNA editing deaminase (ADAR2 mRNA) in BXPC-3 cells and fresh pancreatic carcinoma specimen was evaluated by RT-PCR. Results ADAR1 mRNA expression was positive in human pancreatic carcinoma tissues and BXPC-3 cell lines. After application of 1.0 and 2.0 mmol/L phenylacetate for 24 h and 72 h, BXPC-3 cell growth inhibition rate increased, G0/G1 phase cells decreased, S phase cells increased, ADAR2 mRNA expression decreased ( P < 0.01 ). Conclusion ADAR2 mRNA played a vital role of regulation in the process of pancreatic carcinoma cells growth and differentiation. Phenylacetate could regulate the proliferation and differentiation of pancreatic carcinoma cells through the regulation of ADAR2 mRNA expression.%目的 探讨苯乙酸对胰腺癌诱导分化作用及RNA编辑在胰腺癌细胞增殖分化过程中的作用机理.方法 采用MTT比色法和流式细胞术检测苯乙酸对胰腺癌BXPC-3细胞系增殖抑制作用及对BXPC-3细胞周期的影响.RT-PCR方法检测RNA编辑酶ADAR mRNA在胰腺癌细胞系和胰腺癌组织中的表达以及应用苯乙酸后ADAR mRNA表达的变化.结果 人胰腺癌组织及胰腺癌BXPC-3细胞中均可检测到编辑酶ADAR2.应用1.0及2.0mmol/L苯乙酸后24h及72 h,BXPC-3细胞增殖抑制率增加,G0/G1期比例下降,S期比例显著升高.ADAR2 mRNA表达降低,差异均有统计学意义(P<0.01).结论 ADAR2可能在胰腺细胞增殖分化过程中发挥作用,且苯乙酸可能通过调控ADAR2 mRNA的表达来发挥诱导分化作用.

  15. The CRISPR/Cas genome-editing tool: application in improvement of crops

    Directory of Open Access Journals (Sweden)

    SURENDER eKHATODIA

    2016-04-01

    Full Text Available The Clustered Regularly Interspaced Short Palindromic Repeats (CRISPR associated Cas9/sgRNA system is a novel fledgling targeted genome-editing technique from bacterial immune system, which is a cheap, easy and most rapidly adopted genome editing tool transforming to revolutionary paradigm. Cas9 protein is an RNA guided endonuclease utilized for creating targeted double stranded breaks with only a short RNA sequence to confer recognition of the target in animals and plants. Development of genetically edited (GE crops similar to those developed by conventional or mutation breeding using this potential technique makes it a promising and extremely versatile tool for providing sustainable productive agriculture for better feeding of rapidly growing population in changing climate. The emerging areas of research for the genome editing in plants are like, interrogating gene function, rewiring the regulatory signaling networks, sgRNA library for high-throughput loss-of-function screening. In this review, we will discuss the broad applicability of the Cas9 nuclease mediated targeted plant genome editing for development of designer crops. The regulatory uncertainty and social acceptance of plant breeding by Cas9 genome editing have also been discussed. The non-GM designer genetically edited plants could prospect climate resilient and sustainable energy agriculture in coming future for maximizing the yield by combating abiotic and biotic stresses with this new innovative plant breeding technique.

  16. Cell and molecular biology. 8th Edition

    Energy Technology Data Exchange (ETDEWEB)

    De Robertis

    1987-01-01

    Extensively revised and reorganized, this edition can be considered a new book. The chapters on the cell membrane, cell interactions, cytoskeleton and cell motility, cell secretion, interphase nucleus, the genetic code and genetic engineering, transcription and processing of RNA, ribosomes and protein synthesis, gene regulation and differentiation are completely rewritten. More than half of the 504 illustrations are new. The discussions are concise and easy to read. Each chapter offers an introduction to the broad objectives of that chapter and concluding summary paragraphs which underscore the important points.

  17. Genome Editing in Human Pluripotent Stem Cells.

    Science.gov (United States)

    Smith, Cory; Ye, Zhaohui; Cheng, Linzhao

    2016-01-01

    Pluripotent stem cells (PSCs), defined by their capacity for self-renewal and differentiation into all cell types, are an integral tool for basic biological research and disease modeling. However, full use of PSCs for research and regenerative medicine requires the ability to precisely edit their DNA to correct disease-causing mutations and for functional analysis of genetic variations. Recent advances in DNA editing of human stem cells (including PSCs) have benefited from the use of designer nucleases capable of making double-strand breaks (DSBs) at specific sequences that stimulate endogenous DNA repair. The clustered, regularly interspaced short palindromic repeats (CRISPR)-Cas9 system has become the preferred designer nuclease for genome editing in human PSCs and other cell types. Here we describe the principles for designing a single guide RNA to uniquely target a gene of interest and describe strategies for disrupting, inserting, or replacing a specific DNA sequence in human PSCs. The improvements in efficiency and ease provided by these techniques allow individuals to precisely engineer PSCs in a way previously limited to large institutes and core facilities. PMID:27037079

  18. Psychology. 5th edition

    OpenAIRE

    Martin, G. Neil; Carlson, Neil R.; Buskist, William

    2013-01-01

    A comprehensive, lively and engaging introduction to the fascinating study of the subject. The fifth edition of the best-selling Psychology is a contemporary text that will captivate all psychology students. The authors describe and explore every major area of psychology and present the latest findings, along with clear evaluation of controversial theories and models, to give a rigorous and critical grounding in the subject. Over 420 new references in this thoroughly updated fifth editi...

  19. Efficient Mitochondrial Genome Editing by CRISPR/Cas9

    Directory of Open Access Journals (Sweden)

    Areum Jo

    2015-01-01

    Full Text Available The Clustered Regularly Interspaced Short Palindromic Repeats (CRISPR/Cas9 system has been widely used for nuclear DNA editing to generate mutations or correct specific disease alleles. Despite its flexible application, it has not been determined if CRISPR/Cas9, originally identified as a bacterial defense system against virus, can be targeted to mitochondria for mtDNA editing. Here, we show that regular FLAG-Cas9 can localize to mitochondria to edit mitochondrial DNA with sgRNAs targeting specific loci of the mitochondrial genome. Expression of FLAG-Cas9 together with gRNA targeting Cox1 and Cox3 leads to cleavage of the specific mtDNA loci. In addition, we observed disruption of mitochondrial protein homeostasis following mtDNA truncation or cleavage by CRISPR/Cas9. To overcome nonspecific distribution of FLAG-Cas9, we also created a mitochondria-targeted Cas9 (mitoCas9. This new version of Cas9 localizes only to mitochondria; together with expression of gRNA targeting mtDNA, there is specific cleavage of mtDNA. MitoCas9-induced reduction of mtDNA and its transcription leads to mitochondrial membrane potential disruption and cell growth inhibition. This mitoCas9 could be applied to edit mtDNA together with gRNA expression vectors without affecting genomic DNA. In this brief study, we demonstrate that mtDNA editing is possible using CRISPR/Cas9. Moreover, our development of mitoCas9 with specific localization to the mitochondria should facilitate its application for mitochondrial genome editing.

  20. ADAR2 editing activity in newly diagnosed versus relapsed pediatric high-grade astrocytomas

    International Nuclear Information System (INIS)

    High-grade (WHO grade III and IV) astrocytomas are aggressive malignant brain tumors affecting humans with a high risk of recurrence in both children and adults. To date, limited information is available on the genetic and molecular alterations important in the onset and progression of pediatric high-grade astrocytomas and, even less, on the prognostic factors that influence long-term outcome in children with recurrence. A-to-I RNA editing is an essential post-transcriptional mechanism that can alter the nucleotide sequence of several RNAs and is mediated by the ADAR enzymes. ADAR2 editing activity is particularly important in mammalian brain and is impaired in both adult and pediatric high-grade astrocytomas. Moreover, we have recently shown that the recovered ADAR2 activity in high-grade astrocytomas inhibits in vivo tumor growth. The aim of the present study is to investigate whether changes may occur in ADAR2-mediated RNA editing profiles of relapsed high-grade astrocytomas compared to their respective specimens collected at diagnosis, in four pediatric patients. Total RNAs extracted from all tumor samples and controls were tested for RNA editing levels (by direct sequencing on cDNA pools) and for ADAR2 mRNA expression (by qRT-PCR). A significant loss of ADAR2-editing activity was observed in the newly diagnosed and recurrent astrocytomas in comparison to normal brain. Surprisingly, we found a substantial rescue of ADAR2 editing activity in the relapsed tumor of the only patient showing prolonged survival. High-grade astrocytomas display a generalized loss of ADAR2-mediated RNA editing at both diagnosis and relapse. However, a peculiar Case, in complete remission of disease, displayed a total rescue of RNA editing at relapse, intriguingly suggesting ADAR2 activity/expression as a possible marker for long-term survival of patients with high-grade astrocytomas

  1. In vivo genome editing using Staphylococcus aureus Cas9

    OpenAIRE

    Ran, F Ann; Cong, Le; Yan, Winston X.; Scott, David A.; Gootenberg, Jonathan S.; Kriz, Andrea J.; Zetsche, Bernd; Shalem, Ophir; Wu, Xuebing; Makarova, Kira S.; Koonin, Eugene; Sharp, Phillip A.; Zhang, Feng

    2015-01-01

    The RNA-guided endonuclease Cas9 has emerged as a versatile genome-editing platform. However, the size of the commonly used Cas9 from Streptococcus pyogenes (SpCas9) limits its utility for basic research and therapeutic applications that employ the highly versatile adeno-associated virus (AAV) delivery vehicle. Here, we characterize six smaller Cas9 orthologs and show that Cas9 from Staphylococcus aureus (SaCas9) can edit the genome with efficiencies similar to those of SpCas9, while being >1...

  2. A clique-based method for the edit distance between unordered trees and its application to analysis of glycan structures

    OpenAIRE

    2011-01-01

    Background Measuring similarities between tree structured data is important for analysis of RNA secondary structures, phylogenetic trees, glycan structures, and vascular trees. The edit distance is one of the most widely used measures for comparison of tree structured data. However, it is known that computation of the edit distance for rooted unordered trees is NP-hard. Furthermore, there is almost no available software tool that can compute the exact edit distance for unordered trees. Result...

  3. Revising and editing for translators

    CERN Document Server

    Mossop, Brian

    2014-01-01

    Revising and Editing for Translators provides guidance and learning materials for translation students learning to edit texts written by others, and professional translators wishing to improve their self-revision ability or learning to revise the work of others. Editing is understood as making corrections and improvements to texts, with particular attention to tailoring them to the given readership. Revising is this same task applied to draft translations. The linguistic work of editors and revisers is related to the professional situations in which they work. Mossop offers in-depth coverage of a wide range of topics, including copyediting, style editing, structural editing, checking for consistency, revising procedures and principles, and translation quality assessment. This third edition provides extended coverage of computer aids for revisers, and of the different degrees of revision suited to different texts. The inclusion of suggested activities and exercises, numerous real-world examples, a proposed gra...

  4. Edit Distance with Block Deletions

    OpenAIRE

    Dana Shapira; Storer, James A.

    2011-01-01

    Several variants of the edit distance problem with block deletions are considered. Polynomial time optimal algorithms are presented for the edit distance with block deletions allowing character insertions and character moves, but without block moves. We show that the edit distance with block moves and block deletions is NP-complete (Nondeterministic Polynomial time problems in which any given solution to such problem can be verified in polynomial time, and any NP problem can be converted into...

  5. A contextual normalised edit distance

    OpenAIRE

    Higuera, Colin de la; Micó Andrés, Luisa

    2008-01-01

    In order to better fit a variety of pattern recognition problems over strings, using a normalised version of the edit or Levenshtein distance is considered to be an appropriate approach. The goal of normalisation is to take into account the lengths of the strings. We define a new normalisation, contextual, where each edit operation is divided by the length of the string on which the edit operation takes place. We prove that this contextual edit distance is a metric and that it can be...

  6. Benthic habitat map of U.S. Coral Reef Task Force Faga‘alu Bay priority study area, Tutuila, American Samoa

    Science.gov (United States)

    Cochran, Susan A.; Gibbs, Ann E.; D'Antonio, Nicole L.; Storlazzi, Curt D.

    2016-01-01

    The coral reef in Faga‘alu Bay, Tutuila, American Samoa, has suffered numerous natural and anthropogenic stresses. Areas once dominated by live coral are now mostly rubble surfaces covered with turf or macroalgae. In an effort to improve the health and resilience of the coral reef system, the U.S. Coral Reef Task Force selected Faga‘alu Bay as a priority study area. To support these efforts, the U.S. Geological Survey mapped nearly 1 km2 of seafloor to depths of about 60 m. Unconsolidated sediment (predominantly sand) constitutes slightly greater than 50 percent of the seafloor in the mapped area; reef and other hardbottom potentially available for coral recruitment constitute nearly 50 percent of the mapped area. Of this potentially available hardbottom, only slightly greater than 37 percent is covered with at least 10 percent coral, which is fairly evenly distributed between the reef flat, fore reef, and offshore bank/shelf. 

  7. Dynamic Texture Editing

    Czech Academy of Sciences Publication Activity Database

    Richtr, Radek; Haindl, Michal

    New York, NY, USA: ACM, 2015 - (Jorge, J.; Santos, L.; Durikovic, R.), s. 133-140. (SCCG '15). ISBN 978-1-4503-3693-2. [31st Spring Conference on Computer Graphics. Smolenice (SK), 22.04.2015-24.04.2015] R&D Projects: GA ČR(CZ) GA14-10911S Institutional support: RVO:67985556 Keywords : computer graphics * dynamic texture * dynamic texture editing * dynamic texture modeling * image generation * patch-based texture synthesi Subject RIV: BD - Theory of Information http://library.utia.cas.cz/separaty/2016/RO/haindl-0452757.pdf

  8. Sill emplacement and corresponding ground deformation processes at the Alu-Dalafilla volcanic centre in the Danakil Depression, Ethiopia

    Science.gov (United States)

    Magee, Craig; Bastow, Ian; Hetherington, Rachel; van Wyk de Vries, Ben; Jackson, Christopher

    2016-04-01

    A consensus has emerged from a variety of disciplines over the past 15 years that Quaternary magmatism in Ethiopia is almost entirely dominated by dike intrusion. Focused dike intrusion within 60 km long, 20 km wide, rift zones is considered to mark the present day locus of extension in Ethiopia, and represent the proto-ridge axis location of an incipient ocean spreading centre. However, it has been suggested on the strength of Moho depths and Quaternary eruptive volumes in northernmost Ethiopia, that the final transition from continental rifting to incipient oceanic spreading may instead be characterised by an abrupt, rheologically driven, late-phase of crustal thinning. Development of a sedimentary basin and mantle decompression melting occurring in the Danakil Depression, driven by this late-phase crustal thinning, should result in a markedly different style of magmatism in the upper crust: i.e. field observations, high-resolution seismic reflection studies, and experimental modelling suggest that interconnected networks of sill intrusions dominate in sedimentary basins. Here, we present the first evidence from the Danakil Depression that links surficial structures, observed at the Alu-Dalafilla volcanic centre, to the ongoing emplacement of an underlying sill. In particular, we use satellite imagery to examine a dome-shaped fold, associated fracture patterns, and surrounding lava flows, which we suggest likely formed in response to roof uplift above and extrusion from a saucer-shaped sill; i.e. a sub-horizontal inner sill encircled by an inward-dipping, transgressive inclined rim. InSAR observations by Pagli et al. (2012) of ground uplift and deflation occurring during the eruption of basaltic lava at Alu-Dalafilla in 2008 capture what we believe to be the first real-time evidence for intrusion-induced forced folding dynamics above a saucer-shaped sill. InSAR data further suggest that intrusion occurred at a depth of ~1 km, likely placing the sill within an

  9. The CRISPR/Cas Genome-Editing Tool: Application in Improvement of Crops

    Science.gov (United States)

    Khatodia, Surender; Bhatotia, Kirti; Passricha, Nishat; Khurana, S. M. P.; Tuteja, Narendra

    2016-01-01

    The Clustered Regularly Interspaced Short Palindromic Repeats associated Cas9/sgRNA system is a novel targeted genome-editing technique derived from bacterial immune system. It is an inexpensive, easy, most user friendly and rapidly adopted genome editing tool transforming to revolutionary paradigm. This technique enables precise genomic modifications in many different organisms and tissues. Cas9 protein is an RNA guided endonuclease utilized for creating targeted double-stranded breaks with only a short RNA sequence to confer recognition of the target in animals and plants. Development of genetically edited (GE) crops similar to those developed by conventional or mutation breeding using this potential technique makes it a promising and extremely versatile tool for providing sustainable productive agriculture for better feeding of rapidly growing population in a changing climate. The emerging areas of research for the genome editing in plants include interrogating gene function, rewiring the regulatory signaling networks and sgRNA library for high-throughput loss-of-function screening. In this review, we have described the broad applicability of the Cas9 nuclease mediated targeted plant genome editing for development of designer crops. The regulatory uncertainty and social acceptance of plant breeding by Cas9 genome editing have also been described. With this powerful and innovative technique the designer GE non-GM plants could further advance climate resilient and sustainable agriculture in the future and maximizing yield by combating abiotic and biotic stresses. PMID:27148329

  10. DETECTION OF STRAND BREAKS OF DNA IN HUMAN EARLY CHORIONIC VILLUS CELLS INDUCED BY DIAGNOSTIC ULTRASOUND USING 32p-LABELED ALU HYBRIDIZATION

    Institute of Scientific and Technical Information of China (English)

    Wang Caifeng; Li Xu; Zhang Yunjing

    2006-01-01

    Objective To explore if strand breaks of DNA in human early chorionic villus cells in uterus were induced by diagnostic ultrasound and to evaluate the method used for detection of single-stranded breaks and doublestranded breaks in human DNA. Methods 60 normal pregnant women aged 20-30, who underwent artificial abortion during 6-8 weeks of gestation, were randomly divided into 2 experimental groups: All 30 cases were exposed to diagnostic ultrasound in uterus for 10 minutes, and 24 hours later chorionic villi were extracted; the other 30 cases were taken as the control group. Single-stranded DNA and double-stranded DNA in villus cells in all cases were isolated by the alkaline unwinding combined with hydroxylapatite chromatography, and were quantitatively detected using32 P-labeled Alu probe for dot-blotting hybridization. Results There was no significant difference in quantity and percentage in single-stranded DNA and double-stranded DNA between 2 groups (P>0.05). 32 P-Alu probe could only hybridize with human DNA, and could detect DNA isolated from as few as 2.5 × 103 chorionic villus cells and 0.45 ng DNA in human leukocytes. Conclusion The results suggested that there were no DNA strand damages in human chorionic villus cells when the uterus was exposed to diagnostic ultrasound for 10 minutes. The method, 32P-Alu probe for dot-blotting hybridization, was even more specific, sensitive and accurate than conventional approaches.

  11. Natural Hazards, Second Edition

    Science.gov (United States)

    Rouhban, Badaoui

    Natural disaster loss is on the rise, and the vulnerability of the human and physical environment to the violent forces of nature is increasing. In many parts of the world, disasters caused by natural hazards such as earthquakes, floods, landslides, drought, wildfires, intense windstorms, tsunami, and volcanic eruptions have caused the loss of human lives, injury, homelessness, and the destruction of economic and social infrastructure. Over the last few years, there has been an increase in the occurrence, severity, and intensity of disasters, culminating with the devastating tsunami of 26 December 2004 in South East Asia.Natural hazards are often unexpected or uncontrollable natural events of varying magnitude. Understanding their mechanisms and assessing their distribution in time and space are necessary for refining risk mitigation measures. This second edition of Natural Hazards, (following a first edition published in 1991 by Cambridge University Press), written by Edward Bryant, associate dean of science at Wollongong University, Australia, grapples with this crucial issue, aspects of hazard prediction, and other issues. The book presents a comprehensive analysis of different categories of hazards of climatic and geological origin.

  12. The RNA WikiProject: community annotation of RNA families.

    Science.gov (United States)

    Daub, Jennifer; Gardner, Paul P; Tate, John; Ramsköld, Daniel; Manske, Magnus; Scott, William G; Weinberg, Zasha; Griffiths-Jones, Sam; Bateman, Alex

    2008-12-01

    The online encyclopedia Wikipedia has become one of the most important online references in the world and has a substantial and growing scientific content. A search of Google with many RNA-related keywords identifies a Wikipedia article as the top hit. We believe that the RNA community has an important and timely opportunity to maximize the content and quality of RNA information in Wikipedia. To this end, we have formed the RNA WikiProject (http://en.wikipedia.org/wiki/Wikipedia:WikiProject_RNA) as part of the larger Molecular and Cellular Biology WikiProject. We have created over 600 new Wikipedia articles describing families of noncoding RNAs based on the Rfam database, and invite the community to update, edit, and correct these articles. The Rfam database now redistributes this Wikipedia content as the primary textual annotation of its RNA families. Users can, therefore, for the first time, directly edit the content of one of the major RNA databases. We believe that this Wikipedia/Rfam link acts as a functioning model for incorporating community annotation into molecular biology databases. PMID:18945806

  13. RNA-SSPT: RNA Secondary Structure Prediction Tools.

    Science.gov (United States)

    Ahmad, Freed; Mahboob, Shahid; Gulzar, Tahsin; Din, Salah U; Hanif, Tanzeela; Ahmad, Hifza; Afzal, Muhammad

    2013-01-01

    The prediction of RNA structure is useful for understanding evolution for both in silico and in vitro studies. Physical methods like NMR studies to predict RNA secondary structure are expensive and difficult. Computational RNA secondary structure prediction is easier. Comparative sequence analysis provides the best solution. But secondary structure prediction of a single RNA sequence is challenging. RNA-SSPT is a tool that computationally predicts secondary structure of a single RNA sequence. Most of the RNA secondary structure prediction tools do not allow pseudoknots in the structure or are unable to locate them. Nussinov dynamic programming algorithm has been implemented in RNA-SSPT. The current studies shows only energetically most favorable secondary structure is required and the algorithm modification is also available that produces base pairs to lower the total free energy of the secondary structure. For visualization of RNA secondary structure, NAVIEW in C language is used and modified in C# for tool requirement. RNA-SSPT is built in C# using Dot Net 2.0 in Microsoft Visual Studio 2005 Professional edition. The accuracy of RNA-SSPT is tested in terms of Sensitivity and Positive Predicted Value. It is a tool which serves both secondary structure prediction and secondary structure visualization purposes. PMID:24250115

  14. Introduction to Surgical Technology. Third Edition. Teacher Edition [and] Student Edition.

    Science.gov (United States)

    Bushey, Vicki; Hildebrand, Bob; Hildebrand, Dinah; Johnson, Dave; Sikes, John; Tahah, Ann; Walker, Susan; Zielsdorf, Lani

    These teacher and student editions provide instructional materials for an introduction to surgical technology course. Introductory materials in the teacher edition include information on use, instructional/task analysis, academic and workplace skill classifications and definitions, related academic and workplace skill list, and crosswalk to…

  15. Oxyacetylene Welding and Oxyfuel Cutting. Third Edition. Teacher Edition [and] Student Edition [and] Student Workbook.

    Science.gov (United States)

    Knapp, John; Harper, Eddie

    This Oklahoma curriculum guide, which includes a teacher edition, a student edition, and a student workbook, provides three units for a course on oxyacetylene welding, oxyfuel cutting, and cutting done with alternative fuels such as MAPP, propane, and natural gas. The three units are: "Oxyacetylene Welding"; "Oxyfuel Cutting"; and "Oxyacetylene…

  16. Fundamentals of Welding. Teacher Edition [and] Student Edition [and] Student Workbook. Second Edition.

    Science.gov (United States)

    Fortney, Clarence; Gregory, Mike; New, Larry

    Teacher and student editions and a student workbook for fundamentals of welding comprise the first of six in a series of competency-based instructional materials for welding programs. Introductory pages in the teacher edition are training and competency profile, instructional/task analysis, basic skills icons and classifications, basic skills…

  17. Map Edit Distance vs Graph Edit Distance for Matching Images

    OpenAIRE

    Combier, Camille; Damiand, Guillaume; Solnon, Christine

    2013-01-01

    Generalized maps are widely used to model the topology of nD objects (such as 2D or 3D images) by means of incidence and adjacency relationships between cells (0D vertices, 1D edges, 2D faces, 3D volumes, ...). Recently, we have introduced a map edit distance. This distance compares maps by means of a minimum cost sequence of edit operations that should be performed to transform a map into another map. In this paper, we introduce labelled maps and we show how the map edit distance may be exte...

  18. PAM multiplicity marks genomic target sites as inhibitory to CRISPR-Cas9 editing

    OpenAIRE

    Malina, Abba; Cameron, Christopher J. F.; Robert, Francis; Blanchette, Mathieu; Dostie, Josée; Pelletier, Jerry

    2015-01-01

    In CRISPR-Cas9 genome editing, the underlying principles for selecting guide RNA (gRNA) sequences that would ensure for efficient target site modification remain poorly understood. Here we show that target sites harbouring multiple protospacer adjacent motifs (PAMs) are refractory to Cas9-mediated repair in situ. Thus we refine which substrates should be avoided in gRNA design, implicating PAM density as a novel sequence-specific feature that inhibits in vivo Cas9-driven DNA modification.

  19. CRISPR-ERA: a comprehensive design tool for CRISPR-mediated gene editing, repression and activation

    OpenAIRE

    Liu, Honglei; Wei, Zheng; Dominguez, Antonia; Li, Yanda; Wang, Xiaowo; Qi, Lei S.

    2015-01-01

    Summary: The CRISPR/Cas9 system was recently developed as a powerful and flexible technology for targeted genome engineering, including genome editing (altering the genetic sequence) and gene regulation (without altering the genetic sequence). These applications require the design of single guide RNAs (sgRNAs) that are efficient and specific. However, this remains challenging, as it requires the consideration of many criteria. Several sgRNA design tools have been developed for gene editing, b...

  20. CrisprGE: a central hub of CRISPR/Cas-based genome editing

    OpenAIRE

    Kaur, Karambir; Tandon, Himani; Gupta, Amit Kumar; Kumar, Manoj

    2015-01-01

    CRISPR system is a powerful defense mechanism in bacteria and archaea to provide immunity against viruses. Recently, this process found a new application in intended targeting of the genomes. CRISPR-mediated genome editing is performed by two main components namely single guide RNA and Cas9 protein. Despite the enormous data generated in this area, there is a dearth of high throughput resource. Therefore, we have developed CrisprGE, a central hub of CRISPR/Cas-based genome editing. Presently,...

  1. A non-inheritable maternal Cas9-based multiple-gene editing system in mice

    OpenAIRE

    Takayuki Sakurai; Akiko Kamiyoshi; Hisaka Kawate; Chie Mori; Satoshi Watanabe; Megumu Tanaka; Ryuichi Uetake; Masahiro Sato; Takayuki Shindo

    2016-01-01

    The CRISPR/Cas9 system is capable of editing multiple genes through one-step zygote injection. The preexisting method is largely based on the co-injection of Cas9 DNA (or mRNA) and guide RNAs (gRNAs); however, it is unclear how many genes can be simultaneously edited by this method, and a reliable means to generate transgenic (Tg) animals with multiple gene editing has yet to be developed. Here, we employed non-inheritable maternal Cas9 (maCas9) protein derived from Tg mice with systemic Cas9...

  2. Fast Quasi-Threshold Editing

    CERN Document Server

    Brandes, Ulrik; Strasser, Ben; Wagner, Dorothea

    2015-01-01

    We introduce Quasi-Threshold Mover (QTM), an algorithm to solve the quasi-threshold (also called trivially perfect) graph editing problem with edge insertion and deletion. Given a graph it computes a quasi-threshold graph which is close in terms of edit count. This edit problem is NP-hard. We present an extensive experimental study, in which we show that QTM is the first algorithm that is able to scale to large real-world graphs in practice. As a side result we further present a simple linear-time algorithm for the quasi-threshold recognition problem.

  3. lncRNA-RNA Interactions across the Human Transcriptome

    Science.gov (United States)

    Szcześniak, Michał Wojciech; Makałowska, Izabela

    2016-01-01

    Long non-coding RNAs (lncRNAs) represent a numerous class of non-protein coding transcripts longer than 200 nucleotides. There is possibility that a fraction of lncRNAs are not functional and represent mere transcriptional noise but a growing body of evidence shows they are engaged in a plethora of molecular functions and contribute considerably to the observed diversification of eukaryotic transcriptomes and proteomes. Still, however, only ca. 1% of lncRNAs have well established functions and much remains to be done towards decipherment of their biological roles. One of the least studied aspects of lncRNAs biology is their engagement in gene expression regulation through RNA-RNA interactions. By hybridizing with mate RNA molecules, lncRNAs could potentially participate in modulation of pre-mRNA splicing, RNA editing, mRNA stability control, translation activation, or abrogation of miRNA-induced repression. Here, we implemented a similarity-search based method for transcriptome-wide identification of RNA-RNA interactions, which enabled us to find 18,871,097 lncRNA-RNA base-pairings in human. Further analyses showed that the interactions could be involved in processing, stability control and functions of 57,303 transcripts. An extensive use of RNA-Seq data provided support for approximately one third of the interactions, at least in terms of the two RNA components being co-expressed. The results suggest that lncRNA-RNA interactions are broadly used to regulate and diversify the human transcriptome. PMID:26930590

  4. Selection of tRNA charging quality control mechanisms that increase mistranslation of the genetic code

    DEFF Research Database (Denmark)

    Yadavalli, Srujana S; Ibba, Michael

    2013-01-01

    Mistranslation can follow two events during protein synthesis: production of non-cognate amino acid:transfer RNA (tRNA) pairs by aminoacyl-tRNA synthetases (aaRSs) and inaccurate selection of aminoacyl-tRNAs by the ribosome. Many aaRSs actively edit non-cognate amino acids, but editing mechanisms...... are not evolutionarily conserved, and their physiological significance remains unclear. To address the connection between aaRSs and mistranslation, the evolutionary divergence of tyrosine editing by phenylalanyl-tRNA synthetase (PheRS) was used as a model. Certain PheRSs are naturally error prone...

  5. Edit while watching: home video editing made easy

    Science.gov (United States)

    Campanella, Marco; Weda, Hans; Barbieri, Mauro

    2007-01-01

    In recent years, more and more people capture their experiences in home videos. However, home video editing still is a difficult and time-consuming task. We present the Edit While Watching system that allows users to automatically create and change a summary of a home video in an easy, intuitive and lean-back way. Based on content analysis, video is indexed, segmented, and combined with proper music and editing effects. The result is an automatically generated home video summary that is shown to the user. While watching it, users can indicate whether they like certain content, so that the system will adapt the summary to contain more content that is similar or related to the displayed content. During the video playback users can also modify and enrich the content, seeing immediately the effects of their changes. Edit While Watching does not require a complex user interface: a TV and a few keys of a remote control are sufficient. A user study has shown that it is easy to learn and to use, even if users expressed the need for more control in the editing operations and in the editing process.

  6. A novel web-based TinT application and the chronology of the Primate Alu retroposon activity

    Directory of Open Access Journals (Sweden)

    Makałowski Wojciech

    2010-12-01

    Full Text Available Abstract Background DNA sequences afford access to the evolutionary pathways of life. Particularly mobile elements that constantly co-evolve in genomes encrypt recent and ancient information of their host's history. In mammals there is an extraordinarily abundant activity of mobile elements that occurs in a dynamic succession of active families, subfamilies, types, and subtypes of retroposed elements. The high frequency of retroposons in mammals implies that, by chance, such elements also insert into each other. While inactive elements are no longer able to retropose, active elements retropose by chance into other active and inactive elements. Thousands of such directional, element-in-element insertions are found in present-day genomes. To help analyze these events, we developed a computational algorithm (Transpositions in Transpositions, or TinT that examines the different frequencies of nested transpositions and reconstructs the chronological order of retroposon activities. Results By examining the different frequencies of such nested transpositions, the TinT application reconstructs the chronological order of retroposon activities. We use such activity patterns as a comparative tool to (1 delineate the historical rise and fall of retroposons and their relations to each other, (2 understand the retroposon-induced complexity of recent genomes, and (3 find selective informative homoplasy-free markers of phylogeny. The efficiency of the new application is demonstrated by applying it to dimeric Alu Short INterspersed Elements (SINE to derive a complete chronology of such elements in primates. Conclusion The user-friendly, web-based TinT interface presented here affords an easy, automated screening for nested transpositions from genome assemblies or trace data, assembles them in a frequency-matrix, and schematically displays their chronological activity history.

  7. Feedback inhibition of L1 and alu retrotransposition through altered double strand break repair kinetics

    Directory of Open Access Journals (Sweden)

    Wallace Nicholas A

    2010-10-01

    Full Text Available Abstract Background Cells adapt to various chronic toxic exposures in a multitude of ways to minimize further damage and maximize their growth potential. Expression of L1 elements in the human genome can be greatly deleterious to cells, generating numerous double strand breaks (DSBs. Cells have been reported to respond to chronic DSBs by altering the repair of these breaks, including increasing the rate of homology independent DSB repair. Retrotransposition is strongly affected by proteins involved in DSB repair. Therefore, L1 expression has the potential to be a source of chronic DSBs and thus bring about the changes in cellular environment that could ultimately restrict its own retrotransposition. Results We demonstrate that constitutive L1 expression leads to quicker DSB repair and decreases in the retrotransposition potential of L1 and other retrotransposons dependent on L1 expression for their mobility. This cellular adaptation results in reduced sensitivity to L1 induced toxicity. These effects can be induced by constitutive expression of the functional L1 ORF2 alone, but not by the constitutive expression of an L1 open reading frame 2 with mutations to its endonuclease and reverse transcriptase domains. This adaptation correlates with the relative activity of the L1 introduced into the cells. Conclusions The increased number of DSBs resulting from constitutive expression of L1 results in a more rapid rate of repair. The cellular response to this L1 expression also results in attenuation of retrotransposition and reduced sensitivity of the cells to negative consequences of L1 ORF2 expression. The influence does not appear to be through RNA interference. We believe that the increased rate of DSB repair is the most likely cause of the attenuation of retrotransposition. These alterations act as a fail safe mechanism that allows cells to escape the toxicity associated with the unchecked L1 expression. This gives cells that overexpress L1, such

  8. Circadian RNA expression elicited by 3’-UTR IRAlu-paraspeckle associated elements

    Science.gov (United States)

    Torres, Manon; Becquet, Denis; Blanchard, Marie-Pierre; Guillen, Séverine; Boyer, Bénédicte; Moreno, Mathias; Franc, Jean-Louis; François-Bellan, Anne-Marie

    2016-01-01

    Paraspeckles are nuclear bodies form around the long non-coding RNA, Neat1, and RNA-binding proteins. While their role is not fully understood, they are believed to control gene expression at a post-transcriptional level by means of the nuclear retention of mRNA containing in their 3’-UTR inverted repeats of Alu sequences (IRAlu). In this study, we found that, in pituitary cells, all components of paraspeckles including four major proteins and Neat1 displayed a circadian expression pattern. Furthermore the insertion of IRAlu at the 3’-UTR of the EGFP cDNA led to a rhythmic circadian nuclear retention of the egfp mRNA that was lost when paraspeckles were disrupted whereas insertion of a single antisense Alu had only a weak effect. Using real-time video-microscopy, these IRAlu were further shown to drive a circadian expression of EGFP protein. This study shows that paraspeckles, thanks to their circadian expression, control circadian gene expression at a post-transcriptional level. DOI: http://dx.doi.org/10.7554/eLife.14837.001 PMID:27441387

  9. Natural Editing of Algebraic Expressions

    OpenAIRE

    Nicaud, Jean-François

    2007-01-01

    We call “natural editing of algebraic expressions” the editing of algebraic expressions in their natural representation, the one that is used on paper and blackboard. This is an issue we have investigated in the Aplusix project, a project which develops a system aiming at helping students to learn algebra. The paper summarises first the Aplusix project. Second it presents a notion of algebraic expressions, of representations of algebraic expressions. The last section develops ideas about natu...

  10. Marine botany. Second edition

    Energy Technology Data Exchange (ETDEWEB)

    Dawes, C.J. [Univ. of South Florida, Tampa, FL (United States)

    1998-12-01

    Marine plants are a diverse group that include unicellular algae, seaweeds, seagrasses, salt marshes, and mangrove forests. They carry out a variety of ecological functions and serve as the primary producers in coastal wetlands and oceanic waters. The theme that connects such a wide variety of plants is their ecology, which was also emphasized in the 1981 edition. The goal of this revision is to present taxonomic, physiological, chemical, and ecological aspects of marine plants, their adaptations, and how abiotic and biotic factors interact in their communities. The data are presented in a concise, comparative manner in order to identify similarities and differences between communities such as salt marsh and mangroves or subtidal seaweeds and seagrasses. To accomplish this, the text is organized into five chapters that introduce the marine habitats, consider abiotic and biotic factors, and anthropogenic influences on the communities followed by seven chapters that deal with microalgae, seaweeds, salt marshes, mangroves, seagrasses, and coral reefs. Two appendixes are included; one presents simple field techniques and the other is a summary of seaweed uses.

  11. Maternal phthalate exposure during pregnancy is associated with DNA methylation of LINE-1 and Alu repetitive elements in Mexican-American children.

    Science.gov (United States)

    Huen, Karen; Calafat, Antonia M; Bradman, Asa; Yousefi, Paul; Eskenazi, Brenda; Holland, Nina

    2016-07-01

    Phthalates are frequently used in personal care products and plasticizers and phthalate exposure is ubiquitous in the US population. Exposure to phthalates during critical periods in utero has been associated with a variety of adverse health outcomes but the biological mechanisms linking these exposures with disease are not well characterized. In this study, we examined the relationship of in utero phthalate exposure with repetitive element DNA methylation, an epigenetic marker of genome instability, in children from the longitudinal birth cohort CHAMACOS. Methylation of Alu and long interspersed nucleotide elements (LINE-1) was determined using pyrosequencing of bisulfite-treated DNA isolated from whole blood samples collected from newborns and 9 year old children (n=355). Concentrations of eleven phthalate metabolites were measured in urine collected from pregnant mothers at 13 and 26 weeks gestation. We found a consistent inverse association between prenatal concentrations of monoethyl phthalate, the most frequently detected urinary metabolite, with cord blood methylation of Alu repeats (β(95%CI): -0.14 (-0.28,0.00) and -0.16 (-0.31, -0.02)) for early and late pregnancy, respectively, and a similar but weaker association with LINE-1 methylation. Additionally, increases in urinary concentrations of di-(2-ethylhexyl) phthalate metabolites during late pregnancy were associated with lower levels of methylation of Alu repeats in 9 year old blood (significant p-values ranged from 0.003 to 0.03). Our findings suggest that prenatal exposure to some phthalates may influence differences in repetitive element methylation, highlighting epigenetics as a plausible biological mechanism through which phthalates may affect health. PMID:27019040

  12. Leucyl-tRNA synthetase from the ancestral bacterium Aquifex aeolicus contains relics of synthetase evolution

    OpenAIRE

    Zhao, Ming-Wei; Zhu, Bin; Hao, Rui; Xu, Min-Gang; Eriani, Gilbert; Wang, En-Duo

    2005-01-01

    The editing reactions catalyzed by aminoacyl-tRNA synthetases are critical for the faithful protein synthesis by correcting misactivated amino acids and misaminoacylated tRNAs. We report that the isolated editing domain of leucyl-tRNA synthetase from the deep-rooted bacterium Aquifex aeolicus (αβ-LeuRS) catalyzes the hydrolytic editing of both mischarged tRNALeu and minihelixLeu. Within the domain, we have identified a crucial 20-amino-acid peptide that confers editing capacity when transplan...

  13. CRISPR/Cas9-Mediated Genome Editing in Soybean Hairy Roots

    OpenAIRE

    Yupeng Cai; Li Chen; Xiujie Liu; Shi Sun; Cunxiang Wu; Bingjun Jiang; Tianfu Han; Wensheng Hou

    2015-01-01

    As a new technology for gene editing, the CRISPR (clustered regularly interspaced short palindromic repeat)/Cas (CRISPR-associated) system has been rapidly and widely used for genome engineering in various organisms. In the present study, we successfully applied type II CRISPR/Cas9 system to generate and estimate genome editing in the desired target genes in soybean (Glycine max (L.) Merrill.). The single-guide RNA (sgRNA) and Cas9 cassettes were assembled on one vector to improve transformat...

  14. RNA genetics

    Energy Technology Data Exchange (ETDEWEB)

    Domingo, E. (Instituto de Biologia Molecular, Facultad de Ciencias, Universidad Autonoma de Madrid, Canto Blanco, Madrid (ES)); Holland, J.J. (California Univ., San Diego, La Jolla, CA (USA). Dept. of Biology); Ahlquist, P. (Wisconsin Univ., Madison, WI (USA). Dept. of Plant Pathology)

    1988-01-01

    This book contains the proceedings on RNA genetics: Retroviruses, Viroids, and RNA recombination, Volume 2. Topics covered include: Replication of retrovirus genomes, Hepatitis B virus replication, and Evolution of RNA viruses.

  15. RNA Sociology: Group Behavioral Motifs of RNA Consortia

    Directory of Open Access Journals (Sweden)

    Guenther Witzany

    2014-11-01

    Full Text Available RNA sociology investigates the behavioral motifs of RNA consortia from the social science perspective. Besides the self-folding of RNAs into single stem loop structures, group building of such stem loops results in a variety of essential agents that are highly active in regulatory processes in cellular and non-cellular life. RNA stem loop self-folding and group building do not depend solely on sequence syntax; more important are their contextual (functional needs. Also, evolutionary processes seem to occur through RNA stem loop consortia that may act as a complement. This means the whole entity functions only if all participating parts are coordinated, although the complementary building parts originally evolved for different functions. If complementary groups, such as rRNAs and tRNAs, are placed together in selective pressure contexts, new evolutionary features may emerge. Evolution initiated by competent agents in natural genome editing clearly contrasts with statistical error replication narratives.

  16. GluA2 is rapidly edited at the Q/R site during neural differentiation in vitro

    Directory of Open Access Journals (Sweden)

    Svenja ePachernegg

    2015-03-01

    Full Text Available The majority of AMPA receptors in the adult brain contain GluA2 subunits, which can be edited at the Q/R site, changing a glutamine to an arginine within the ion pore. Q/R editing renders AMPARs virtually Ca2+-impermeable, which is important for normal AMPA receptor function. Thus, all GluA2 subunits are Q/R-edited in the adult brain. However, it has remained controversial precisely when editing sets in during development. In the present study, we show that GluA2 mRNA is very rapidly Q/R-edited immediately after its appearance, which is after 4.5 days of differentiation from 46C embryonic stem cells (ESCs to neuroepithelial precursor cells (NEPs. At this time point, most of the GluA2 transcripts were already edited, with only a small fraction remaining unedited, and half a day later all GluA2 transcripts were edited. This can be explained by the observation that the enzyme that Q/R-edits GluA2 transcripts, ADAR2, is already expressed in the cell well before GluA2 transcription starts, and later is not significantly upregulated any more. Editing at another site works differently: The R/G site within the ligand-binding domain was never completely edited at any of the developmental stages tested, and the enzyme that performs this editing, ADAR1, was significantly upregulated during neural differentiation. This confirms previous data suggesting that R/G editing, in contrast to Q/R editing, progresses gradually during development.

  17. Connectivity editing for quadrilateral meshes

    KAUST Repository

    Peng, Chihan

    2011-12-12

    We propose new connectivity editing operations for quadrilateral meshes with the unique ability to explicitly control the location, orientation, type, and number of the irregular vertices (valence not equal to four) in the mesh while preserving sharp edges. We provide theoretical analysis on what editing operations are possible and impossible and introduce three fundamental operations to move and re-orient a pair of irregular vertices. We argue that our editing operations are fundamental, because they only change the quad mesh in the smallest possible region and involve the fewest irregular vertices (i.e., two). The irregular vertex movement operations are supplemented by operations for the splitting, merging, canceling, and aligning of irregular vertices. We explain how the proposed highlevel operations are realized through graph-level editing operations such as quad collapses, edge flips, and edge splits. The utility of these mesh editing operations are demonstrated by improving the connectivity of quad meshes generated from state-of-art quadrangulation techniques. © 2011 ACM.

  18. Connectivity editing for quadrilateral meshes

    KAUST Repository

    Peng, Chihan

    2011-12-01

    We propose new connectivity editing operations for quadrilateral meshes with the unique ability to explicitly control the location, orientation, type, and number of the irregular vertices (valence not equal to four) in the mesh while preserving sharp edges. We provide theoretical analysis on what editing operations are possible and impossible and introduce three fundamental operations to move and re-orient a pair of irregular vertices. We argue that our editing operations are fundamental, because they only change the quad mesh in the smallest possible region and involve the fewest irregular vertices (i.e., two). The irregular vertex movement operations are supplemented by operations for the splitting, merging, canceling, and aligning of irregular vertices. We explain how the proposed high-level operations are realized through graph-level editing operations such as quad collapses, edge flips, and edge splits. The utility of these mesh editing operations are demonstrated by improving the connectivity of quad meshes generated from state-of-art quadrangulation techniques.

  19. Newer Gene Editing Technologies toward HIV Gene Therapy

    Directory of Open Access Journals (Sweden)

    Premlata Shankar

    2013-11-01

    Full Text Available Despite the great success of highly active antiretroviral therapy (HAART in ameliorating the course of HIV infection, alternative therapeutic approaches are being pursued because of practical problems associated with life-long therapy. The eradication of HIV in the so-called “Berlin patient” who received a bone marrow transplant from a CCR5-negative donor has rekindled interest in genome engineering strategies to achieve the same effect. Precise gene editing within the cells is now a realistic possibility with recent advances in understanding the DNA repair mechanisms, DNA interaction with transcription factors and bacterial defense mechanisms. Within the past few years, four novel technologies have emerged that can be engineered for recognition of specific DNA target sequences to enable site-specific gene editing: Homing Endonuclease, ZFN, TALEN, and CRISPR/Cas9 system. The most recent CRISPR/Cas9 system uses a short stretch of complementary RNA bound to Cas9 nuclease to recognize and cleave target DNA, as opposed to the previous technologies that use DNA binding motifs of either zinc finger proteins or transcription activator-like effector molecules fused to an endonuclease to mediate sequence-specific DNA cleavage. Unlike RNA interference, which requires the continued presence of effector moieties to maintain gene silencing, the newer technologies allow permanent disruption of the targeted gene after a single treatment. Here, we review the applications, limitations and future prospects of novel gene-editing strategies for use as HIV therapy.

  20. Growth and carcass traits associated with GH1/Alu I and POU1F1/Hinf I gene polymorphisms in Zebu and crossbred beef cattle

    Directory of Open Access Journals (Sweden)

    Rogério A. Curi

    2006-01-01

    Full Text Available The objectives of the present study were to estimate the allele and genotype frequencies of the GH1/Alu I and POU1F1/Hinf I polymorphisms in beef cattle belonging to different genetic groups and to determine the effects of these polymorphisms on growth and carcass traits in cattle submitted to feedlot management, an intensive production model. Genotyping was performed on 384 animals, including 79 Nellore, 30 Canchim (5/8 Charolais + 3/8 Zebu, 30 Simmental x Nellore crossbred and 245 Angus x Nellore crossbred cattle. Body weight, weight gain, dressing percentage, Longissimus dorsi area and backfat thickness were fitted using the General Linear Model (GLM procedure of the SAS program and the least square means of the genotypes were compared using the F test. The results showed significant associations between the LL genotype of the GH1/Alu I polymorphism and higher weight gain and body weight at slaughter (p < 0.05. The POU1F1/Hinf I polymorphism did not have any effect on the growth and carcass traits analyzed.

  1. Understanding Editing Behaviors in Multilingual Wikipedia

    Science.gov (United States)

    Hale, Scott A.; Kim, Sooyoung; Byun, Jeongmin; Oh, Alice H.

    2016-01-01

    Multilingualism is common offline, but we have a more limited understanding of the ways multilingualism is displayed online and the roles that multilinguals play in the spread of content between speakers of different languages. We take a computational approach to studying multilingualism using one of the largest user-generated content platforms, Wikipedia. We study multilingualism by collecting and analyzing a large dataset of the content written by multilingual editors of the English, German, and Spanish editions of Wikipedia. This dataset contains over two million paragraphs edited by over 15,000 multilingual users from July 8 to August 9, 2013. We analyze these multilingual editors in terms of their engagement, interests, and language proficiency in their primary and non-primary (secondary) languages and find that the English edition of Wikipedia displays different dynamics from the Spanish and German editions. Users primarily editing the Spanish and German editions make more complex edits than users who edit these editions as a second language. In contrast, users editing the English edition as a second language make edits that are just as complex as the edits by users who primarily edit the English edition. In this way, English serves a special role bringing together content written by multilinguals from many language editions. Nonetheless, language remains a formidable hurdle to the spread of content: we find evidence for a complexity barrier whereby editors are less likely to edit complex content in a second language. In addition, we find that multilinguals are less engaged and show lower levels of language proficiency in their second languages. We also examine the topical interests of multilingual editors and find that there is no significant difference between primary and non-primary editors in each language. PMID:27171158

  2. Understanding Editing Behaviors in Multilingual Wikipedia.

    Directory of Open Access Journals (Sweden)

    Suin Kim

    Full Text Available Multilingualism is common offline, but we have a more limited understanding of the ways multilingualism is displayed online and the roles that multilinguals play in the spread of content between speakers of different languages. We take a computational approach to studying multilingualism using one of the largest user-generated content platforms, Wikipedia. We study multilingualism by collecting and analyzing a large dataset of the content written by multilingual editors of the English, German, and Spanish editions of Wikipedia. This dataset contains over two million paragraphs edited by over 15,000 multilingual users from July 8 to August 9, 2013. We analyze these multilingual editors in terms of their engagement, interests, and language proficiency in their primary and non-primary (secondary languages and find that the English edition of Wikipedia displays different dynamics from the Spanish and German editions. Users primarily editing the Spanish and German editions make more complex edits than users who edit these editions as a second language. In contrast, users editing the English edition as a second language make edits that are just as complex as the edits by users who primarily edit the English edition. In this way, English serves a special role bringing together content written by multilinguals from many language editions. Nonetheless, language remains a formidable hurdle to the spread of content: we find evidence for a complexity barrier whereby editors are less likely to edit complex content in a second language. In addition, we find that multilinguals are less engaged and show lower levels of language proficiency in their second languages. We also examine the topical interests of multilingual editors and find that there is no significant difference between primary and non-primary editors in each language.

  3. Understanding Editing Behaviors in Multilingual Wikipedia.

    Science.gov (United States)

    Kim, Suin; Park, Sungjoon; Hale, Scott A; Kim, Sooyoung; Byun, Jeongmin; Oh, Alice H

    2016-01-01

    Multilingualism is common offline, but we have a more limited understanding of the ways multilingualism is displayed online and the roles that multilinguals play in the spread of content between speakers of different languages. We take a computational approach to studying multilingualism using one of the largest user-generated content platforms, Wikipedia. We study multilingualism by collecting and analyzing a large dataset of the content written by multilingual editors of the English, German, and Spanish editions of Wikipedia. This dataset contains over two million paragraphs edited by over 15,000 multilingual users from July 8 to August 9, 2013. We analyze these multilingual editors in terms of their engagement, interests, and language proficiency in their primary and non-primary (secondary) languages and find that the English edition of Wikipedia displays different dynamics from the Spanish and German editions. Users primarily editing the Spanish and German editions make more complex edits than users who edit these editions as a second language. In contrast, users editing the English edition as a second language make edits that are just as complex as the edits by users who primarily edit the English edition. In this way, English serves a special role bringing together content written by multilinguals from many language editions. Nonetheless, language remains a formidable hurdle to the spread of content: we find evidence for a complexity barrier whereby editors are less likely to edit complex content in a second language. In addition, we find that multilinguals are less engaged and show lower levels of language proficiency in their second languages. We also examine the topical interests of multilingual editors and find that there is no significant difference between primary and non-primary editors in each language. PMID:27171158

  4. A non-inheritable maternal Cas9-based multiple-gene editing system in mice.

    Science.gov (United States)

    Sakurai, Takayuki; Kamiyoshi, Akiko; Kawate, Hisaka; Mori, Chie; Watanabe, Satoshi; Tanaka, Megumu; Uetake, Ryuichi; Sato, Masahiro; Shindo, Takayuki

    2016-01-01

    The CRISPR/Cas9 system is capable of editing multiple genes through one-step zygote injection. The preexisting method is largely based on the co-injection of Cas9 DNA (or mRNA) and guide RNAs (gRNAs); however, it is unclear how many genes can be simultaneously edited by this method, and a reliable means to generate transgenic (Tg) animals with multiple gene editing has yet to be developed. Here, we employed non-inheritable maternal Cas9 (maCas9) protein derived from Tg mice with systemic Cas9 overexpression (Cas9 mice). The maCas9 protein in zygotes derived from mating or in vitro fertilization of Tg/+ oocytes and +/+ sperm could successfully edit the target genome. The efficiency of such maCas9-based genome editing was comparable to that of zygote microinjection-based genome editing widely used at present. Furthermore, we demonstrated a novel approach to create "Cas9 transgene-free" gene-modified mice using non-Tg (+/+) zygotes carrying maCas9. The maCas9 protein in mouse zygotes edited nine target loci simultaneously after injection with nine different gRNAs alone. Cas9 mouse-derived zygotes have the potential to facilitate the creation of genetically modified animals carrying the Cas9 transgene, enabling repeatable genome engineering and the production of Cas9 transgene-free mice. PMID:26817415

  5. Intronic Alus influence alternative splicing.

    Directory of Open Access Journals (Sweden)

    Galit Lev-Maor

    Full Text Available Examination of the human transcriptome reveals higher levels of RNA editing than in any other organism tested to date. This is indicative of extensive double-stranded RNA (dsRNA formation within the human transcriptome. Most of the editing sites are located in the primate-specific retrotransposed element called Alu. A large fraction of Alus are found in intronic sequences, implying extensive Alu-Alu dsRNA formation in mRNA precursors. Yet, the effect of these intronic Alus on splicing of the flanking exons is largely unknown. Here, we show that more Alus flank alternatively spliced exons than constitutively spliced ones; this is especially notable for those exons that have changed their mode of splicing from constitutive to alternative during human evolution. This implies that Alu insertions may change the mode of splicing of the flanking exons. Indeed, we demonstrate experimentally that two Alu elements that were inserted into an intron in opposite orientation undergo base-pairing, as evident by RNA editing, and affect the splicing patterns of a downstream exon, shifting it from constitutive to alternative. Our results indicate the importance of intronic Alus in influencing the splicing of flanking exons, further emphasizing the role of Alus in shaping of the human transcriptome.

  6. Gas Metal Arc Welding and Flux-Cored Arc Welding. Third Edition. Teacher Edition [and] Student Edition [and] Student Workbook.

    Science.gov (United States)

    Knapp, John; Harper, Eddie

    This packet, containing a teacher's edition, a student edition, and a student workbook, introduces students to high deposition welding and processes for "shielding" a weld. In addition to general information, the teacher edition consists of introductory pages and teacher pages, as well as unit information that corresponds to the materials in the…

  7. RNA-DNA Differences Are Generated in Human Cells within Seconds after RNA Exits Polymerase II

    Directory of Open Access Journals (Sweden)

    Isabel X. Wang

    2014-03-01

    Full Text Available RNA sequences are expected to be identical to their corresponding DNA sequences. Here, we found all 12 types of RNA-DNA sequence differences (RDDs in nascent RNA. Our results show that RDDs begin to occur in RNA chains ∼55 nt from the RNA polymerase II (Pol II active site. These RDDs occur so soon after transcription that they are incompatible with known deaminase-mediated RNA-editing mechanisms. Moreover, the 55 nt delay in appearance indicates that they do not arise during RNA synthesis by Pol II or as a direct consequence of modified base incorporation. Preliminary data suggest that RDD and R-loop formations may be coupled. These findings identify sequence substitution as an early step in cotranscriptional RNA processing.

  8. Beginning XML, 5th Edition

    CERN Document Server

    Fawcett, Joe; Quin, Liam R E

    2012-01-01

    A complete update covering the many advances to the XML language The XML language has become the standard for writing documents on the Internet and is constantly improving and evolving. This new edition covers all the many new XML-based technologies that have appeared since the previous edition four years ago, providing you with an up-to-date introductory guide and reference. Packed with real-world code examples, best practices, and in-depth coverage of the most important and relevant topics, this authoritative resource explores both the advantages and disadvantages of XML and addresses the mo

  9. Introducing ZBrush 3rd Edition

    CERN Document Server

    Keller, Eric

    2012-01-01

    Learn ZBrush inside and out with this updated new edition Get totally comfortable sculpting in a digital environment with the latest edition of this bestselling beginner's guide to ZBrush. Fully updated for the newest version of the software, ZBrush 4R3, this book dispels any fears you might have about the difficulty of using ZBrush and soon has you creating realistic, cartoon, and organic models with flair. Learn all the essentials, as you complete fun tutorials on painting, meshes, organic scripting, hard surface sculpting, lighting, rendering, and more. Introduces you to ZBrush, the sculpt

  10. DNA Editing by APOBECs: A Genomic Preserver and Transformer.

    Science.gov (United States)

    Knisbacher, Binyamin A; Gerber, Doron; Levanon, Erez Y

    2016-01-01

    Information warfare is not limited to the cyber world because it is waged within our cells as well. The unique AID (activation-induced cytidine deaminase)/APOBEC (apolipoprotein B mRNA editing enzyme, catalytic polypeptide) family comprises proteins that alter DNA sequences by converting deoxycytidines to deoxyuridines through deamination. This C-to-U DNA editing enables them to inhibit parasitic viruses and retrotransposons by disrupting their genomic content. In addition to attacking genomic invaders, APOBECs can target their host genome, which can be beneficial by initiating processes that create antibody diversity needed for the immune system or by accelerating the rate of evolution. AID can also alter gene regulation by removing epigenetic modifications from genomic DNA. However, when uncontrolled, these powerful agents of change can threaten genome stability and eventually lead to cancer. PMID:26608778

  11. Methods for Optimizing CRISPR-Cas9 Genome Editing Specificity.

    Science.gov (United States)

    Tycko, Josh; Myer, Vic E; Hsu, Patrick D

    2016-08-01

    Advances in the development of delivery, repair, and specificity strategies for the CRISPR-Cas9 genome engineering toolbox are helping researchers understand gene function with unprecedented precision and sensitivity. CRISPR-Cas9 also holds enormous therapeutic potential for the treatment of genetic disorders by directly correcting disease-causing mutations. Although the Cas9 protein has been shown to bind and cleave DNA at off-target sites, the field of Cas9 specificity is rapidly progressing, with marked improvements in guide RNA selection, protein and guide engineering, novel enzymes, and off-target detection methods. We review important challenges and breakthroughs in the field as a comprehensive practical guide to interested users of genome editing technologies, highlighting key tools and strategies for optimizing specificity. The genome editing community should now strive to standardize such methods for measuring and reporting off-target activity, while keeping in mind that the goal for specificity should be continued improvement and vigilance. PMID:27494557

  12. [An introduction on the editions of Zhang Gao's Yi shuo (Medical Narrations)].

    Science.gov (United States)

    Wang, Xuguang; Lu, Xiang

    2014-11-01

    Zhang Gao's Yi shuo (Medical Narrations) of the Southern Song Dynasty had 2 kinds of editions: domestic editions and foreign editions. The former includes 1 Song edition, 14 Ming editions, 3 Qing editions and 25 editions after the Republic of China. The latter, mainly 2 classes, the Japanese edition and Korean printing type edition. In the Ming Dynasty, the editions of Yi shuo generated 2 branches: inherited edition and supplementary edition. The inherited editions include Gu Dingfang's edition, Zhang Yaode's edition, Wu Mianxue's edition, Wu Zhongheng's edition, Wang Kentang's edition, editions from Si ku quan shu (Imperial Collection of Four), stereotype edition of the 3th year of Xuantong reign (1911) from Shanghai Civilization Bookstore, edition of the 2(nd) year of Manji of Japan etc. The supplemental editions include Zhang Zili's edition, Shen Fan's edition, Fu Feng'ao's edition, transcript of the late Ming Dynasty preserved in the Library of Peking University, and Korean printing type edition etc. PMID:25620361

  13. Civil Technology Applications. Teacher Edition [and] Student Edition.

    Science.gov (United States)

    Schertz, Karen

    Teacher and student editions of Civil Technology Applications are one in a series of competency-based instructional materials for drafting and civil technology programs. It includes the technical content and tasks necessary for a student to be employed as a drafter or civil technician in a civil engineering firm. Introductory pages in the teacher…

  14. Controlled flexibility in technical editing - The levels-of-edit concept at JPL

    Science.gov (United States)

    Buehler, M. F.

    1977-01-01

    The levels-of-edit concept, which can be used to specify the amount of editorial effort involved in the preparation of a manuscript for publication, is discussed. Nine types of editing are identified and described. These include coordination edit (preparing estimates, gathering cost data, monitoring production processes), policy edit, integrity edit (making sure that parts of a publication match in a physical or numerical sense), screening edit (ensuring that the quality of camera-ready copy is sufficient for external publication), copy clarification edit, format edit, mechanical style edit, language edit, and substantive edit (reviewing the manuscript for content coherence, emphasis, subordination and parallelism). These functions are grouped into five levels of edit. An edit-level number is assigned to each manuscript, providing a quantitative and qualitative indicator of the editing to be done which is clearly understood by authors, managers, and editors alike. In addition, clear boundaries are drawn between normal and extraordinary editing tasks. Individual organizations will group various edits in different ways to reflect their needs and priorities; the essential element of the system is unambiguous definition and coding of the types and amount of work to be done.

  15. New vectors for simple and streamlined CRISPR-Cas9 genome editing in Saccharomyces cerevisiae.

    Science.gov (United States)

    Laughery, Marian F; Hunter, Tierra; Brown, Alexander; Hoopes, James; Ostbye, Travis; Shumaker, Taven; Wyrick, John J

    2015-12-01

    Clustered regularly interspaced short palindromic repeats (CRISPR)-Cas9 technology is an important tool for genome editing because the Cas9 endonuclease can induce targeted DNA double-strand breaks. Targeting of the DNA break is typically controlled by a single-guide RNA (sgRNA), a chimeric RNA containing a structural segment important for Cas9 binding and a 20mer guide sequence that hybridizes to the genomic DNA target. Previous studies have demonstrated that CRISPR-Cas9 technology can be used for efficient, marker-free genome editing in Saccharomyces cerevisiae. However, introducing the 20mer guide sequence into yeast sgRNA expression vectors often requires cloning procedures that are complex, time-consuming and/or expensive. To simplify this process, we have developed a new sgRNA expression cassette with internal restriction enzyme sites that permit rapid, directional cloning of 20mer guide sequences. Here we describe a flexible set of vectors based on this design for cloning and expressing sgRNAs (and Cas9) in yeast using different selectable markers. We anticipate that the Cas9-sgRNA expression vector with the URA3 selectable marker (pML104) will be particularly useful for genome editing in yeast, since the Cas9 machinery can be easily removed by counter-selection using 5-fluoro-orotic acid (5-FOA) following successful genome editing. The availability of new vectors that simplify and streamline the technical steps required for guide sequence cloning should help accelerate the use of CRISPR-Cas9 technology in yeast genome editing. PMID:26305040

  16. Parameter-Free Extended Edit Distance

    OpenAIRE

    Muhammad Fuad, Muhammad Marwan

    2014-01-01

    The edit distance is the most famous distance to compute the similarity between two strings of characters. The main drawback of the edit distance is that it is based on local procedures which reflect only a local view of similarity. To remedy this problem we presented in a previous work the extended edit distance, which adds a global view of similarity between two strings. However, the extended edit distance includes a parameter whose computation requires a long training time. In this paper w...

  17. RNA-Editing with Combined Insertion and Deletion Preserves Regularity

    OpenAIRE

    Vink, E.P.; Zantema, H.; Bosnacki, D.

    2013-01-01

    We consider two elementary forms of string rewriting called guided insertion/deletion and guided rewriting. The original strings are modified depending on the match with a given set of auxiliary strings, called guides. Guided insertion/deletion considers matching of a string and a guide with respect to a specific correspondence of strings. Guided rewriting considers matching of a string and a guide with respect to an equivalence relation on the alphabet. Guided insertion/deletion is inspired ...

  18. Efficient In Vivo Genome Editing Using RNA-Guided Nucleases

    OpenAIRE

    Hwang, Woong Y.; Fu, Yanfang; Reyon, Deepak; Maeder, Morgan L.; Tsai, Shengdar Q.; Sander, Jeffry D.; Peterson, Randall T.; Yeh, J.-R. Joanna; Joung, J. Keith

    2013-01-01

    Clustered regularly interspaced short palindromic repeats (CRISPR)/CRISPR-associated (Cas) systems have evolved in bacteria and archaea as a defense mechanism to silence foreign nucleic acids of viruses and plasmids. Recent work has shown that bacterial type II CRISPR systems can be adapted to create guide RNAs (gRNAs) capable of directing site-specific DNA cleavage by the Cas9 nuclease in vitro. Here we show that this system can function in vivo to induce targeted genetic modifications in ze...

  19. Strategies of Qualitative Inquiry. Third Edition

    Science.gov (United States)

    Denzin, Norman K., Ed.; Lincoln, Yvonna S., Ed.

    2007-01-01

    "Strategies of Qualitative Inquiry, Third Edition," the second volume in the paperback version of "The SAGE Handbook of Qualitative Research, 3rd Edition," consists of Part III of the handbook ("Strategies of Inquiry"). "Strategies of Qualitative Inquiry, Third Edition" presents the major tactics--historically, the research methods--that…

  20. Efficient CRISPR/Cas9 Plasmids for Rapid and Versatile Genome Editing in Drosophila

    OpenAIRE

    Gokcezade, Joseph; Sienski, Grzegorz; Duchek, Peter

    2014-01-01

    The CRISPR-associated RNA-guided nuclease Cas9 has emerged as a powerful tool for genome engineering in a variety of organisms. To achieve efficient gene targeting rates in Drosophila, current approaches require either injection of in vitro transcribed RNAs or injection into transgenic Cas9-expressing embryos. We report a simple and versatile alternative method for CRISPR-mediated genome editing in Drosophila using bicistronic Cas9/sgRNA expression vectors. Gene targeting with this single-pla...

  1. Transgene-Free Genome Editing in Caenorhabditis elegans Using CRISPR-Cas

    OpenAIRE

    Chiu, Hui; Schwartz, Hillel T.; Antoshechkin, Igor; Paul W Sternberg

    2013-01-01

    CRISPR-Cas is an efficient method for genome editing in organisms from bacteria to human cells. We describe a transgene-free method for CRISPR-Cas-mediated cleavage in nematodes, enabling RNA-homology-targeted deletions that cause loss of gene function; analysis of whole-genome sequencing indicates that the nuclease activity is highly specific.

  2. Money and Schools. Fifth Edition

    Science.gov (United States)

    Thompson, David C.; Crampton, Faith E.; Wood, R. Craig

    2012-01-01

    In the new edition of this essential, all-inclusive text, the authors provide more important research for future principals and others enrolled in graduate-level school finance courses. Written in a style that is highly readable, the book offers strong connections to real-world experiences. Readers get both a broad overview of funding concepts and…

  3. Medical writing, revising and editing

    DEFF Research Database (Denmark)

    Pilegaard, Morten

    2006-01-01

    The globalization of science makes medical writing, editing and revision a rapidly growing field of linguistic study and practice. Medical science texts are written according to uniform, general guidelines and medical genres have become highly conventionalized in terms of structure and linguistic...

  4. Tea and cake [second edition

    OpenAIRE

    Wood, Philippa; MacLellan, Tamar

    2011-01-01

    Tea & Cake is an edited version of the original book produced by Philippa Wood in 2007. The book takes a nostalgic look at childhood memories of ‘baking with mother’ or special tea-time treats. The book combines ink-jet printing with typewritten text and rubber stamps; doilly end-papers and embroidered traycloth covers.

  5. Veterinary Microbiology, 3rd Edition

    Science.gov (United States)

    Veterinary Microbiology, Third Edition is organized into four sections and begins with an updated and expanded introductory section on infectious disease pathogenesis, diagnosis and clinical management. The second section covers bacterial and fungal pathogens, and the third section describes viral d...

  6. Testing post-editing guidelines

    DEFF Research Database (Denmark)

    Flanagan, Marian; Christensen, Tina Paulsen

    2014-01-01

    There is a growing interest in machine translation (MT) and post-editing (PE). MT has been around for decades, but the use of the technology has grown significantly in the language industry in recent years, while PE is still a relatively new task. Consequently, there are currently no standard PE...

  7. Strengthening Family Resilience, Second Edition

    Science.gov (United States)

    Walsh, Froma

    2006-01-01

    In a fully revised, updated, and expanded second edition, this informative clinical resource and text presents Froma Walsh's family resilience framework for intervention and prevention with clients dealing with adversity. Drawing on extensive research and clinical experience, the author describes key processes in resilience for practitioners to…

  8. Teaching Reading Sourcebook, Second Edition

    Science.gov (United States)

    Honig, Bill; Diamond, Linda; Gutlohn, Linda

    2008-01-01

    The "Teaching Reading Sourcebook, Second Edition" is a comprehensive reference about reading instruction. Organized according to the elements of explicit instruction (what? why? when? and how?), the "Sourcebook" includes both a research-informed knowledge base and practical sample lesson models. It teaches the key elements of an effective reading…

  9. Coupling and Coordination in Gene Expression Processes with Pre-mRNA Splicing

    OpenAIRE

    Kewu Pan; Jimmy Tsz Hang Lee; Zhe Huang; Chi-Ming Wong

    2015-01-01

    RNA processing is a tightly regulated and highly complex pathway which includes transcription, splicing, editing, transportation, translation and degradation. It has been well-documented that splicing of RNA polymerase II medicated nascent transcripts occurs co-transcriptionally and is functionally coupled to other RNA processing. Recently, increasing experimental evidence indicated that pre-mRNA splicing influences RNA degradation and vice versa. In this review, we summarized the recent find...

  10. BRASERO: A Resource for Benchmarking RNA Secondary Structure Comparison Algorithms

    OpenAIRE

    Chauve, Cedric; Allali, Julien; Saule, Cedric

    2012-01-01

    The pairwise comparison of RNA secondary structures is a fundamental problem, with direct application in mining databases for annotating putative noncoding RNA candidates in newly sequenced genomes. An increasing number of software tools are available for comparing RNA secondary structures, based on different models (such as ordered trees or forests, arc annotated sequences, and multilevel trees) and computational principles (edit distance, alignment). We describe here the website BRASERO tha...

  11. BRASERO: A Resource for Benchmarking RNA Secondary Structure Comparison Algorithms.

    Science.gov (United States)

    Allali, Julien; Saule, Cédric; Chauve, Cédric; d'Aubenton-Carafa, Yves; Denise, Alain; Drevet, Christine; Ferraro, Pascal; Gautheret, Daniel; Herrbach, Claire; Leclerc, Fabrice; de Monte, Antoine; Ouangraoua, Aida; Sagot, Marie-France; Termier, Michel; Thermes, Claude; Touzet, Hélène

    2012-01-01

    The pairwise comparison of RNA secondary structures is a fundamental problem, with direct application in mining databases for annotating putative noncoding RNA candidates in newly sequenced genomes. An increasing number of software tools are available for comparing RNA secondary structures, based on different models (such as ordered trees or forests, arc annotated sequences, and multilevel trees) and computational principles (edit distance, alignment). We describe here the website BRASERO that offers tools for evaluating such software tools on real and synthetic datasets. PMID:22675348

  12. Recent Advances in Genome Editing Using CRISPR/Cas9

    Science.gov (United States)

    Ding, Yuduan; Li, Hong; Chen, Ling-Ling; Xie, Kabin

    2016-01-01

    The CRISPR (clustered regularly interspaced short palindromic repeat)-Cas9 (CRISPR-associated nuclease 9) system is a versatile tool for genome engineering that uses a guide RNA (gRNA) to target Cas9 to a specific sequence. This simple RNA-guided genome-editing technology has become a revolutionary tool in biology and has many innovative applications in different fields. In this review, we briefly introduce the Cas9-mediated genome-editing method, summarize the recent advances in CRISPR/Cas9 technology, and discuss their implications for plant research. To date, targeted gene knockout using the Cas9/gRNA system has been established in many plant species, and the targeting efficiency and capacity of Cas9 has been improved by optimizing its expression and that of its gRNA. The CRISPR/Cas9 system can also be used for sequence-specific mutagenesis/integration and transcriptional control of target genes. We also discuss off-target effects and the constraint that the protospacer-adjacent motif (PAM) puts on CRISPR/Cas9 genome engineering. To address these problems, a number of bioinformatic tools are available to help design specific gRNAs, and new Cas9 variants and orthologs with high fidelity and alternative PAM specificities have been engineered. Owing to these recent efforts, the CRISPR/Cas9 system is becoming a revolutionary and flexible tool for genome engineering. Adoption of the CRISPR/Cas9 technology in plant research would enable the investigation of plant biology at an unprecedented depth and create innovative applications in precise crop breeding. PMID:27252719

  13. The Landscape of A-to-I RNA Editome Is Shaped by Both Positive and Purifying Selection

    Science.gov (United States)

    Kong, Yimeng; Pan, Bohu; Chen, Longxian; Wang, Hongbing; Hao, Pei; Li, Xuan

    2016-01-01

    The hydrolytic deamination of adenosine to inosine (A-to-I editing) in precursor mRNA induces variable gene products at the post-transcription level. How and to what extent A-to-I RNA editing diversifies transcriptome is not fully characterized in the evolution, and very little is known about the selective constraints that drive the evolution of RNA editing events. Here we present a study on A-to-I RNA editing, by generating a global profile of A-to-I editing for a phylogeny of seven Drosophila species, a model system spanning an evolutionary timeframe of approximately 45 million years. Of totally 9281 editing events identified, 5150 (55.5%) are located in the coding sequences (CDS) of 2734 genes. Phylogenetic analysis places these genes into 1,526 homologous families, about 5% of total gene families in the fly lineages. Based on conservation of the editing sites, the editing events in CDS are categorized into three distinct types, representing events on singleton genes (type I), and events not conserved (type II) or conserved (type III) within multi-gene families. While both type I and II events are subject to purifying selection, notably type III events are positively selected, and highly enriched in the components and functions of the nervous system. The tissue profiles are documented for three editing types, and their critical roles are further implicated by their shifting patterns during holometabolous development and in post-mating response. In conclusion, three A-to-I RNA editing types are found to have distinct evolutionary dynamics. It appears that nervous system functions are mainly tested to determine if an A-to-I editing is beneficial for an organism. The coding plasticity enabled by A-to-I editing creates a new class of binary variations, which is a superior alternative to maintain heterozygosity of expressed genes in a diploid mating system. PMID:27467689

  14. The emerging role of viral vectors as vehicles for DMD gene editing.

    Science.gov (United States)

    Maggio, Ignazio; Chen, Xiaoyu; Gonçalves, Manuel A F V

    2016-01-01

    Duchenne muscular dystrophy (DMD) is a genetic disorder caused by mutations in the dystrophin-encoding DMD gene. The DMD gene, spanning over 2.4 megabases along the short arm of the X chromosome (Xp21.2), is the largest genetic locus known in the human genome. The size of DMD, combined with the complexity of the DMD phenotype and the extent of the affected tissues, begs for the development of novel, ideally complementary, therapeutic approaches. Genome editing based on the delivery of sequence-specific programmable nucleases into dystrophin-defective cells has recently enriched the portfolio of potential therapies under investigation. Experiments involving different programmable nuclease platforms and target cell types have established that the application of genome-editing principles to the targeted manipulation of defective DMD loci can result in the rescue of dystrophin protein synthesis in gene-edited cells. Looking towards translation into the clinic, these proof-of-principle experiments have been swiftly followed by the conversion of well-established viral vector systems into delivery agents for DMD editing. These gene-editing tools consist of zinc-finger nucleases (ZFNs), engineered homing endoculeases (HEs), transcription activator-like effector nucleases (TALENs), and RNA-guided nucleases (RGNs) based on clustered, regularly interspaced, short palindromic repeats (CRISPR)-Cas9 systems. Here, we succinctly review these fast-paced developments and technologies, highlighting their relative merits and potential bottlenecks, when used as part of in vivo and ex vivo gene-editing strategies. PMID:27215286

  15. Quantifying Genome-Editing Outcomes at Endogenous Loci with SMRT Sequencing

    Directory of Open Access Journals (Sweden)

    Ayal Hendel

    2014-04-01

    Full Text Available Targeted genome editing with engineered nucleases has transformed the ability to introduce precise sequence modifications at almost any site within the genome. A major obstacle to probing the efficiency and consequences of genome editing is that no existing method enables the frequency of different editing events to be simultaneously measured across a cell population at any endogenous genomic locus. We have developed a method for quantifying individual genome-editing outcomes at any site of interest with single-molecule real-time (SMRT DNA sequencing. We show that this approach can be applied at various loci using multiple engineered nuclease platforms, including transcription-activator-like effector nucleases (TALENs, RNA-guided endonucleases (CRISPR/Cas9, and zinc finger nucleases (ZFNs, and in different cell lines to identify conditions and strategies in which the desired engineering outcome has occurred. This approach offers a technique for studying double-strand break repair, facilitates the evaluation of gene-editing technologies, and permits sensitive quantification of editing outcomes in almost every experimental system used.

  16. Photography, production, design and editing

    OpenAIRE

    Sánchez-Biosca, Vicente

    2013-01-01

    The formal features that have made Spanish cinema what it is and the technical processes behind them (camerawork, production design, editing) have passed through the same stages of trial and error, hurdles, crises, and conflicts as has been the case with other national cinemas. The notion of the exceptionalism of Spanish cinema is not supported by evidence, unless by “anomaly” we just mean “cultural specificity.” Cultural specificities there have been, as there are today: the h...

  17. Edition des Corpus areopagiticum slavicum

    Directory of Open Access Journals (Sweden)

    Dieter Fahl

    2005-12-01

    Full Text Available An Edition of the Corpus areopagiticum slavicum In the fourteenth century, the monk Isaiah of the holy Mount Athos translated the writings of pseudo-Dionysius the Areopagite (c. end of the 5th century, core texts for Eastern and Western European theological and philosophical thought, from Greek into Church Slavonic. This first Slavic translation of Dionysius’ oeuvre (“De Coelesti Hierarchia,” “De Ecclesiastica Hierarchia,” “De Divinis Nominibus,” “De Mystica Theologia,” the epistles and scholia, which played a significant role in the development of Slavic culture, Orthodox Slavic socio-political theory and praxis, is still central to the study of Slavia Orthodoxa. A working group of German and Russian scholars has completed an edition of the translator’s Church Slavonic autograph with an en face reconstruction of the Greek text used by the translator and philological commentary. A Church Slavonic-Greek and Greek-Church Slavonic dictionary of this edition, currently in preparation, plans to make the terminology used in this influential translation accessible to interdisciplinary researchers. For the first time, the Church Slavonic lexica of this corpus, a substantial part of which was coined by the translator, will be registered in an index of words and forms.

  18. Short RNA guides cleavage by eukaryotic RNase III.

    Directory of Open Access Journals (Sweden)

    Bruno Lamontagne

    Full Text Available In eukaryotes, short RNAs guide a variety of enzymatic activities that range from RNA editing to translation repression. It is hypothesized that pre-existing proteins evolved to bind and use guide RNA during evolution. However, the capacity of modern proteins to adopt new RNA guides has never been demonstrated. Here we show that Rnt1p, the yeast orthologue of the bacterial dsRNA-specific RNase III, can bind short RNA transcripts and use them as guides for sequence-specific cleavage. Target cleavage occurred at a constant distance from the Rnt1p binding site, leaving the guide RNA intact for subsequent cleavage. Our results indicate that RNase III may trigger sequence-specific RNA degradation independent of the RNAi machinery, and they open the road for a new generation of precise RNA silencing tools that do not trigger a dsRNA-mediated immune response.

  19. Editing of mouse and human immunoglobulin genes by CRISPR-Cas9 system

    Science.gov (United States)

    Cheong, Taek-Chin; Compagno, Mara; Chiarle, Roberto

    2016-01-01

    Applications of the CRISPR-Cas9 system to edit the genome have widely expanded to include DNA gene knock-out, deletions, chromosomal rearrangements, RNA editing and genome-wide screenings. Here we show the application of CRISPR-Cas9 technology to edit the mouse and human immunoglobulin (Ig) genes. By delivering Cas9 and guide-RNA (gRNA) with retro- or lenti-virus to IgM+ mouse B cells and hybridomas, we induce class-switch recombination (CSR) of the IgH chain to the desired subclass. Similarly, we induce CSR in all human B cell lines tested with high efficiency to targeted IgH subclass. Finally, we engineer mouse hybridomas to secrete Fab′ fragments instead of the whole Ig. Our results indicate that Ig genes in mouse and human cells can be edited to obtain any desired IgH switching helpful to study the biology of normal and lymphoma B cells. We also propose applications that could transform the technology of antibody production. PMID:26956543

  20. Editing of mouse and human immunoglobulin genes by CRISPR-Cas9 system.

    Science.gov (United States)

    Cheong, Taek-Chin; Compagno, Mara; Chiarle, Roberto

    2016-01-01

    Applications of the CRISPR-Cas9 system to edit the genome have widely expanded to include DNA gene knock-out, deletions, chromosomal rearrangements, RNA editing and genome-wide screenings. Here we show the application of CRISPR-Cas9 technology to edit the mouse and human immunoglobulin (Ig) genes. By delivering Cas9 and guide-RNA (gRNA) with retro- or lenti-virus to IgM(+) mouse B cells and hybridomas, we induce class-switch recombination (CSR) of the IgH chain to the desired subclass. Similarly, we induce CSR in all human B cell lines tested with high efficiency to targeted IgH subclass. Finally, we engineer mouse hybridomas to secrete Fab' fragments instead of the whole Ig. Our results indicate that Ig genes in mouse and human cells can be edited to obtain any desired IgH switching helpful to study the biology of normal and lymphoma B cells. We also propose applications that could transform the technology of antibody production. PMID:26956543

  1. Application of the genome editing tool CRISPR/Cas9 in non-human primates

    Science.gov (United States)

    LUO, Xin; LI, Min; SU, Bing

    2016-01-01

    In the past three years, RNA-guided Cas9 nuclease from the microbial clustered regularly interspaced short palindromic repeats (CRISPR) adaptive immune system has been used to facilitate efficient genome editing in many model and non-model animals. However, its application in nonhuman primates is still at the early stage, though in view of the similarities in anatomy, physiology, behavior and genetics, closely related nonhuman primates serve as optimal models for human biology and disease studies. In this review, we summarize the current proceedings of gene editing using CRISPR/Cas9 in nonhuman primates. PMID:27469252

  2. Connectivity editing for quad-dominant meshes

    KAUST Repository

    Peng, Chihan

    2013-08-01

    We propose a connectivity editing framework for quad-dominant meshes. In our framework, the user can edit the mesh connectivity to control the location, type, and number of irregular vertices (with more or fewer than four neighbors) and irregular faces (non-quads). We provide a theoretical analysis of the problem, discuss what edits are possible and impossible, and describe how to implement an editing framework that realizes all possible editing operations. In the results, we show example edits and illustrate the advantages and disadvantages of different strategies for quad-dominant mesh design. © 2013 The Author(s) Computer Graphics Forum © 2013 The Eurographics Association and John Wiley & Sons Ltd.

  3. Edit Distance between Unlabeled Ordered Trees

    OpenAIRE

    Micheli, Anne; Rossin, Dominique

    2005-01-01

    There exists a bijection between one stack sortable permutations --permutations which avoid the pattern $231$-- and planar trees. We define an edit distance between permutations which is coherent with the standard edit distance between trees. This one-to-one correspondence yields a polynomial algorithm for the subpermutation problem for $(231)$ avoiding permutations. Moreover, we obtain the generating function of the edit distance between ordered trees and some special ones. For the general c...

  4. Electronic editions – possibilities, limitations, perspectives

    OpenAIRE

    Mertens, Martin

    2010-01-01

    The thesis "Electronic Editions – Possibilities, Limitations, Perspectives" presents concepts and technologies for electronic editions in the field of German language and literature. The thesis focuses on describing forms of information contained by written texts that extend beyond the literal information content, and on the possibilities that exist for encoding this information in electronic editions and thus opening it up to electronic evaluation. It begins with a survey of existing electro...

  5. Operational Transformation In Co-Operative Editing

    Directory of Open Access Journals (Sweden)

    Mandeep Kaur

    2015-08-01

    Full Text Available Cooperative Editing Systems in real-time allows a virtual team to view and edit a shared document at the same time. The document shared must be synchronized in order to ensure consistency for all the participants. This paper describes the Operational Transformation the evolution of its techniques its various applications major issues and achievements. In addition this paper will present working of a platform where two users can edit a code programming file at the same time.

  6. Endonucleases: new tools to edit the mouse genome.

    Science.gov (United States)

    Wijshake, Tobias; Baker, Darren J; van de Sluis, Bart

    2014-10-01

    Mouse transgenesis has been instrumental in determining the function of genes in the pathophysiology of human diseases and modification of genes by homologous recombination in mouse embryonic stem cells remains a widely used technology. However, this approach harbors a number of disadvantages, as it is time-consuming and quite laborious. Over the last decade a number of new genome editing technologies have been developed, including zinc finger nucleases (ZFNs), transcription activator-like effector nucleases (TALENs) and clustered regularly interspaced short palindromic repeats/CRISPR-associated (CRISPR/Cas). These systems are characterized by a designed DNA binding protein or RNA sequence fused or co-expressed with a non-specific endonuclease, respectively. The engineered DNA binding protein or RNA sequence guides the nuclease to a specific target sequence in the genome to induce a double strand break. The subsequent activation of the DNA repair machinery then enables the introduction of gene modifications at the target site, such as gene disruption, correction or insertion. Nuclease-mediated genome editing has numerous advantages over conventional gene targeting, including increased efficiency in gene editing, reduced generation time of mutant mice, and the ability to mutagenize multiple genes simultaneously. Although nuclease-driven modifications in the genome are a powerful tool to generate mutant mice, there are concerns about off-target cleavage, especially when using the CRISPR/Cas system. Here, we describe the basic principles of these new strategies in mouse genome manipulation, their inherent advantages, and their potential disadvantages compared to current technologies used to study gene function in mouse models. This article is part of a Special Issue entitled: From Genome to Function. PMID:24794718

  7. Potential pitfalls of CRISPR/Cas9-mediated genome editing.

    Science.gov (United States)

    Peng, Rongxue; Lin, Guigao; Li, Jinming

    2016-04-01

    Recently, a novel technique named the clustered regularly interspaced short palindromic repeat (CRISPR)/CRISPR-associated protein (Cas)9 system has been rapidly developed. This genome editing tool has improved our ability tremendously with respect to exploring the pathogenesis of diseases and correcting disease mutations, as well as phenotypes. With a short guide RNA, Cas9 can be precisely directed to target sites, and functions as an endonuclease to efficiently produce breaks in DNA double strands. Over the past 30 years, CRISPR has evolved from the 'curious sequences of unknown biological function' into a promising genome editing tool. As a result of the incessant development in the CRISPR/Cas9 system, Cas9 co-expressed with custom guide RNAs has been successfully used in a variety of cells and organisms. This genome editing technology can also be applied to synthetic biology, functional genomic screening, transcriptional modulation and gene therapy. However, although CRISPR/Cas9 has a broad range of action in science, there are several aspects that affect its efficiency and specificity, including Cas9 activity, target site selection and short guide RNA design, delivery methods, off-target effects and the incidence of homology-directed repair. In the present review, we highlight the factors that affect the utilization of CRISPR/Cas9, as well as possible strategies for handling any problems. Addressing these issues will allow us to take better advantage of this technique. In addition, we also review the history and rapid development of the CRISPR/Cas system from the time of its initial discovery in 2012. PMID:26535798

  8. Versatility of chemically synthesized guide RNAs for CRISPR-Cas9 genome editing.

    Science.gov (United States)

    Kelley, Melissa L; Strezoska, Žaklina; He, Kaizhang; Vermeulen, Annaleen; Smith, Anja van Brabant

    2016-09-10

    The CRISPR-Cas9 system has become the most popular and efficient method for genome engineering in mammalian cells. The Streptococcus pyogenes Cas9 nuclease can function with two types of guide RNAs: the native dual crRNA and tracrRNA (crRNA:tracrRNA) or a chimeric single guide RNA (sgRNA). Although sgRNAs expressed from a DNA vector are predominant in the literature, guide RNAs can be rapidly generated by chemical synthesis and provide equivalent functionality in gene editing experiments. This review highlights the attributes and advantages of chemically synthesized guide RNAs including the incorporation of chemical modifications to enhance gene editing efficiencies in certain applications. The use of synthetic guide RNAs is also uniquely suited to genome-scale high throughput arrayed screening, particularly when using complex phenotypic assays for functional genomics studies. Finally, the use of synthetic guide RNAs along with DNA-free sources of Cas9 (mRNA or protein) allows for transient CRISPR-Cas9 presence in the cell, thereby resulting in a decreased probability of off-target events. PMID:27374403

  9. Evaluation of microRNA alignment techniques.

    Science.gov (United States)

    Ziemann, Mark; Kaspi, Antony; El-Osta, Assam

    2016-08-01

    Genomic alignment of small RNA (smRNA) sequences such as microRNAs poses considerable challenges due to their short length (∼21 nucleotides [nt]) as well as the large size and complexity of plant and animal genomes. While several tools have been developed for high-throughput mapping of longer mRNA-seq reads (>30 nt), there are few that are specifically designed for mapping of smRNA reads including microRNAs. The accuracy of these mappers has not been systematically determined in the case of smRNA-seq. In addition, it is unknown whether these aligners accurately map smRNA reads containing sequence errors and polymorphisms. By using simulated read sets, we determine the alignment sensitivity and accuracy of 16 short-read mappers and quantify their robustness to mismatches, indels, and nontemplated nucleotide additions. These were explored in the context of a plant genome (Oryza sativa, ∼500 Mbp) and a mammalian genome (Homo sapiens, ∼3.1 Gbp). Analysis of simulated and real smRNA-seq data demonstrates that mapper selection impacts differential expression results and interpretation. These results will inform on best practice for smRNA mapping and enable more accurate smRNA detection and quantification of expression and RNA editing. PMID:27284164

  10. Fuel Cell Handbook, Fourth Edition

    Energy Technology Data Exchange (ETDEWEB)

    Stauffer, D.B; Hirschenhofer, J.H.; Klett, M.G.; Engleman, R.R.

    1998-11-01

    Robust progress has been made in fuel cell technology since the previous edition of the Fuel Cell Handbook was published in January 1994. This Handbook provides a foundation in fuel cells for persons wanting a better understanding of the technology, its benefits, and the systems issues that influence its application. Trends in technology are discussed, including next-generation concepts that promise ultra high efficiency and low cost, while providing exceptionally clean power plant systems. Section 1 summarizes fuel cell progress since the last edition and includes existing power plant nameplate data. Section 2 addresses the thermodynamics of fuel cells to provide an understanding of fuel cell operation at two levels (basic and advanced). Sections 3 through 6 describe the four major fuel cell types and their performance based on cell operating conditions. The section on polymer electrolyte membrane fuel cells has been added to reflect their emergence as a significant fuel cell technology. Phosphoric acid, molten carbonate, and solid oxide fuel cell technology description sections have been updated from the previous edition. New information indicates that manufacturers have stayed with proven cell designs, focusing instead on advancing the system surrounding the fuel cell to lower life cycle costs. Section 7, Fuel Cell Systems, has been significantly revised to characterize near-term and next-generation fuel cell power plant systems at a conceptual level of detail. Section 8 provides examples of practical fuel cell system calculations. A list of fuel cell URLs is included in the Appendix. A new index assists the reader in locating specific information quickly.

  11. GPU Computing Gems Emerald Edition

    CERN Document Server

    Hwu, Wen-mei W

    2011-01-01

    ".the perfect companion to Programming Massively Parallel Processors by Hwu & Kirk." -Nicolas Pinto, Research Scientist at Harvard & MIT, NVIDIA Fellow 2009-2010 Graphics processing units (GPUs) can do much more than render graphics. Scientists and researchers increasingly look to GPUs to improve the efficiency and performance of computationally-intensive experiments across a range of disciplines. GPU Computing Gems: Emerald Edition brings their techniques to you, showcasing GPU-based solutions including: Black hole simulations with CUDA GPU-accelerated computation and interactive display of

  12. 6. Electronic Editions for Everyone

    OpenAIRE

    Robinson, Peter

    2013-01-01

    1. Books Defy the Digital Revolution In January 2004, I gave a lecture on electronic scholarly editing at the University of Virginia. At the beginning of the lecture I asked the audience, of around 60 people, three questions. The first question was: who among them had bought a movie on DVD in the last year; who had bought a piece of music on CD-ROM or by download in the last year; who had taken digital photographs? Almost everyone in the audience had done all three. The second question: who i...

  13. Lentivirus pre-packed with Cas9 protein for safer gene editing.

    Science.gov (United States)

    Choi, J G; Dang, Y; Abraham, S; Ma, H; Zhang, J; Guo, H; Cai, Y; Mikkelsen, J G; Wu, H; Shankar, P; Manjunath, N

    2016-07-01

    The CRISPR/Cas9 system provides an easy way to edit specific site/s in the genome and thus offers tremendous opportunity for human gene therapy for a wide range of diseases. However, one major concern is off-target effects, particularly with long-term expression of Cas9 nuclease when traditional expression methods such as via plasmid/viral vectors are used. To overcome this limitation, we pre-packaged Cas9 protein (Cas9P LV) in lentiviral particles for transient exposure and showed its effectiveness for gene disruption in cells, including primary T cells expressing specific single guide RNAs (sgRNAs). We then constructed an 'all in one virus' to express sgRNAs in association with pre-packaged Cas9 protein (sgRNA/Cas9P LV). We successfully edited CCR5 in TZM-bl cells by this approach. Using an sgRNA-targeting HIV long terminal repeat, we also were able to disrupt HIV provirus in the J-LAT model of viral latency. Moreover, we also found that pre-packaging Cas9 protein in LV particle reduced off-target editing of chromosome 4:-29134166 locus by CCR5 sgRNA, compared with continued expression from the vector. These results show that sgRNA/Cas9P LV can be used as a safer approach for human gene therapy applications. PMID:27052803

  14. CRISPR/Cas9-Mediated Genome Editing in Soybean Hairy Roots.

    Directory of Open Access Journals (Sweden)

    Yupeng Cai

    Full Text Available As a new technology for gene editing, the CRISPR (clustered regularly interspaced short palindromic repeat/Cas (CRISPR-associated system has been rapidly and widely used for genome engineering in various organisms. In the present study, we successfully applied type II CRISPR/Cas9 system to generate and estimate genome editing in the desired target genes in soybean (Glycine max (L. Merrill.. The single-guide RNA (sgRNA and Cas9 cassettes were assembled on one vector to improve transformation efficiency, and we designed a sgRNA that targeted a transgene (bar and six sgRNAs that targeted different sites of two endogenous soybean genes (GmFEI2 and GmSHR. The targeted DNA mutations were detected in soybean hairy roots. The results demonstrated that this customized CRISPR/Cas9 system shared the same efficiency for both endogenous and exogenous genes in soybean hairy roots. We also performed experiments to detect the potential of CRISPR/Cas9 system to simultaneously edit two endogenous soybean genes using only one customized sgRNA. Overall, generating and detecting the CRISPR/Cas9-mediated genome modifications in target genes of soybean hairy roots could rapidly assess the efficiency of each target loci. The target sites with higher efficiencies can be used for regular soybean transformation. Furthermore, this method provides a powerful tool for root-specific functional genomics studies in soybean.

  15. Site-specific genome editing for correction of induced pluripotent stem cells derived from dominant dystrophic epidermolysis bullosa.

    Science.gov (United States)

    Shinkuma, Satoru; Guo, Zongyou; Christiano, Angela M

    2016-05-17

    Genome editing with engineered site-specific endonucleases involves nonhomologous end-joining, leading to reading frame disruption. The approach is applicable to dominant negative disorders, which can be treated simply by knocking out the mutant allele, while leaving the normal allele intact. We applied this strategy to dominant dystrophic epidermolysis bullosa (DDEB), which is caused by a dominant negative mutation in the COL7A1 gene encoding type VII collagen (COL7). We performed genome editing with TALENs and CRISPR/Cas9 targeting the mutation, c.8068_8084delinsGA. We then cotransfected Cas9 and guide RNA expression vectors expressed with GFP and DsRed, respectively, into induced pluripotent stem cells (iPSCs) generated from DDEB fibroblasts. After sorting, 90% of the iPSCs were edited, and we selected four gene-edited iPSC lines for further study. These iPSCs were differentiated into keratinocytes and fibroblasts secreting COL7. RT-PCR and Western blot analyses revealed gene-edited COL7 with frameshift mutations degraded at the protein level. In addition, we confirmed that the gene-edited truncated COL7 could neither associate with normal COL7 nor undergo triple helix formation. Our data establish the feasibility of mutation site-specific genome editing in dominant negative disorders. PMID:27143720

  16. Origin and evolution of a placental-specific microRNA family in the human genome

    Directory of Open Access Journals (Sweden)

    Gong Lejun

    2010-11-01

    Full Text Available Abstract Background MicroRNAs (miRNAs are a class of short regulatory RNAs encoded in the genome of DNA viruses, some single cell organisms, plants and animals. With the rapid development of technology, more and more miRNAs are being discovered. However, the origin and evolution of most miRNAs remain obscure. Here we report the origin and evolution dynamics of a human miRNA family. Results We have shown that all members of the miR-1302 family are derived from MER53 elements. Although the conservation scores of the MER53-derived pre-miRNA sequences are low, we have identified 36 potential paralogs of MER53-derived miR-1302 genes in the human genome and 58 potential orthologs of the human miR-1302 family in placental mammals. We suggest that in placental species, this miRNA family has evolved following the birth-and-death model of evolution. Three possible mechanisms that can mediate miRNA duplication in evolutionary history have been proposed: the transposition of the MER53 element, segmental duplications and Alu-mediated recombination. Finally, we have found that the target genes of miR-1302 are over-represented in transportation, localization, and system development processes and in the positive regulation of cellular processes. Many of them are predicted to function in binding and transcription regulation. Conclusions The members of miR-1302 family that are derived from MER53 elements are placental-specific miRNAs. They emerged at the early stage of the recent 180 million years since eutherian mammals diverged from marsupials. Under the birth-and-death model, the miR-1302 genes have experienced a complex expansion with some members evolving by segmental duplications and some by Alu-mediated recombination events.

  17. Kinetoplastid guide RNA biogenesis is dependent on subunits of the mitochondrial RNA binding complex 1 and mitochondrial RNA polymerase

    Czech Academy of Sciences Publication Activity Database

    Hashimi, Hassan; Číčová, Zdeňka; Novotná, Lucie; Wen, Y.-Z.; Lukeš, Julius

    2009-01-01

    Roč. 15, č. 4 (2009), s. 588-599. ISSN 1355-8382 R&D Projects: GA ČR GA204/09/1667; GA AV ČR IAA500960705; GA MŠk LC07032; GA MŠk 2B06129; GA ČR GD524/03/H133 Institutional research plan: CEZ:AV0Z60220518 Keywords : RNA editing * guide RNA * mitochondrion * trypanosome Subject RIV: EB - Genetics ; Molecular Biology Impact factor: 5.198, year: 2009

  18. Sex Differences in Cognitive Abilities. Fourth Edition

    Science.gov (United States)

    Halpern, Diane F.

    2011-01-01

    The fourth edition of "Sex Differences in Cognitive Abilities" critically examines the breadth of research on this complex and controversial topic, with the principal aim of helping the reader to understand where sex differences are found--and where they are not. Since the publication of the third edition, there have been many exciting and…

  19. Applications of genome editing in insects

    Science.gov (United States)

    Insect genome editing was first reported 1991 in Drosophila melanogaster but the technology used was not portable to other species. Not until the recent development of facile, engineered DNA endonuclease systems has gene editing become widely available to insect scientists. Most applications in inse...

  20. Editing Technical Proposals (The Friendly Editor).

    Science.gov (United States)

    Bush, Don

    1994-01-01

    Makes suggestions for editing technical proposals. Discusses the marketeers, the hierarchy of hype, how to save days, managing story boards, expediting a laborious process, teaching engineers to write, writing incrementally, the art group, and the editing task. Argues that the best proposals come from starting to write early. (SR)

  1. Fusion Canada issue 32. Final edition

    International Nuclear Information System (INIS)

    Fusion Canada is a bulletin of the National Fusion Program, this is the last edition. Included in this July edition are articles on Funding for Canada's fusion program, Research and Development on TdeV-96 , Divertor Maintenance Robotics and reference listing for Canada's Fusion research and development sites

  2. Fuel Cell Handbook, Fifth Edition

    Energy Technology Data Exchange (ETDEWEB)

    Energy and Environmental Solutions

    2000-10-31

    Progress continues in fuel cell technology since the previous edition of the Fuel Cell Handbook was published in November 1998. Uppermost, polymer electrolyte fuel cells, molten carbonate fuel cells, and solid oxide fuel cells have been demonstrated at commercial size in power plants. The previously demonstrated phosphoric acid fuel cells have entered the marketplace with more than 220 power plants delivered. Highlighting this commercial entry, the phosphoric acid power plant fleet has demonstrated 95+% availability and several units have passed 40,000 hours of operation. One unit has operated over 49,000 hours. Early expectations of very low emissions and relatively high efficiencies have been met in power plants with each type of fuel cell. Fuel flexibility has been demonstrated using natural gas, propane, landfill gas, anaerobic digester gas, military logistic fuels, and coal gas, greatly expanding market opportunities. Transportation markets worldwide have shown remarkable interest in fuel cells; nearly every major vehicle manufacturer in the U.S., Europe, and the Far East is supporting development. This Handbook provides a foundation in fuel cells for persons wanting a better understanding of the technology, its benefits, and the systems issues that influence its application. Trends in technology are discussed, including next-generation concepts that promise ultrahigh efficiency and low cost, while providing exceptionally clean power plant systems. Section 1 summarizes fuel cell progress since the last edition and includes existing power plant nameplate data. Section 2 addresses the thermodynamics of fuel cells to provide an understanding of fuel cell operation at two levels (basic and advanced). Sections 3 through 8 describe the six major fuel cell types and their performance based on cell operating conditions. Alkaline and intermediate solid state fuel cells were added to this edition of the Handbook. New information indicates that manufacturers have stayed

  3. Endonuclease V cleaves at inosines in RNA.

    Science.gov (United States)

    Vik, Erik Sebastian; Nawaz, Meh Sameen; Strøm Andersen, Pernille; Fladeby, Cathrine; Bjørås, Magnar; Dalhus, Bjørn; Alseth, Ingrun

    2013-01-01

    Endonuclease V orthologues are highly conserved proteins found in all kingdoms of life. While the prokaryotic enzymes are DNA repair proteins for removal of deaminated adenosine (inosine) from the genome, no clear role for the eukaryotic counterparts has hitherto been described. Here we report that human endonuclease V (ENDOV) and also Escherichia coli endonuclease V are highly active ribonucleases specific for inosine in RNA. Inosines are normal residues in certain RNAs introduced by specific deaminases. Adenosine-to-inosine editing is essential for proper function of these transcripts and defects are linked to various human disease. Here we show that human ENDOV cleaves an RNA substrate containing inosine in a position corresponding to a biologically important site for deamination in the Gabra-3 transcript of the GABA(A) neurotransmitter. Further, human ENDOV specifically incises transfer RNAs with inosine in the wobble position. This previously unknown RNA incision activity may suggest a role for endonuclease V in normal RNA metabolism. PMID:23912683

  4. Branched DNA-based Alu quantitative assay for cell-free plasma DNA levels in patients with sepsis or systemic inflammatory response syndrome.

    Science.gov (United States)

    Hou, Yan-Qiang; Liang, Dong-Yu; Lou, Xiao-Li; Zhang, Mei; Zhang, Zhen-huan; Zhang, Lu-rong

    2016-02-01

    Cell-free circulating DNA (cf-DNA) can be detected by various of laboratory techniques. We described a branched DNA-based Alu assay for measuring cf-DNA in septic patients. Compared to healthy controls and systemic inflammatory response syndrome (SIRS) patients, serum cf-DNA levels were significantly higher in septic patients (1426.54 ± 863.79 vs 692.02 ± 703.06 and 69.66 ± 24.66 ng/mL). The areas under the receiver operating characteristic curve of cf-DNA for normal vs sepsis and SIRS vs sepsis were 0.955 (0.884-1.025), and 0.856 (0.749-0.929), respectively. There was a positive correlation between cf-DNA and interleukin 6 or procalcitonin or Acute Physiology and Chronic Health Evaluation II. The cf-DNA concentration was higher in intensive care unit nonsurviving patients compared to surviving patients (2183.33 ± 615.26 vs 972.46 ± 648.36 ng/mL; P format. Cell-free circulating DNA might be a new marker in discrimination of sepsis and SIRS. PMID:26589770

  5. Identification of a 4.9-kilo base-pair Alu-mediated founder SDHD deletion in two extended paraganglioma families from Austria.

    Science.gov (United States)

    Janecke, Andreas R; Willett-Brozick, Joan E; Karas, Christoph; Hasipek, Metis; Loeffler-Ragg, Judith; Baysal, Bora E

    2010-03-01

    Hereditary paraganglioma (PGL) is characterized by the development of highly vascularized paraganglionic tumors as a result of germline mutations in the SDHB, SDHC or SDHD subunit genes of succinate dehydrogenase (SDH; mitochondrial complex II), or in the Von Hippel-Lindau tumor-suppressor gene. Although many PGL mutations have been described, gross SDHD deletions have not yet been implicated as founder mutations and are rarely characterized at the DNA sequence level. We investigated the genetic basis of head and neck PGLs observed in 20 subjects from two unrelated multiplex pedigrees from Austria and identified a 4944-base pair partial SDHD deletion, which escaped PCR-based detection methods. The deletion occurred between Alu elements and was present within the same haplotype context in both pedigrees, indicating a founder effect. The deletion caused tumors only after a paternal transmission similar to other conventional SDHD mutations, suggesting preservation of genomic imprinting mechanisms operating at this locus. These data describe a large SDHD deletion at the genomic sequence level and indicate that gross SDHD deletions could be a founder PGL mutation in certain populations. PMID:20111059

  6. Partial protoporphyrinogen oxidase (PPOX gene deletions, due to different Alu-mediated mechanisms, identified by MLPA analysis in patients with variegate porphyria

    Directory of Open Access Journals (Sweden)

    Barbaro Michela

    2013-01-01

    Full Text Available Abstract Variegate porphyria (VP is an autosomal dominantly inherited hepatic porphyria. The genetic defect in the PPOX gene leads to a partial defect of protoporphyrinogen oxidase, the penultimate enzyme of heme biosynthesis. Affected individuals can develop cutaneous symptoms in sun-exposed areas of the skin and/or neuropsychiatric acute attacks. The identification of the genetic defect in VP families is of crucial importance to detect the carrier status which allows counseling to prevent potentially life threatening neurovisceral attacks, usually triggered by factors such as certain drugs, alcohol or fasting. In a total of 31 Swedish VP families sequence analysis had identified a genetic defect in 26. In the remaining five families an extended genetic investigation was necessary. After the development of a synthetic probe set, MLPA analysis to screen for single exon deletions/duplications was performed. We describe here, for the first time, two partial deletions within the PPOX gene detected by MLPA analysis. One deletion affects exon 5 and 6 (c.339-197_616+320del1099 and has been identified in four families, most probably after a founder effect. The other extends from exon 5 to exon 9 (c.339-350_987+229del2609 and was found in one family. We show that both deletions are mediated by Alu repeats. Our findings emphasize the usefulness of MLPA analysis as a complement to PPOX gene sequencing analysis for comprehensive genetic diagnostics in patients with VP.

  7. Exploiting CRISPR-Cas immune systems for genome editing in bacteria.

    Science.gov (United States)

    Barrangou, Rodolphe; van Pijkeren, Jan-Peter

    2016-02-01

    The CRISPR-Cas immune system is a DNA-encoded, RNA-mediated, DNA-targeting defense mechanism, which provides sequence-specific targeting of DNA. This molecular machinery can be engineered into the sgRNA:Cas9 technology, for programmable cleavage of DNA. Following the genesis of double-stranded DNA breaks, the DNA repair machinery generates mutations at the cleavage site using various pathways. This technology has revolutionized eukaryotic genome editing, and we are at the cusp of full exploitation in bacteria. Here, we discuss the potential of CRISPR-based technologies for use in bacteria, and highlight the application of single stranded DNA recombineering combined with CRISPR-Cas selection to edit the genome of a probiotic organism. We envision that CRISPR-Cas technologies will play a key role in the development of next-generation industrial bacteria. PMID:26629846

  8. Targeted viral-mediated plant genome editing using crispr/cas9

    KAUST Repository

    Mahfouz, Magdy M.

    2015-12-17

    The present disclosure provides a viral-mediated genome-editing platform that facilitates multiplexing, obviates stable transformation, and is applicable across plant species. The RNA2 genome of the tobacco rattle virus (TRV) was engineered to carry and systemically deliver a guide RNA molecules into plants overexpressing Cas9 endonuclease. High genomic modification frequencies were observed in inoculated as well as systemic leaves including the plant growing points. This system facilitates multiplexing and can lead to germinal transmission of the genomic modifications in the progeny, thereby obviating the requirements of repeated transformations and tissue culture. The editing platform of the disclosure is useful in plant genome engineering and applicable across plant species amenable to viral infections for agricultural biotechnology applications.

  9. Transfer RNA-derived small RNAs in the cancer transcriptome

    OpenAIRE

    Green, Darrell; Fraser, William; Dalmay, Tamas

    2016-01-01

    The cellular lifetime includes stages such as differentiation, proliferation, division, senescence and apoptosis.These stages are driven by a strictly ordered process of transcription dynamics. Molecular disruption to RNA polymerase assembly, chromatin remodelling and transcription factor binding through to RNA editing, splicing, post-transcriptional regulation and ribosome scanning can result in significant costs arising from genome instability. Cancer development is one example of when such...

  10. AAO Observer - February 2011 Edition

    CERN Document Server

    Brough, Sarah

    2011-01-01

    This edition of the Australian Astronomical Observatory Observer contains articles on the detection of the baryonic acoustic oscillation signal over a wide range of redshifts by the WiggleZ dark energy survey; results and future plans for the Galaxy And Mass Assembly survey; an update on the HERMES instrument; a report on the use of the AAT as a testbed for experiments using a laser frequency comb to achieve ultra-precise calibration of spectrographs; an innovative project combining professional astronomers and highly professional 'amateurs' to discover and characterise planets orbiting binary stars and finding out what it's like to be present when the comet you discovered encounters a spacecraft! The AAO's Distinguished Visitors for 2011 are introduced and an announcement is made for the upcoming Southern Cross Conference on 'Supernovae and their Host Galaxies' to be held in Sydney in June 2011.

  11. AAO Observer - August 2011 Edition

    CERN Document Server

    Brough, Sarah

    2011-01-01

    This edition of the Australian Astronomical Observatory Observer contains articles on the commissioning of the new SAMI instrument giving the first hexabundle galaxy spectra; galaxy parameter variations across and through the 6dFGS Fundamental Plane; an introduction to the new Dragonfly stellar interferometer; an update on the RAdial VElocity (RAVE) survey at half a million spectra; the Magellanic Quasars Survey; the Integrated Photonic Spectrograph's first look at the heart of the Scorpion; using AAOMega to measure the age of the young open cluster IC2602; making MANIFEST fibres for the Giant Magellan Telescope and a Voyage through Filaments of Galaxies. The Observer also contains thoughts on diversity in the astronomy community and reports on the recent Supernovae and their Host Galaxies conference and the 2011 Science Meets Parliament. In addition there are the usual features of the AUSGO Corner, Epping News and Letter from Coona.

  12. Association between Alu insertion polymorphisms and HLA class T alleles in Chinese Lisu and Nu ethnic populations%中国傈僳族和怒族群体人类白细胞抗原Ⅰ类基因区Alu插入多态性研究

    Institute of Scientific and Technical Information of China (English)

    董兆梅; 姚宇峰; 史磊; 陶玉芬; 林克勤; 黄小琴; 杨昭庆; 褚嘉祐; 史荔

    2012-01-01

    Objective To investigate the frequencies of HLA-Alu repeat polymorphisms (AluMICB,AluTF,AluHJ,AluHG and AluHF) in Chinese Lisu and Nu ethnic populations.Methods The frequencies of HLA-Alu repeat polymorphisms in above populations were determined with polymerase chain reaction (PCR).The associations between HLA-Alu repeat polymorphisms and HLA-A,HLA-B and HLA-C alleles were also analyzed.Phylogenetic trees were constructed with genetic distance calculated from the frequencies of HLA-Alu repeat polymorphisms.Results Frequencies of AluTF * 2 and AluHF * 2 were different between the two populations (P<0.05),while those of other three insertions were similar.The strength of association between HLA-Alus and HLA alleles were different (P<0.05) in the two populations.Although AluMICB * 2 were associated with HLA-B* 56:01 in both populations,the association was stronger in Lisu population (74.0%) but moderate in Nu population (30.7%).HLA-Alus were associated with particular HLA subtypes,e.g.,AluHG * 2 with certain HLA-A * 02 subtypes.By phylogenetic analysis,Lisu and Nu were clustered together with southern Chinese and Thai populations.Conclusion The distribution of HLA-Alus and the strength of associations between HLA-Alus and HLA class I alleles have varied between the two populations.Study of this association may facilitate identification of origins,evolution,progenitor haplotypes and recombination within the HLA class I region.%目的 研究中国两个隔离群体(傈僳族和怒族)人类白细胞抗原(human leukocyte antigen,HLA)Ⅰ类基因区域内5个HLA-Alu插入多态性(AluMICB、AluTF、AluHJ、AluHG和AluHF)的分布特征.方法 应用聚合酶链反应技术对中国两个隔离群体傈僳族(107人)和怒族(104人)进行HLA-Alu多态性分型.结合HLA基因分型数据,分析这两个群体中HLA-Alu插入与HLA-A、HLA-B和HLA-C基因的关系.根据HLA-Alu频率计算各群体间遗传距离,构建系统进化树.结果 AluTF和AluHF插入

  13. Creation of a split-cas9 system for effective plant genome editing

    OpenAIRE

    Costa Carrió, Pedro

    2015-01-01

    Precise and efficient genome-targeting technologies are needed to enable both systematic genome editing and interrogation of gene function. Although technologies such as designer zinc fingers (ZFs), transcription activator-like effectors (TALENs), and homing meganucleases have begun to enable targeted genome modifications, there is a need for new technologies that are scalable, affordable, and easy to engineer. Easy programmability of the RNA-guided DNA-endonuclease Cas9 of the type II CRISPR...

  14. The Neisseria meningitidis CRISPR-Cas9 System Enables Specific Genome Editing in Mammalian Cells

    OpenAIRE

    Lee, Ciaran M; Cradick, Thomas J.; Bao, Gang

    2016-01-01

    The clustered regularly-interspaced short palindromic repeats (CRISPR)—CRISPR-associated (Cas) system from Streptococcus pyogenes (Spy) has been successfully adapted for RNA-guided genome editing in a wide range of organisms. However, numerous reports have indicated that Spy CRISPR-Cas9 systems may have significant off-target cleavage of genomic DNA sequences differing from the intended on-target site. Here, we report the performance of the Neisseria meningitidis (Nme) CRISPR-Cas9 system that...

  15. Efficient genome editing in Caenorhabditis elegans by CRISPR-targeted homologous recombination

    OpenAIRE

    Chen, C.; Fenk, L. A.; Bono, M.

    2013-01-01

    Cas9 is an RNA-guided double-stranded DNA nuclease that participates in clustered regularly interspaced short palindromic repeats (CRISPR)-mediated adaptive immunity in prokaryotes. CRISPR–Cas9 has recently been used to generate insertion and deletion mutations in Caenorhabditis elegans, but not to create tailored changes (knock-ins). We show that the CRISPR–CRISPR-associated (Cas) system can be adapted for efficient and precise editing of the C. elegans genome. The targeted double-strand bre...

  16. NRP 7th Edition: Are You Prepared?

    Science.gov (United States)

    Zaichkin, Jeanette; Mccarney, Linda; Weiner, Gary

    2016-01-01

    The seventh edition of the American Academy of Pediatrics/American Heart Association Neonatal Resuscitation Program (NRP) materials must be in use by January 1, 2017. As in previous editions, changes in resuscitation science are based on an international review and consensus of current resuscitation science. The seventh edition NRP materials also include enhancements to training materials aimed at improving the quality of NRP instruction and providing the opportunity for ongoing education. A standardized approach to instructor training, an online Instructor Toolkit, eSim cases, and a new learning management system are among the new resources. PMID:27461196

  17. Graph edit distance from spectral seriation

    OpenAIRE

    Robles-Kelly, A; Hancock, E.R.

    2005-01-01

    This paper is concerned with computing graph edit distance. One of the criticisms that can be leveled at existing methods for computing graph edit distance is that they lack some of the formality and rigor of the computation of string edit distance. Hence, our aim is to convert graphs to string sequences so that string matching techniques can be used. To do this, we use a graph spectral seriation method to convert the adjacency matrix into a string or sequence order. We show how the serial or...

  18. On the computation of edit distance functions

    OpenAIRE

    Martin, Ryan R.

    2010-01-01

    The edit distance between two graphs on the same labeled vertex set is the size of the symmetric difference of the edge sets. The edit distance function of the hereditary property, $\\mathcal{H}$, is a function of $p\\in[0,1]$ and is the limit of the maximum normalized distance between a graph of density $p$ and $\\mathcal{H}$. This paper uses the symmetrization method of Sidorenko in order to compute the edit distance function of various hereditary properties. For any graph $H$, ${\\rm Forb}(H)$...

  19. Optimization of CRISPR/Cas9 genome editing to modify abiotic stress responses in plants

    Science.gov (United States)

    Osakabe, Yuriko; Watanabe, Takahito; Sugano, Shigeo S; Ueta, Risa; Ishihara, Ryosuke; Shinozaki, Kazuo; Osakabe, Keishi

    2016-01-01

    Genome editing using the CRISPR/Cas9 system can be used to modify plant genomes, however, improvements in specificity and applicability are still needed in order for the editing technique to be useful in various plant species. Here, using genome editing mediated by a truncated gRNA (tru-gRNA)/Cas9 combination, we generated new alleles for OST2, a proton pump in Arabidopsis, with no off-target effects. By following expression of Cas9 and the tru-gRNAs, newly generated mutations in CRIPSR/Cas9 transgenic plants were detected with high average mutation rates of up to 32.8% and no off-target effects using constitutive promoter. Reducing nuclear localization signals in Cas9 decreased the mutation rate. In contrast, tru-gRNA Cas9 cassettes driven by meristematic- and reproductive-tissue-specific promoters increased the heritable mutation rate in Arabidopsis, showing that high expression in the germ line can produce bi-allelic mutations. Finally, the new mutant alleles obtained for OST2 exhibited altered stomatal closing in response to environmental conditions. These results suggest further applications in molecular breeding to improve plant function using optimized plant CRISPR/Cas9 systems. PMID:27226176

  20. Optimization of CRISPR/Cas9 genome editing to modify abiotic stress responses in plants.

    Science.gov (United States)

    Osakabe, Yuriko; Watanabe, Takahito; Sugano, Shigeo S; Ueta, Risa; Ishihara, Ryosuke; Shinozaki, Kazuo; Osakabe, Keishi

    2016-01-01

    Genome editing using the CRISPR/Cas9 system can be used to modify plant genomes, however, improvements in specificity and applicability are still needed in order for the editing technique to be useful in various plant species. Here, using genome editing mediated by a truncated gRNA (tru-gRNA)/Cas9 combination, we generated new alleles for OST2, a proton pump in Arabidopsis, with no off-target effects. By following expression of Cas9 and the tru-gRNAs, newly generated mutations in CRIPSR/Cas9 transgenic plants were detected with high average mutation rates of up to 32.8% and no off-target effects using constitutive promoter. Reducing nuclear localization signals in Cas9 decreased the mutation rate. In contrast, tru-gRNA Cas9 cassettes driven by meristematic- and reproductive-tissue-specific promoters increased the heritable mutation rate in Arabidopsis, showing that high expression in the germ line can produce bi-allelic mutations. Finally, the new mutant alleles obtained for OST2 exhibited altered stomatal closing in response to environmental conditions. These results suggest further applications in molecular breeding to improve plant function using optimized plant CRISPR/Cas9 systems. PMID:27226176

  1. Marketing/Planning Library and Information Services. Second Edition.

    Science.gov (United States)

    Weingand, Darlene E.

    In the first edition of this book, the concepts of marketing and planning library and information services were presented as effective managerial strategies. Several paragraphs from the introduction to the first edition are reproduced, with author commentary, in this edition as an affirmation that the message is still true. In this second edition,…

  2. CRISPR/Cas9 for plant genome editing: accomplishments, problems and prospects.

    Science.gov (United States)

    Paul, Joseph W; Qi, Yiping

    2016-07-01

    The increasing burden of the world population on agriculture requires the development of more robust crops. Dissecting the basic biology that underlies plant development and stress responses will inform the design of better crops. One powerful tool for studying plants at the molecular level is the RNA-programmed genome editing system composed of a clustered regularly interspaced short palindromic repeats (CRISPR)-encoded guide RNA and the nuclease Cas9. Here, some of the recent advances in CRISPR/Cas9 technology that have profound implications for improving the study of plant biology are described. These tools are also paving the way towards new horizons for biotechnologies and crop development. PMID:27114166

  3. Negative Feedback Regulation of HIV-1 by Gene Editing Strategy.

    Science.gov (United States)

    Kaminski, Rafal; Chen, Yilan; Salkind, Julian; Bella, Ramona; Young, Won-Bin; Ferrante, Pasquale; Karn, Jonathan; Malcolm, Thomas; Hu, Wenhui; Khalili, Kamel

    2016-01-01

    The CRISPR/Cas9 gene editing method is comprised of the guide RNA (gRNA) to target a specific DNA sequence for cleavage and the Cas9 endonuclease for introducing breaks in the double-stranded DNA identified by the gRNA. Co-expression of both a multiplex of HIV-1-specific gRNAs and Cas9 in cells results in the modification and/or excision of the segment of viral DNA, leading to replication-defective virus. In this study, we have personalized the activity of CRISPR/Cas9 by placing the gene encoding Cas9 under the control of a minimal promoter of HIV-1 that is activated by the HIV-1 Tat protein. We demonstrate that functional activation of CRISPR/Cas9 by Tat during the course of viral infection excises the designated segment of the integrated viral DNA and consequently suppresses viral expression. This strategy was also used in a latently infected CD4+ T-cell model after treatment with a variety of HIV-1 stimulating agents including PMA and TSA. Controlled expression of Cas9 by Tat offers a new strategy for safe implementation of the Cas9 technology for ablation of HIV-1 at a very early stage of HIV-1 replication during the course of the acute phase of infection and the reactivation of silent proviral DNA in latently infected cells. PMID:27528385

  4. Negative Feedback Regulation of HIV-1 by Gene Editing Strategy

    Science.gov (United States)

    Kaminski, Rafal; Chen, Yilan; Salkind, Julian; Bella, Ramona; Young, Won-bin; Ferrante, Pasquale; Karn, Jonathan; Malcolm, Thomas; Hu, Wenhui; Khalili, Kamel

    2016-01-01

    The CRISPR/Cas9 gene editing method is comprised of the guide RNA (gRNA) to target a specific DNA sequence for cleavage and the Cas9 endonuclease for introducing breaks in the double-stranded DNA identified by the gRNA. Co-expression of both a multiplex of HIV-1-specific gRNAs and Cas9 in cells results in the modification and/or excision of the segment of viral DNA, leading to replication-defective virus. In this study, we have personalized the activity of CRISPR/Cas9 by placing the gene encoding Cas9 under the control of a minimal promoter of HIV-1 that is activated by the HIV-1 Tat protein. We demonstrate that functional activation of CRISPR/Cas9 by Tat during the course of viral infection excises the designated segment of the integrated viral DNA and consequently suppresses viral expression. This strategy was also used in a latently infected CD4+ T-cell model after treatment with a variety of HIV-1 stimulating agents including PMA and TSA. Controlled expression of Cas9 by Tat offers a new strategy for safe implementation of the Cas9 technology for ablation of HIV-1 at a very early stage of HIV-1 replication during the course of the acute phase of infection and the reactivation of silent proviral DNA in latently infected cells. PMID:27528385

  5. Getting Started with Isabelle/jEdit

    OpenAIRE

    Sternagel, Christian

    2012-01-01

    We give a beginner-oriented introduction to Isabelle/jEdit, providing motivation for using it as well as pointing at some differences to the traditional Proof General interface and current limitations.

  6. KWIC Index of nuclear codes (1975 edition)

    International Nuclear Information System (INIS)

    It is a KWIC Index for 254 nuclear codes in the Nuclear Code Abstracts (1975 edition). The classification of nuclear codes and the form of index are the same as those in the Computer Programme Library at Ispra, Italy. (auth.)

  7. A genome editing primer for the hematologist.

    Science.gov (United States)

    Hoban, Megan D; Bauer, Daniel E

    2016-05-26

    Gene editing enables the site-specific modification of the genome. These technologies have rapidly advanced such that they have entered common use in experimental hematology to investigate genetic function. In addition, genome editing is becoming increasingly plausible as a treatment modality to rectify genetic blood disorders and improve cellular therapies. Genome modification typically ensues from site-specific double-strand breaks and may result in a myriad of outcomes. Even single-strand nicks and targeted biochemical modifications that do not permanently alter the DNA sequence (epigenome editing) may be powerful instruments. In this review, we examine the various technologies, describe their advantages and shortcomings for engendering useful genetic alterations, and consider future prospects for genome editing to impact hematology. PMID:27053532

  8. Specification Editing and Discovery Assistant Project

    Data.gov (United States)

    National Aeronautics and Space Administration — The project will prototype a specification editing and discovery tool (SPEEDY) for C/C++ that will assist software developers with modular formal verification tasks...

  9. LabVIEW 8 student edition

    CERN Document Server

    Bishop, Robert H

    2007-01-01

    For courses in Measurement and Instrumentation, Electrical Engineering lab, and Physics and Chemistry lab. This revised printing has been updated to include new LabVIEW 8.2 Student Edition. National Instruments' LabVIEW is the defacto industry standard for test, measurement, and automation software solutions. With the Student Edition of LabVIEW, students can design graphical programming solutions to their classroom problems and laboratory experiments with software that delivers the graphical programming capabilites of the LabVIEW professional version. . The Student Edition is also compatible with all National Instruments data acquisition and instrument control hardware. Note: The LabVIEW Student Edition is available to students, faculty, and staff for personal educational use only. It is not intended for research, institutional, or commercial use. For more information about these licensing options, please visit the National Instruments website at (http:www.ni.com/academic/)

  10. Prescott’s Microbiology, Eighth Edition

    OpenAIRE

    Joanne J. Dobbins

    2010-01-01

    Review of: Prescott’s Microbiology, Eighth Edition. Joanne M. Willey, Linda M. Sherwood, and Christopher J. Woolverton. 2011. McGraw-Hill Higher Education, NewYork, NY. 1070 pages. ISBN- 978-0-07-337526-7.

  11. Good edit similarity learning by loss minimization

    OpenAIRE

    Bellet, Aurélien; Habrard, Amaury; Sebban, Marc

    2012-01-01

    Similarity functions are a fundamental component of many learning algorithms. When dealing with string or tree-structured data, edit distancebased measures are widely used, and there exists a few methods for learning them from data. However, these methods offer no theoretical guarantee as to the generalization ability and discriminative power of the learned similarities. In this paper, we propose a loss minimization-based edit similarity learning approach, called GESL. It is driven by the not...

  12. Nuclear electronics laboratory manual 1989 edition

    International Nuclear Information System (INIS)

    This manual is a joint product of several electronics experts who have been associated with IAEA activity in this field for many years. It is based on the experience of conducting twenty-three training courses on nuclear electronics. Compared with the first edition, published 1984, this edition contains many new experiments, mainly on the advanced technical level. The total number of experiments and special projects is 58. Tabs and figs

  13. Discovery-based edit assistance for spreadsheets

    OpenAIRE

    Cunha, Jácome Miguel Costa; Saraiva, João Alexandre; Visser, Joost

    2009-01-01

    Spreadsheets can be viewed as a highly flexible endusers programming environment which enjoys wide-spread adoption. But spreadsheets lack many of the structured programming concepts of regular programming paradigms. In particular, the lack of data structures in spreadsheets may lead spreadsheet users to cause redundancy, loss, or corruption of data during edit actions. In this paper, we demonstrate how implicit structural properties of spreadsheet data can be exploited to offer edit assistanc...

  14. Portfolio of compositions and scholarly edition

    OpenAIRE

    Caponnetto, Alfredo

    2011-01-01

    A. SCHOLARLY EDITION 1. Zoilo Annibale, Ave Regina a Quattro Chori, ed. by Caponnetto A. and O’Regan N. in Caponnetto A. Ph.D. dissertation, item 2‐ Scholarly Edition, University of Edinburgh, 2011. B. PORTFOLIO OF COMPOSITIONS 1. Caponnetto Alfredo, Ave Regina, for eight solo voices or choir and organ, in Caponnetto A. Ph.D. dissertation, item 1, University of Edinburgh, 2011. 2. Caponnetto Alfredo, Dialectic of the Many and the One: Protagoras‐Parmenides‐Gorgias, for pi...

  15. Wetland Ecology Principles and Conservation, Second Edition

    OpenAIRE

    Richard Smardon

    2014-01-01

    This is a book review of Wetland Ecology Principles and Conservation, second edition, by Paul Keddy. This review focuses on the book’s content as it relates to wetland sustainability for both science and management. Besides overall comments, comparisons are made with the first edition of the book and then very specific chapter-by-chapter relationships to wetland sustainability are made to illustrate specific applications toward wetland sustainability.

  16. Looking forward to genetically edited fruit crops.

    Science.gov (United States)

    Nagamangala Kanchiswamy, Chidananda; Sargent, Daniel James; Velasco, Riccardo; Maffei, Massimo E; Malnoy, Mickael

    2015-02-01

    The availability of genome sequences for many fruit crops has redefined the boundaries of genetic engineering and genetically modified (GM) crop plants. However commercialization of GM crops is hindered by numerous regulatory and social hurdles. Here, we focus on recently developed genome-editing tools for fruit crop improvement and their importance from the consumer perspective. Challenges and opportunities for the deployment of new genome-editing tools for fruit plants are also discussed. PMID:25129425

  17. Improving OCR Accuracy for Classical Critical Editions

    OpenAIRE

    Boschetti, Federico; Romanello, Matteo; Babeu, Alison; Bamman, David; Crane, Gregory

    2009-01-01

    This paper describes a work-flow designed to populate a digital library of ancient Greek critical editions with highly accurate OCR scanned text. While the most recently available OCR engines are now able after suitable training to deal with the polytonic Greek fonts used in 19th and 20th century editions, further improvements can also be achieved with postprocessing. In particular, the progressive multiple alignment method applied to different OCR outputs based on the same ...

  18. [Textual research on the edition of WU Kun's books].

    Science.gov (United States)

    Wang, Xu-Guang; Lu, Xiang

    2013-03-01

    WU Kun is a famous physician of the Xin'an School. He wrote a 6-volume Yi fang kao (Textual Research on Recipes) in 1584, a 24-volume Wu zhu su wen (Wu's Annotation of Plain Questions) in 1594, a 6-volume Zhen fang liu ji (Six Collections of Acupuncture and Prescription) in 1618 block-printed and funded by CHENG Biao, and a 2-volume Mai yu (Language of Pulse) with writing date unknown. For Yi fang kao, there were 4 editions, including Ming-dynasty block-printed edition, Japanese and Korean block-printed edition, old hand-copied edition and modern printing edition. For Wu zhu su wen, there were 5 editions, Ming-Qing-dynasty block-printed edition, Japanese block-printed edition, old hand-copied edition, photocopy and modern edition. For Zhen fang liu ji, there were Ming-dynasty block-printed edition, hand-copied edition, photocopy and modern edition. Mai yu always was printed together with Yi fang kao, and its editions were same as those of Yi fang kao, except some modern printing edition with Mai yu excluded. PMID:24135480

  19. Applications of genome editing in insects.

    Science.gov (United States)

    Reid, William; O'Brochta, David A

    2016-02-01

    Insect genome editing was first reported 1991 in Drosophila melanogaster but the technology used was not portable to other species. Not until the recent development of facile, engineered DNA endonuclease systems has gene editing become widely available to insect scientists. Most applications in insects to date have been technical in nature but this is rapidly changing. Functional genomics and genetics-based insect control efforts will be major beneficiaries of the application of contemporary gene editing technologies. Engineered endonucleases like Cas9 make it possible to create powerful and effective gene drive systems that could be used to reduce or even eradicate specific insect populations. 'Best practices' for using Cas9-based editing are beginning to emerge making it easier and more effective to design and use but gene editing technologies still require traditional means of delivery in order to introduce them into somatic and germ cells of insects-microinjection of developing embryos. This constrains the use of these technologies by insect scientists. Insects created using editing technologies challenge existing governmental regulatory structures designed to manage genetically modified organisms. PMID:27436552

  20. RNA oxidation

    DEFF Research Database (Denmark)

    Kjaer, L. K.; Cejvanovic, V.; Henriken, T.;

    2015-01-01

    .9 significant hazard ratio for death compared with the quartile with the lowest 8oxoGuo excretion when adjusted for age, sex, BMI, smoker status, s-HbA1c, urine protein excretion and s-cholesterol. We conclude that it is now established that RNA oxidation is an independent risk factor for death in type 2...

  1. Enhanced genome editing in mammalian cells with a modified dual-fluorescent surrogate system.

    Science.gov (United States)

    Zhou, Yan; Liu, Yong; Hussmann, Dianna; Brøgger, Peter; Al-Saaidi, Rasha Abdelkadhem; Tan, Shuang; Lin, Lin; Petersen, Trine Skov; Zhou, Guang Qian; Bross, Peter; Aagaard, Lars; Klein, Tino; Rønn, Sif Groth; Pedersen, Henrik Duelund; Bolund, Lars; Nielsen, Anders Lade; Sørensen, Charlotte Brandt; Luo, Yonglun

    2016-07-01

    Programmable DNA nucleases such as TALENs and CRISPR/Cas9 are emerging as powerful tools for genome editing. Dual-fluorescent surrogate systems have been demonstrated by several studies to recapitulate DNA nuclease activity and enrich for genetically edited cells. In this study, we created a single-strand annealing-directed, dual-fluorescent surrogate reporter system, referred to as C-Check. We opted for the Golden Gate Cloning strategy to simplify C-Check construction. To demonstrate the utility of the C-Check system, we used the C-Check in combination with TALENs or CRISPR/Cas9 in different scenarios of gene editing experiments. First, we disrupted the endogenous pIAPP gene (3.0 % efficiency) by C-Check-validated TALENs in primary porcine fibroblasts (PPFs). Next, we achieved gene-editing efficiencies of 9.0-20.3 and 4.9 % when performing single- and double-gene targeting (MAPT and SORL1), respectively, in PPFs using C-Check-validated CRISPR/Cas9 vectors. Third, fluorescent tagging of endogenous genes (MYH6 and COL2A1, up to 10.0 % frequency) was achieved in human fibroblasts with C-Check-validated CRISPR/Cas9 vectors. We further demonstrated that the C-Check system could be applied to enrich for IGF1R null HEK293T cells and CBX5 null MCF-7 cells with frequencies of nearly 100.0 and 86.9 %, respectively. Most importantly, we further showed that the C-Check system is compatible with multiplexing and for studying CRISPR/Cas9 sgRNA specificity. The C-Check system may serve as an alternative dual-fluorescent surrogate tool for measuring DNA nuclease activity and enrichment of gene-edited cells, and may thereby aid in streamlining programmable DNA nuclease-mediated genome editing and biological research. PMID:26755436

  2. Ca2+ permeability of unedited and edited versions of the kainate selective glutamate receptor GluR6.

    OpenAIRE

    Egebjerg, J; Heinemann, S F

    1993-01-01

    The Ca2+ permeability of the kainate selective glutamate receptor GluR6 depends on the editing of the RNA (or DNA). The unedited version of GluR6, GluR6Q, encodes a glutamine at position 621 (Q/R site) and exhibits a Ca2+/monovalent ion permeability ratio of 1.2, while the edited version of GluR6, GluR6R, encodes an arginine at position 621 and exhibits a permeability ratio of 0.47. Kainate activation of the GluR6 receptor results in currents that are modulated by extracellular calcium ions. ...

  3. Book Review: New Perspectives on Technical Editing

    Science.gov (United States)

    Murphy, A. J. (Ed.); Sterken, Christiaan

    2012-08-01

    New Perspectives on Technical Editing by Avon J. Murphy (ed.) ISBN : 978-0895033949 (2010) Baywood Publishing Company Inc, Hardcover, 210 pages, 35.5 GBP This book presents a collection of 10 chapters dealing with diverse aspects of technical editing (ie, editorial planning, and analysis and structural changes made to other people's technological documents): research in technical editing, trends and teaching of technical editing, copyediting, and technical journal editing. The role and function of the modern journal and book editor is also dealt with in detail. Each chapter is written by an expert in the field: senior editors, university professors in technical communication, technical writers and linguists. The ever-evolving role of the editor is clearly elucidated in several historical reviews, and in the descriptions of the expectations for the future. A very striking aspect of this book is its extensive collection of bibliographic resources: every chapter lists dozens of very useful references, and the closing chapter, and annotated bibliography, contain many not so well known references, and are most useful. All in all, the book is a treasure trove listing more than 400 references, in addition to numerous webpage URLs embedded in the texts. The book is designed to help the reader to understand current practices and norms in technical editing, and to help to take action in editing as well as in teaching and educating would-be editors. The audience for this book thus includes editors and teachers, but also writers, researchers and students. A deep reading of this book will result in a better understanding of the difference between full technical editing and its much narrower component so well known as copyediting, and will convince any prospective editor that editing should not be undertaken if the people involved do not master the art of precision and accuracy in technical (as well as in human) communication, do not possess the technical know how and computer

  4. Effects of Trypanosoma brucei tryptophanyl-tRNA synthetases silencing by RNA interference

    Directory of Open Access Journals (Sweden)

    Liliana Torcoroma García

    2007-09-01

    Full Text Available The kinetoplast genetic code deviates from the universal code in that 90% of mitochondrial tryptophans are specified by UGA instead of UGG codons. A single nucleus-encoded tRNA Trp(CCA is used by both nuclear and mitochondria genes, since all kinetoplast tRNAs are imported into the mitochondria from the cytoplasm. To allow decoding of the mitochondrial UGA codons as tryptophan, the tRNA Trp(CCA anticodon is changed to UCA by an editing event. Two tryptophanyl tRNA synthetases (TrpRSs have been identified in Trypanosoma brucei: TbTrpRS1 and TbTrpRS2 which localize to the cytoplasm and mitochondria respectively. We used inducible RNA interference (RNAi to assess the role of TbTrpRSs. Our data validates previous observations of TrpRS as potential drug design targets and investigates the RNAi effect on the mitochondria of the parasite.

  5. Detailed phenotypic and molecular analyses of genetically modified mice generated by CRISPR-Cas9-mediated editing.

    Directory of Open Access Journals (Sweden)

    Bijal A Parikh

    Full Text Available The bacterial CRISPR-Cas9 system has been adapted for use as a genome editing tool. While several recent reports have indicated that successful genome editing of mice can be achieved, detailed phenotypic and molecular analyses of the mutant animals are limited. Following pronuclear micro-injection of fertilized eggs with either wild-type Cas9 or the nickase mutant (D10A and single or paired guide RNA (sgRNA for targeting of the tyrosinase (Tyr gene, we assessed genome editing in mice using rapid phenotypic readouts (eye and coat color. Mutant mice with insertions or deletions (indels in Tyr were efficiently generated without detectable off-target cleavage events. Gene correction of a single nucleotide by homologous recombination (HR could only occur when the sgRNA recognition sites in the donor DNA were modified. Gene repair did not occur if the donor DNA was not modified because Cas9 catalytic activity was completely inhibited. Our results indicate that allelic mosaicism can occur following -Cas9-mediated editing in mice and appears to correlate with sgRNA cleavage efficiency at the single-cell stage. We also show that larger than expected deletions may be overlooked based on the screening strategy employed. An unbiased analysis of all the deleted nucleotides in our experiments revealed that the highest frequencies of nucleotide deletions were clustered around the predicted Cas9 cleavage sites, with slightly broader distributions than expected. Finally, additional analysis of founder mice and their offspring indicate that their general health, fertility, and the transmission of genetic changes were not compromised. These results provide the foundation to interpret and predict the diverse outcomes following CRISPR-Cas9-mediated genome editing experiments in mice.

  6. Louse (Insecta : Phthiraptera) mitochondrial 12S rRNA secondary structure is highly variable

    OpenAIRE

    Page, R.D.M.; Cruickshank, R.; Johnson, K P

    2002-01-01

    Lice are ectoparasitic insects hosted by birds and mammals. Mitochondrial 12S rRNA sequences obtained from lice show considerable length variation and are very difficult to align. We show that the louse 12S rRNA domain III secondary structure displays considerable variation compared to other insects, in both the shape and number of stems and loops. Phylogenetic trees constructed from tree edit distances between louse 12S rRNA structures do not closely resemble trees constructed from sequence ...

  7. The Express-Lane Edit: Making Editing Useful for Young Adolescents

    Science.gov (United States)

    Anderson, Jeff

    2008-01-01

    Editing is a powerful tool for writers, but are our methods of teaching it really demonstrating that power for young adolescents? The author, frustrated with students' inability to edit, blames his own approach and, beginning with a grocery store epiphany, works to develop a more effective system. Elements of his successful approach include time…

  8. CRISPR-based genome editing and expression control systems in Clostridium acetobutylicum and Clostridium beijerinckii.

    Science.gov (United States)

    Li, Qi; Chen, Jun; Minton, Nigel P; Zhang, Ying; Wen, Zhiqiang; Liu, Jinle; Yang, Haifeng; Zeng, Zhe; Ren, Xiaodan; Yang, Junjie; Gu, Yang; Jiang, Weihong; Jiang, Yu; Yang, Sheng

    2016-07-01

    Solventogenic clostridia are important industrial microorganisms that produce various chemicals and fuels. Effective genetic tools would facilitate physiological studies aimed both at improving our understanding of metabolism and optimizing solvent productivity through metabolic engineering. Here we have developed an all-in-one, CRISPR-based genome editing plasmid, pNICKclos, that can be used to achieve successive rounds of gene editing in Clostridium acetobutylicum ATCC 824 and Clostridium beijerinckii NCIMB 8052 with efficiencies varying from 6.7% to 100% and 18.8% to 100%, respectively. The plasmid specifies the requisite target-specific guide RNA, the gene encoding the Streptococcus pyogenes Cas9 nickase and the genome editing template encompassing the gene-specific homology arms. It can be used to create single target mutants within three days, with a further two days required for the curing of the pNICKclos plasmid ready for a second round of mutagenesis. A S. pyogenes dCas9-mediated gene regulation control system, pdCASclos, was also developed and used in a CRISPRi strategy to successfully repress the expression of spo0A in C. acetobutylicum and C. beijerinckii. The combined application of the established high efficiency CRISPR-Cas9 based genome editing and regulation control systems will greatly accelerate future progress in the understanding and manipulation of metabolism in solventogenic clostridia. PMID:27213844

  9. The genome editing revolution: A CRISPR-Cas TALE off-target story.

    Science.gov (United States)

    Stella, Stefano; Montoya, Guillermo

    2016-07-01

    In the last 10 years, we have witnessed a blooming of targeted genome editing systems and applications. The area was revolutionized by the discovery and characterization of the transcription activator-like effector proteins, which are easier to engineer to target new DNA sequences than the previously available DNA binding templates, zinc fingers and meganucleases. Recently, the area experimented a quantum leap because of the introduction of the clustered regularly interspaced short palindromic repeats (CRISPR)-associated protein (Cas) system (clustered regularly interspaced short palindromic sequence). This ribonucleoprotein complex protects bacteria from invading DNAs, and it was adapted to be used in genome editing. The CRISPR ribonucleic acid (RNA) molecule guides to the specific DNA site the Cas9 nuclease to cleave the DNA target. Two years and more than 1000 publications later, the CRISPR-Cas system has become the main tool for genome editing in many laboratories. Currently the targeted genome editing technology has been used in many fields and may be a possible approach for human gene therapy. Furthermore, it can also be used to modifying the genomes of model organisms for studying human pathways or to improve key organisms for biotechnological applications, such as plants, livestock genome as well as yeasts and bacterial strains. PMID:27417121

  10. Identification of the long, edited dsRNAome of LPS-stimulated immune cells.

    Science.gov (United States)

    Blango, Matthew G; Bass, Brenda L

    2016-06-01

    Endogenous double-stranded RNA (dsRNA) must be intricately regulated in mammals to prevent aberrant activation of host inflammatory pathways by cytosolic dsRNA binding proteins. Here, we define the long, endogenous dsRNA repertoire in mammalian macrophages and monocytes during the inflammatory response to bacterial lipopolysaccharide. Hyperediting by adenosine deaminases that act on RNA (ADAR) enzymes was quantified over time using RNA-seq data from activated mouse macrophages to identify 342 Editing Enriched Regions (EERs), indicative of highly structured dsRNA. Analysis of publicly available data sets for samples of human peripheral blood monocytes resulted in discovery of 3438 EERs in the human transcriptome. Human EERs had predicted secondary structures that were significantly more stable than those of mouse EERs and were located primarily in introns, whereas nearly all mouse EERs were in 3' UTRs. Seventy-four mouse EER-associated genes contained an EER in the orthologous human gene, although nucleotide sequence and position were only rarely conserved. Among these conserved EER-associated genes were several TNF alpha-signaling genes, including Sppl2a and Tnfrsf1b, important for processing and recognition of TNF alpha, respectively. Using publicly available data and experimental validation, we found that a significant proportion of EERs accumulated in the nucleus, a strategy that may prevent aberrant activation of proinflammatory cascades in the cytoplasm. The observation of many ADAR-edited dsRNAs in mammalian immune cells, a subset of which are in orthologous genes of mouse and human, suggests a conserved role for these structured regions. PMID:27197207

  11. Triplex-mediated genome targeting and editing.

    Science.gov (United States)

    Reza, Faisal; Glazer, Peter M

    2014-01-01

    Genome targeting and editing in vitro and in vivo can be achieved through an interplay of exogenously introduced molecules and the induction of endogenous recombination machinery. The former includes a repertoire of sequence-specific binding molecules for targeted induction and appropriation of this machinery, such as by triplex-forming oligonucleotides (TFOs) or triplex-forming peptide nucleic acids (PNAs) and recombinagenic donor DNA, respectively. This versatile targeting and editing via recombination approach facilitates high-fidelity and low-off-target genome mutagenesis, repair, expression, and regulation. Herein, we describe the current state-of-the-art in triplex-mediated genome targeting and editing with a perspective towards potential translational and therapeutic applications. We detail several materials and methods for the design, delivery, and use of triplex-forming and recombinagenic molecules for mediating and introducing specific, heritable, and safe genomic modifications. Furthermore we denote some guidelines for endogenous genome targeting and editing site identification and techniques to test targeting and editing efficiency. PMID:24557900

  12. CRISPR/Cas9: A powerful tool for crop genome editing

    Directory of Open Access Journals (Sweden)

    Gaoyuan Song

    2016-04-01

    Full Text Available The CRISPR/Cas9 technology is evolved from a type II bacterial immune system and represents a new generation of targeted genome editing technology that can be applied to nearly all organisms. Site-specific modification is achieved by a single guide RNA (usually about 20 nucleotides that is complementary to a target gene or locus and is anchored by a protospacer-adjacent motif. Cas9 nuclease then cleaves the targeted DNA to generate double-strand breaks (DSBs, which are subsequently repaired by non-homologous end joining (NHEJ or homology-directed repair (HDR mechanisms. NHEJ may introduce indels that cause frame shift mutations and hence the disruption of gene functions. When combined with double or multiplex guide RNA design, NHEJ may also introduce targeted chromosome deletions, whereas HDR can be engineered for target gene correction, gene replacement, and gene knock-in. In this review, we briefly survey the history of the CRISPR/Cas9 system invention and its genome-editing mechanism. We also describe the most recent innovation of the CRISPR/Cas9 technology, particularly the broad applications of modified Cas9 variants, and discuss the potential of this system for targeted genome editing and modification for crop improvement.

  13. CRISPR/Cas9:A powerful tool for crop genome editing

    Institute of Scientific and Technical Information of China (English)

    Gaoyuan Song; Meiling Jia; Kai Chen; Xingchen Kong; Bushra Khattak; Chuanxiao Xie; Aili Li; Long Mao

    2016-01-01

    The CRISPR/Cas9 technology is evolved from a type II bacterial immune system and represents a new generation of targeted genome editing technology that can be applied to nearly all organisms. Site-specific modification is achieved by a single guide RNA (usually about 20 nucleotides) that is complementary to a target gene or locus and is anchored by a protospacer-adjacent motif. Cas9 nuclease then cleaves the targeted DNA to generate double-strand breaks (DSBs), which are subsequently repaired by non-homologous end joining (NHEJ) or homology-directed repair (HDR) mechanisms. NHEJ may introduce indels that cause frame shift mutations and hence the disruption of gene functions. When combined with double or multiplex guide RNA design, NHEJ may also introduce targeted chromosome deletions, whereas HDR can be engineered for target gene correction, gene replacement, and gene knock-in. In this review, we briefly survey the history of the CRISPR/Cas9 system invention and its genome-editing mechanism. We also describe the most recent innovation of the CRISPR/Cas9 technology, particularly the broad applications of modified Cas9 variants, and discuss the potential of this system for targeted genome editing and modification for crop improvement.

  14. Ethical and regulatory aspects of genome editing.

    Science.gov (United States)

    Kohn, Donald B; Porteus, Matthew H; Scharenberg, Andrew M

    2016-05-26

    Gene editing is a rapidly developing area of biotechnology in which the nucleotide sequence of the genome of living cells is precisely changed. The use of genome-editing technologies to modify various types of blood cells, including hematopoietic stem cells, has emerged as an important field of therapeutic development for hematopoietic disease. Although these technologies offer the potential for generation of transformative therapies for patients suffering from myriad disorders of hematopoiesis, their application for therapeutic modification of primary human cells is still in its infancy. Consequently, development of ethical and regulatory frameworks that ensure their safe and effective use is an increasingly important consideration. Here, we review a number of issues that have the potential to impact the clinical implementation of genome-editing technologies, and suggest paths forward for resolving them such that new therapies can be safely and rapidly translated to the clinic. PMID:27053531

  15. Introduction to nuclear science, second edition

    CERN Document Server

    Bryan, Jeff C.

    2013-01-01

    This book was written to provide students who have limited backgrounds in the physical sciences and math with an accessible textbook on nuclear science. Expanding on the foundation of the bestselling first edition, Introduction to Nuclear Science, Second Edition provides a clear and complete introduction to nuclear chemistry and physics, from basic concepts to nuclear power and medical applications. Incorporating suggestions from professors using this book for their courses, the author has created a new text that is approximately 60 percent larger and more comprehensive and flexible than the first.New to This Edition: Thorough review of nuclear forensics, radiology, gamma cameras, and decay through proton or neutron emission More detailed explanations of the necessary mathematics A chapter on dosimetry of radiation fields Expanded discussion of applications, introduced earlier in the text More in-depth coverage of nuclear reactors, including a new chapter examining more reactor types, their safety systems,...

  16. Transportation Energy Data Book, Edition 19; TOPICAL

    International Nuclear Information System (INIS)

    The Transportation Energy Data Book: Edition 19 is a statistical compendium prepared and published by Oak Ridge National Laboratory (ORNL) under contract with the Office of Transportation Technologies in the Department of Energy (DOE). Designed for use as a desk-top reference, the data book represents an assembly and display of statistics and information that characterize transportation activity, and presents data on other factors that influence transportation energy use. The purpose of this document is to present relevant statistical data in the form of tables and graphs. The latest editions of the Data Book are available to a larger audience via the Internet (http://www-cta.ornl.gov/data/tedb.htm)

  17. Transportation Energy Data Book, Edition 19

    Energy Technology Data Exchange (ETDEWEB)

    Davis, S.C.

    1999-09-01

    The Transportation Energy Data Book: Edition 19 is a statistical compendium prepared and published by Oak Ridge National Laboratory (ORNL) under contract with the Office of Transportation Technologies in the Department of Energy (DOE). Designed for use as a desk-top reference, the data book represents an assembly and display of statistics and information that characterize transportation activity, and presents data on other factors that influence transportation energy use. The purpose of this document is to present relevant statistical data in the form of tables and graphs. The latest editions of the Data Book are available to a larger audience via the Internet (http://www-cta.ornl.gov/data/tedb.htm).

  18. Radiation protection - general regulations. 1992 edition

    International Nuclear Information System (INIS)

    The edition 1977 of the fire brigade service regulation 9/1 has been revised completely and passed as ''edition 1992''. As far as it has not been done, the Federal uniform introduction through the countries is realized at short-term. Though the application of radioactive materials (also reactor fuels and basic material) in research, technology, medicine, additional endangering possibilities may occur for application forces in fire company service. FwDV 9/91 contains regulations according to which these dangers can be met through corresponding measures. (orig.)

  19. Prince2 2009 edition a pocket guide

    CERN Document Server

    Hedeman, Bert

    2010-01-01

    This Pocket Guide supplies a summary of the PRINCE2 method, to provide a quick introduction as well as a structured overview of the method;Main target Group for this pocket guide is anyone who wants to get to know the method PRINCE2 or a methodical approach for project management. The book is also very useful for members of a project management team on a project using the PRINCE2 method. Furthermore this pocket guide can be used as literature for the preparation of the PRINCE2 2009 Edition Foundation exam;This pocket guide is based on PRINCE2 2009 Edition;This pocket book deals with processes,

  20. European Corporate Law, 2nd edition

    DEFF Research Database (Denmark)

    Werlauff, Erik; Dorresteijn, Adriaan; Monteiro, Tiago Pereira;

    As in the First Edition (1995) of this well-known book, the authors demonstrate that analysis and comparison of national corporate laws on a number of issues yield highly valuable general principles and observations, not least because business organisations, wherever located, tend to show a...... fundamentally similar set of legal characteristics. To its original selection of six representative jurisdictions - Belgium, France, Germany, The Netherlands, Spain, and the United Kingdom - the Second Edition now adds Poland, thus including an Eastern European perspective to supplement those of continental...

  1. 76 FR 68740 - Trademark Manual of Examining Procedure, Eighth Edition

    Science.gov (United States)

    2011-11-07

    ... attorneys and representatives for trademark applicants with a reference on the practices and procedures for... reference. Links to the seventh edition, as well as to the fourth, fifth, and sixth editions, are on...

  2. [On two editions of Yunyu Xuanji and their source].

    Science.gov (United States)

    Deng, Yue-E

    2013-01-01

    Yunyu Xuanji is considered as one of the 400 key ancient books in the 'TCM Ancient Books Protection and Use Project' by the State Bureau of TCM. Two editions of this book are hand-written copies. The Qianyao edition was finished in the 52(nd) year of the Kangxi Period (1713). The other edition (Taoshi Xianyishukoucongshu) was probably done in the Late Qing Dynasty or the period of the Republic of China. By careful comparison, the contents of the two editions are basically the same, and only the preface of the former edition has been given some changes, which were probably made by later generations. The second manuscript was written in a more standard mode. Therefore the two editions were written according to the same literature and it is possible that the latter one is the copied edition of the Qianyao edition. PMID:23879984

  3. Generation of an Oocyte-Specific Cas9 Transgenic Mouse for Genome Editing.

    Directory of Open Access Journals (Sweden)

    Linlin Zhang

    Full Text Available The CRISPR/Cas9 system has been developed as an easy-handle and multiplexable approach for engineering eukaryotic genomes by zygote microinjection of Cas9 and sgRNA, while preparing Cas9 for microinjection is laborious and introducing inconsistency into the experiment. Here, we describe a modified strategy for gene targeting through using oocyte-specific Cas9 transgenic mouse. With this mouse line, we successfully achieve precise gene targeting by injection of sgRNAs only into one-cell-stage embryos. Through comprehensive analysis, we also show allele complexity and off-target mutagenesis induced by this strategy is obviously lower than Cas9 mRNA/sgRNA injection. Thus, injection of sgRNAs into oocyte-specific Cas9 transgenic mouse embryo provides a convenient, efficient and reliable approach for mouse genome editing.

  4. Efficient Genome Editing in Caenorhabditis elegans with a Toolkit of Dual-Marker Selection Cassettes.

    Science.gov (United States)

    Norris, Adam D; Kim, Hyun-Min; Colaiácovo, Mónica P; Calarco, John A

    2015-10-01

    Use of the CRISPR/Cas9 RNA-guided endonuclease complex has recently enabled the generation of double-strand breaks virtually anywhere in the C. elegans genome. Here, we present an improved strategy that makes all steps in the genome editing process more efficient. We have created a toolkit of template-mediated repair cassettes that contain an antibiotic resistance gene to select for worms carrying the repair template and a fluorescent visual marker that facilitates identification of bona fide recombinant animals. Homozygous animals can be identified as early as 4-5 days post-injection, and minimal genotyping by PCR is required. We demonstrate that our toolkit of dual-marker vectors can generate targeted disruptions, deletions, and endogenous tagging with fluorescent proteins and epitopes. This strategy should be useful for a wide variety of additional applications and will provide researchers with increased flexibility when designing genome editing experiments. PMID:26232410

  5. Delivery of Genome Editing Reagents to Hematopoietic Stem/Progenitor Cells.

    Science.gov (United States)

    Hoban, Megan D; Romero, Zulema; Cost, Gregory J; Mendel, Matthew; Holmes, Michael; Kohn, Donald B

    2016-01-01

    This unit describes the protocol for the delivery of reagents for targeted genome editing to CD34(+) hematopoietic stem/progenitor cells (HSPCs). Specifically, this unit focuses on the process of thawing and pre-stimulating CD34(+) HSPCs, as well as the details of their electroporation with in vitro-transcribed mRNA-encoding site-specific nucleases [in this case zinc-finger nucleases (ZFNs)]. In addition, discussed is delivery of a gene editing donor template in the form of an oligonucleotide or integrase-defective lentiviral vector (IDLV). Finally, an analysis of cell survival following treatment and downstream culture conditions are presented. While optimization steps might be needed for each specific application with respect to nuclease and donor template amount, adherence to this protocol will serve as an excellent starting point for this further work. PMID:26840227

  6. Structure and expression of the human Lysyl hydroxylase gene (PLOD): Introns 9 and 16 contain Alu sequences at the sites of recombination in Ehlers-Danlos syndrome type VI patients

    Energy Technology Data Exchange (ETDEWEB)

    Heikkinen, J.; Hautala, T.; Kivirikko, K.I. [Univ. of Oulu (Finland)] [and others

    1994-12-01

    Lysyl hydroxylase (EC 1.14.11.4) catalyzes the formation of hydroxylysine in collagens by the hydroxylation of lysine residues in peptide linkages. This enzyme activity is known to be reduced in patients with the type VI variant of the Ehlers-Danlos syndrome, and the first mutations in the lysyl hydroxylase gene (PLOD) have recently been identified. We have now isolated genomic clones for human lysyl hydroxylase and determined the complete structure of the gene, which contains 19 exons and a 5{prime} flanking region with characteristics shared by housekeeping genes. The constitutive expression of the gene in different tissues further suggests that lysyl hydroxylase has an essential function. We have sequenced the introns of the gene in the region where many mutations and rearrangements analyzed to date are concentrated. Intron 9 and intron 16 show extensive homology resulting from the many Alu sequences found in these introns. Intron 9 contains five and intron 16 eight Alu sequences. The high homology and many short identical or complementary sequences in these introns generate many potential recombination sites with the gene. The delineation of the structure of the lysyl hydroxylase gene contributes significantly to our understanding of the rearrangements in the genome of Ehlers-Danlos type VI patients. 21 refs., 2 figs., 2 tabs.

  7. Experimental Stochatics (2nd edition)

    International Nuclear Information System (INIS)

    for teachers of computational stochastic methods, is the main contribution of this electronic monograph. However, both the book and software suffer from several severe problems. Firstly, I feel that the structure of the text is weak. Probably this is partly the result of the text from the CD-ROM being put into a book format, but the short paragraphs and poorly structured sentences destroy the reading experience. Secondly, although the software is functional, I believe that, like me, many users will be disappointed by the quality of the user interface and the visualizations. The opportunities to interact with the simulations are limited. Thirdly, the presentation is slightly old fashioned and lacking in pedagogical structure. For example, flow charts and Pascal programs are used to present algorithms. To conclude, I am surprised that this electronic monograph warranted a second edition in this form. Teachers may find the examples useful as a starting point, but students and researchers are advised to look elsewhere. (book review)

  8. 基于多能干细胞的Alport综合征的microRNA差异性表达与碱基编辑功能分析%Identification of different expression microRNA and base edit function in Alport syndrome base on iPSCs

    Institute of Scientific and Technical Information of China (English)

    陈文标; 彭武建; 林小聪; 戴勇

    2014-01-01

    目的 分析Alport综合征(AS)的诱导多能干细胞(iPSCs)全基因微小核糖核酸(microRNA)的表达谱,筛选出具有差异性表达的microRNA与发生碱基突变的microRNA.方法 收集1例AS患者与1例健康人的尿液,从尿液中分离尿肾脏管细胞,尿肾脏管细胞分化诱导成iPSCs.运用高通量测序方法对iPSCs的microRNA进行测序与表达量测定,比较分析两人microRNA的表达差异,寻找具有特异性的microRNA靶点.同时,将mi-croRNA序列与miRBase数据库(http://www.mirbase.org/)中已知成熟microRNA序列进行比对,比较两标本mi-croRNA碱基编辑数量,找出发生了碱基突变的microRNA.结果 在两人标本中发现30个microRNA具有显著的差异性表达,其中19个microRNAs表达上调,11个microRNA表达下调.has-miR-3117-3p的倍比值(log2 Ratio)为7.79,在表达上调的microRNA最具有特异性;has-miR-544b的倍比值为-9.09,在表达下调的microRNA最具有特异性.同时在两标本中共有208个microRNA发生了碱基突变,其中104个microRNA,AS患者发生碱基编辑概率比健康人大;77个microRNA,健康人发生碱基编辑概率比AS患者大.结论 AS患者与健康人肾脏组织中的microRNA存在差异性,AS发生碱基编辑引起基因突变的microRNA比健康人更常见.这些差异性表达的microRNAs与microRNA碱基突变的概率可能在AS发病机制中起重要作用.

  9. Annotated Bibliography on Offender Job Training and Placement. Second Edition.

    Science.gov (United States)

    Clem, Constance, Ed.

    This second edition of an annotated bibliography on offender job training and placement lists nearly 400 items from the collection of the National Institute of Corrections (NIC) Information Center in Longmont, Colorado, including 85 items added since the first edition in 1997. Items new to this edition are marked with an asterisk. Citations have…

  10. Editing modifies the GABA(A) receptor subunit alpha3

    DEFF Research Database (Denmark)

    Ohlson, Johan; Pedersen, Jakob Skou; Haussler, David;

    2007-01-01

    to find selectively edited sites and combined it with bioinformatic techniques that find stem-loop structures suitable for editing. We present here the first verified editing candidate detected by this screening procedure. We show that Gabra-3, which codes for the alpha3 subunit of the GABA...

  11. FeynEdit - a tool for drawing Feynman diagrams

    OpenAIRE

    Hahn, T.; Lang, P.

    2007-01-01

    We describe the FeynEdit tool for drawing Feynman diagrams. Input and output is done using the LaTeX macros of FeynArts, which also implies that diagrams drawn by FeynArts can be edited with FeynEdit. The LaTeX code can be conveniently transferred using copy-and-paste.

  12. Human Resources Administration: A School-Based Perspective. Fourth Edition

    Science.gov (United States)

    Smith, Richard

    2009-01-01

    Enhanced and updated, this Fourth Edition of Richard E. Smith's highly successful text examines the growing role of the principal in planning, hiring, staff development, supervision, and other human resource functions. The Fourth Edition includes new sections on ethics, induction, and the role of the mentor teacher. This edition also introduces…

  13. The Academic Job Search Handbook. Third Edition.

    Science.gov (United States)

    Heiberger, Mary Morris; Vick, Julia Miller

    This handbook provides specific advice on all aspects of job-seeking in an increasingly tight academic market, from the timetable for the application process through seeking tenure. New material in this edition provides more information for candidates in scientific and technical fields and more advice for those applying to technical and community…

  14. Does Money Matter in Education? Second Edition

    Science.gov (United States)

    Baker, Bruce D.

    2016-01-01

    This second edition policy brief revisits the long and storied literature on whether money matters in providing a quality education. It includes research released since the original brief in 2012 and covers a handful of additional topics. Increasingly, political rhetoric adheres to the unfounded certainty that money does not make a difference in…

  15. Critical Social Theories. 2nd Edition

    Science.gov (United States)

    Agger, Ben

    2006-01-01

    Praised for its clarity and accessibility, this fully updated edition of "Critical Social Theories" presents a comprehensive analysis of leading social and cultural theories today. Diverse perspectives are addressed from feminism and cultural studies to postmodernism and critical theory. Written accessibly for students and faculty, the second…

  16. The staging system: Display and edit module

    Science.gov (United States)

    Edwards, E.; Bernier, L.

    1976-01-01

    The Display and Edit (D and E) Module described is one of six major modules being developed for the STAGING (STructural Analysis through Generalized INteractive Graphics) System. Several remarks are included concerning the computer environment and the architecture of the data base. The utility of this module is emphasized.

  17. The NMC Horizon Report: 2014 Library Edition

    NARCIS (Netherlands)

    Johnson, L; Adams Becker, S.; Estrada, V.; Freeman, A.; van den Brekel, Guus

    2014-01-01

    The NMC Horizon Report: 2014 Library Edition, examines key trends, significant challenges, and emerging technologies for their potential impact on academic and research libraries worldwide. While there are many local factors affecting libraries, there are also issues that transcend regional boundari

  18. Maine Dance Curriculum Guide. Second Edition.

    Science.gov (United States)

    Harms, Patricia; Nolan, Kathleen; Ross, Ann; Salmon, Nancy; Schaper, MaryEllen; Wescott, Margaret Gould; Weisman, Eleanor

    This second edition updates and aligns the original 1996 publication with the Maine Learning Results, which were enacted by the state legislature in 1997. The Learning Results articulate what knowledge and skills all Maine students should have at various benchmarks throughout the preK-12 continuum. Dance is identified in the Learning Results as…

  19. The Art of Electronics - 2nd Edition

    Science.gov (United States)

    Horowitz, Paul; Hill, Winfield

    1989-09-01

    This is the thoroughly revised and updated second edition of the hugely successful The Art of Electronics. Widely accepted as the single authoritative text and reference on electronic circuit design, both analog and digital, the original edition sold over 125,000 copies worldwide and was translated into eight languages. The book revolutionized the teaching of electronics by emphasizing the methods actually used by citcuit designers - a combination of some basic laws, rules to thumb, and a large nonmathematical treatment that encourages circuit values and performance. The new Art of Electronics retains the feeling of informality and easy access that helped make the first edition so successful and popular. It is an ideal first textbook on electronics for scientists and engineers and an indispensable reference for anyone, professional or amateur, who works with electronic circuits. The best self-teaching book and reference book in electronics Simply indispensable, packed with essential information for all scientists and engineers who build electronic circuits Totally rewritten chapters on microcomputers and microprocessors The first edition of this book has sold over 100,000 copies in seven years, it has a market in virtually all research centres where electronics is important

  20. The Landscape of Qualitative Research. Third Edition

    Science.gov (United States)

    Denzin, Norman K., Ed.; Lincoln, Yvonna, Ed.

    2007-01-01

    This book, the first volume of the paperback versions of the "The SAGE Handbook of Qualitative Research, Third Edition," takes a look at the field from a broadly theoretical perspective, and is composed of the Handbook's Parts I ("Locating the Field"), II ("Major Paradigms and Perspectives"), and VI ("The Future of Qualitative Research"). "The…

  1. Handbook of Qualitative Research. Second Edition.

    Science.gov (United States)

    Denzin, Norman K., Ed.; Lincoln, Yvonna S., Ed.

    This handbook's second edition represents the state of the art for the theory and practice of qualitative inquiry. It features eight new topics, including autoethnography, critical race theory, applied ethnography, queer theory, and "testimonio"every chapter in the handbook has been thoroughly revised and updated. The book contains:"Preface" (1…

  2. Collecting and Interpreting Qualitative Materials. Third Edition

    Science.gov (United States)

    Denzin, Norman K., Ed.; Lincoln, Yvonna, Ed.

    2007-01-01

    This book is the third volume of the paperback versions of "The SAGE Handbook of Qualitative Research, Third Edition." This portion of the handbook considers the tasks of collecting, analyzing, and interpreting empirical materials, and comprises the Handbook's Parts IV ("Methods of Collecting and Analyzing Empirical Materials") and V ("The Art and…

  3. Introduction to Energy - 2nd Edition

    Science.gov (United States)

    Cassedy, Edward S.; Grossman, Peter Z.

    1998-12-01

    Energy issues such as pollution, resource depletion, global warming, nuclear power and waste are problems that demand timely solutions. This book provides a critical examination of the resources, market forces, and social impacts of modern energy production. The book addresses the dilemmas that have arisen due to society's crucial dependence on energy, particularly fossil fuels, and explores the available alternative energy producing technologies. The second edition has increased emphasis on those issues at the forefront of the current energy debate: energy sustainability, climate change, and the radical restructuring of the power industry due to de-regulation. Assuming no prior technical expertise and avoiding complex mathematical formulation, it is directed at a broad readership. The second edition will follow the first in proving especially useful as a textbook for undergraduate programs in Science, Technology and Society (STS), and as a supplementary text in a variety of courses which touch upon energy studies, including environmental and technology policy, environmental, mineral and business law, energy and resource economics. Fully updated second edition of successful first edition that was adopted on Science, Technology and Society courses Provides a critical examination of all aspects of modern energy production for non-technical readers For a broad readership from a variety of backgrounds

  4. A Dictionary of American Idioms. Third Edition.

    Science.gov (United States)

    Makkai, Adam; And Others

    This revised, updated, and expanded edition defines more than 8000 idiomatic words and phrases of American English. Each alphabetical entry has a grammatical explanation and an example sentence. The dictionary, which includes a preface in nine different languages, is a tool to help increase fluency in English for students, business travelers, and…

  5. ETC 408/508: Technical Editing

    Science.gov (United States)

    Charlton, Michael

    2013-01-01

    ETC 408/508: Technical Editing is a cross-listed undergraduate and graduate course at Missouri Western State University, an open admissions public university with approximately 6,000 students. 508 is an elective course for students in the Master of Applied Arts in Written Communication degree and highly recommended for those in the Technical…

  6. The NMC Horizon Report: 2013 Museum Edition

    Science.gov (United States)

    Johnson, L.; Adams Becker, S.; Freeman, A.

    2013-01-01

    The "NMC Horizon Report: 2013 Museum Edition," is a co-production with the Marcus Institute for Digital Education in the Arts (MIDEA), and examines six emerging technologies for their potential impact on and use in education and interpretation within the museum environment: BYOD (Bring Your Own Device), crowdsourcing, electronic…

  7. Prescott’s Microbiology, Eighth Edition

    Directory of Open Access Journals (Sweden)

    Joanne J. Dobbins

    2010-04-01

    Full Text Available Review of: Prescott’s Microbiology, Eighth Edition. Joanne M. Willey, Linda M. Sherwood, and Christopher J. Woolverton. 2011. McGraw-Hill Higher Education, NewYork, NY. 1070 pages. ISBN- 978-0-07-337526-7.

  8. Edit propagation using geometric relationship functions

    KAUST Repository

    Guerrero, Paul

    2014-03-01

    We propose a method for propagating edit operations in 2D vector graphics, based on geometric relationship functions. These functions quantify the geometric relationship of a point to a polygon, such as the distance to the boundary or the direction to the closest corner vertex. The level sets of the relationship functions describe points with the same relationship to a polygon. For a given query point, we first determine a set of relationships to local features, construct all level sets for these relationships, and accumulate them. The maxima of the resulting distribution are points with similar geometric relationships. We show extensions to handle mirror symmetries, and discuss the use of relationship functions as local coordinate systems. Our method can be applied, for example, to interactive floorplan editing, and it is especially useful for large layouts, where individual edits would be cumbersome. We demonstrate populating 2D layouts with tens to hundreds of objects by propagating relatively few edit operations. © 2014 ACM 0730-0301/2014/03- ART15 $15.00.

  9. Thermodynamics of Fluids Under Flow Second Edition

    CERN Document Server

    Jou, David; Criado-Sancho, Manuel

    2011-01-01

    This is the second edition of the book “Thermodynamics of Fluids under Flow,” which was published in 2000 and has now been corrected, expanded and updated. This is a companion book to our other title Extended irreversible thermodynamics (D. Jou, J. Casas-Vázquez and G. Lebon, Springer, 4th edition 2010), and of the textbook Understanding non-equilibrium thermodynamics (G. Lebon, D. Jou and J. Casas-Vázquez, Springer, 2008. The present book is more specialized than its counterpart, as it focuses its attention on the non-equilibrium thermodynamics of flowing fluids, incorporating non-trivial thermodynamic contributions of the flow, going beyond local equilibrium theories, i.e., including the effects of internal variables and of external forcing due to the flow. Whereas the book's first edition was much more focused on polymer solutions, with brief glimpses into ideal and real gases, the present edition covers a much wider variety of systems, such as: diluted and concentrated polymer solutions, polymer ble...

  10. Part C Updates. 11th Edition

    Science.gov (United States)

    Danaher, Joan; Goode, Sue; Lazara, Alex

    2010-01-01

    "Part C Updates" is a compilation of information on various aspects of the Early Intervention Program for Infants and Toddlers with Disabilities (Part C) of the Individuals with Disabilities Education Act (IDEA). This is the eleventh volume in a series of compilations, which included two editions of Part H Updates, the former name of the program.…

  11. European Code against Cancer 4th Edition

    DEFF Research Database (Denmark)

    Friis, Søren; Kesminiene, Ausrele; Espina, Carolina;

    2015-01-01

    were given for carcinogenic drugs and medical radiation in the 4th edition of European Code against Cancer. It is crucial that the application of these measures relies on medical expertise and thorough benefit-risk evaluation. This also pertains to cancer-preventive drugs, and self-medication with...

  12. Science Editing in America: An Overview

    Institute of Scientific and Technical Information of China (English)

    Barbara Gastel

    2003-01-01

    @@ In America, science editing appears to be an increasingly recognized field. In what settings do American science editors work, and what kinds of work do they do ? What is their educational background? What style manuals and other resources do they use? What organizations serve them? What topics and issues do they find of professional interest?

  13. Nanoparticles for Site Specific Genome Editing

    Science.gov (United States)

    McNeer, Nicole Ali

    Triplex-forming peptide nucleic acids (PNAs) can be used to coordinate the recombination of short 50-60 by "donor DNA" fragments into genomic DNA, resulting in site-specific correction of genetic mutations or the introduction of advantageous genetic modifications. Site-specific gene editing in hematopoietic stem and progenitor cells (HSPCs) could result in treatment or cure of inherited disorders of the blood such as beta-thalassemia. Gene editing in HSPCs and differentiated T cells could help combat HIV/AIDs by modifying receptors, such as CCR5, necessary for R5-tropic HIV entry. However, translation of genome modification technologies to clinical practice is limited by challenges in intracellular delivery, especially in difficult-to-transfect hematolymphoid cells. In vivo gene editing could also provide novel treatment for systemic monogenic disorders such as cystic fibrosis, an autosomal recessive disorder caused by mutations in the cystic fibrosis transmembrane receptor. Here, we have engineered biodegradable nanoparticles to deliver oligonucleotides for site-specific genome editing of disease-relevant genes in human cells, with high efficiency, low toxicity, and editing of clinically relevant cell types. We designed nanoparticles to edit the human beta-globin and CCR5 genes in hematopoietic cells. We show that poly(lactic-co-glycolic acid) (PLGA) nanoparticles can delivery PNA and donor DNA for site-specific gene modification in human hematopoietic cells in vitro and in vivo in NOD-scid IL2rgammanull mice. Nanoparticles delivered by tail vein localized to hematopoietic compartments in the spleen and bone marrow of humanized mice, resulting in modification of the beta-globin and CCR5 genes. Modification frequencies ranged from 0.005 to 20% of cells depending on the organ and cell type, without detectable toxicity. This project developed highly versatile methods for delivery of therapeutics to hematolymphoid cells and hematopoietic stem cells, and will help to

  14. Nuclear-physical data editing and analysis

    International Nuclear Information System (INIS)

    Full text: At the modern stage of high energy physics development, investigation of multiple production processes of particles and nucleus-fragments and their automatical editing becomes one of the central problems of high energy physics (elementary-particles). Enormous experimental information accumulated from various registering installation put global physical problems before physicists-experimenters. One of these problems -automatic editing and receiving of correct physical data and thereby to make scientific conclusions [1]. In the article, some questions of automatic editing of experimental data are presented. Modern automatic systems of mathematical editing of data are complex programme, consisting of various components, which are called in definite sequence [2]. The editing of experimental results on ECM is a complex process, connected with using of tens thousands of information storage devices (discs and cartridges) and other external memory equipment. For processing of experimental data the programme system complexes are prepared and created. This system is used for study of hypernuclear and multiparticle processes in different scientific research institutions of Commonwealth of Independent Countries and other countries abroad [3]. At the present in Andijan State University some of processors and methodic systems are used by students of Physics Department in preparing graduation papers. In particular, the dependence of average number of fragments from energy of particles bombarding nucleus-target was investigated. The investigations made in the framework of different models of nuclei, show slow increase of the number of nuclei-fragments, depending of the energy of bombarding particle [4]. In the further the similar systems for investigation and automation of rapid and superrapid reactions, as well as processes in solid and fluid crystals will be created [5]. (author)

  15. Scalable XML Collaborative Editing with Undo short paper

    CERN Document Server

    Martin, Stéphane; Weiss, Stéphane

    2010-01-01

    Commutative Replicated Data-Type (CRDT) is a new class of algorithms that ensures scalable consistency of replicated data. It has been successfully applied to collaborative editing of texts without complex concurrency control. In this paper, we present a CRDT to edit XML data. Compared to existing approaches for XML collaborative editing, our approach is more scalable and handles all the XML editing aspects : elements, contents, attributes and undo. Indeed, undo is recognized as an important feature for collaborative editing that allows to overcome system complexity through error recovery or collaborative conflict resolution.

  16. A Biophysical Model of CRISPR/Cas9 Activity for Rational Design of Genome Editing and Gene Regulation

    OpenAIRE

    Farasat, Iman; Salis, Howard M.

    2016-01-01

    The ability to precisely modify genomes and regulate specific genes will greatly accelerate several medical and engineering applications. The CRISPR/Cas9 (Type II) system binds and cuts DNA using guide RNAs, though the variables that control its on-target and off-target activity remain poorly characterized. Here, we develop and parameterize a system-wide biophysical model of Cas9-based genome editing and gene regulation to predict how changing guide RNA sequences, DNA superhelical densities, ...

  17. A Biophysical Model of CRISPR/Cas9 Activity for Rational Design of Genome Editing and Gene Regulation.

    OpenAIRE

    Iman Farasat; Salis, Howard M.

    2016-01-01

    The ability to precisely modify genomes and regulate specific genes will greatly accelerate several medical and engineering applications. The CRISPR/Cas9 (Type II) system binds and cuts DNA using guide RNAs, though the variables that control its on-target and off-target activity remain poorly characterized. Here, we develop and parameterize a system-wide biophysical model of Cas9-based genome editing and gene regulation to predict how changing guide RNA sequences, DNA superhelical densities, ...

  18. Vitamin D and the RNA transcriptome: more than mRNA regulation

    Directory of Open Access Journals (Sweden)

    MorayJCampbell

    2014-05-01

    Full Text Available The GRCh37.p13 primary assembly of the human genome contains 20805 protein coding mRNA, and 37147 non-protein coding genes and pseudogenes that as a result of RNA processing and editing generate 196501 gene transcripts. Given the size and diversity of the human transcriptome, it is timely to revisit what is known of VDR function in the regulation and targeting of transcription.Early transcriptomic studies using microarray approaches focused on the protein coding mRNA that were regulated by the VDR, usually following treatment with ligand. These studies quickly established the approxamte size, and surprising diversity of the VDR transcriptome, revealing it to be highly heterogenous and cell type and time dependent. With the discovery of microRNA, investigators also considered VDR regulation of these non-protein coding RNA. Again, cell and time dependency has emerged. Attempts to integrate mRNA and miRNA regulation patterns are beginning to reveal patterns of co-regulation and interaction that allow for greater control of mRNA expression, and the capacity to govern more complex cellular events. As the awareness of the diversity of non-coding RNA increases, it is evident that VDR actions are mediated through these molecules also. Key knowledge gaps remain over the VDR transcriptome. The causes for the cell and type dependent transcriptional heterogenetiy remain enigmatic. ChIP-Seq approaches have confirmed that VDR binding choices differ very significantly by cell type, but as yet the underlying causes distilling VDR binding choices are unclear. Similarly, it is clear that many of the VDR binding sites are non-canonical in nature but again the mechanisms underlying these interactions are unclear. Finally, although alternative splicing is clearly a very significant process in cellular transcriptional control, the lack of RNA-Seq data centered on VDR function are currently limiting the global assessment of the VDR transcriptome. VDR focused research

  19. [CRISPR/Cas9-based genome editing systems and the analysis of targeted genome mutations in plants].

    Science.gov (United States)

    Xingliang, Ma; Yaoguang, Liu

    2016-02-01

    Targeted genomic editing technologies use programmable DNA nucleases to cleave genomic target sites, thus inducing targeted mutations in the genomes. The newly prevailed clustered regularly interspaced short palindromic repeats/CRISPR-associated protein 9 (CRISPR/Cas9) system that consists of the Cas9 nuclease and single guide RNA (sgRNA) has the advantages of simplicity and high efficiency as compared to other programmable DNA nuclease systems such as zinc finger nucleases (ZFNs) and transcription activator like effector nucleases (TALENs). Currently, a number of cases have been reported on the application of the CRISPR/Cas9 genomic editing technology in plants. In this review, we summarize the strategies for preparing the Cas9 and sgRNA expression constructs, the transformation method for obtaining targeted mutations, the efficiency and features of the resulting mutations and the methods for detecting or genotyping of the mutation sites. We also discuss the existing problems and perspectives of CRISPR/Cas9-based genomic editing in plants. PMID:26907775

  20. Generating Mouse Models Using CRISPR-Cas9-Mediated Genome Editing.

    Science.gov (United States)

    Qin, Wenning; Kutny, Peter M; Maser, Richard S; Dion, Stephanie L; Lamont, Jeffrey D; Zhang, Yingfan; Perry, Greggory A; Wang, Haoyi

    2016-01-01

    The CRISPR-Cas9 system in bacteria and archaea has recently been exploited for genome editing in various model organisms, including mice. The CRISPR-Cas9 reagents can be delivered directly into the mouse zygote to derive a mutant animal carrying targeted genetic modifications. The major components of the system include the guide RNA, which provides target specificity, the Cas9 nuclease that creates the DNA double-strand break, and the donor oligonucleotide or plasmid carrying the intended mutation flanked by sequences homologous to the target site. Here we describe the general considerations and experimental protocols for creating genetically modified mice using the CRISPR-Cas9 system. PMID:26928663

  1. Long non-coding RNA APTR promotes the activation of hepatic stellate cells and the progression of liver fibrosis

    International Nuclear Information System (INIS)

    In this study, we aimed at assessing a role of Alu-mediated p21 transcriptional regulator (APTR) in hepatofibrogenesis. APTR was upregulated in fibrotic liver samples and activated hepatic stellate cells (HSCs). Knockdown of APTR inhibited the activation of HSCs in vitro and mitigated the accumulation of collagen in vivo. Importantly, APTR silencing could abrogate TGF-β1-induced upregulation of α-SMA in HSCs. In addition, inhibition of cell cycle and cell proliferation by APTR knockdown was attenuated by p21 siRNA1 in primary HSCs. Finally, serum APTR levels were increased in patients with liver cirrhosis, indicating a potential biomarker for liver cirrhosis. Collectively, evidence is proposed for a new biological role of APTR in hepatofibrogenesis. - Highlights: • APTR is upregulated in fibrotic liver tissues and activated HSCs. • APTR silencing inhibits HSC activation and the progression of liver fibrosis. • Antifibrotic effect of APTR silencing is achieved by increasing p21

  2. Genome Editing with CRISPR-Cas9: Can It Get Any Better?

    Science.gov (United States)

    Haeussler, Maximilian; Concordet, Jean-Paul

    2016-05-20

    The CRISPR-Cas revolution is taking place in virtually all fields of life sciences. Harnessing DNA cleavage with the CRISPR-Cas9 system of Streptococcus pyogenes has proven to be extraordinarily simple and efficient, relying only on the design of a synthetic single guide RNA (sgRNA) and its co-expression with Cas9. Here, we review the progress in the design of sgRNA from the original dual RNA guide for S. pyogenes and Staphylococcus aureus Cas9 (SpCas9 and SaCas9). New assays for genome-wide identification of off-targets have provided important insights into the issue of cleavage specificity in vivo. At the same time, the on-target activity of thousands of guides has been determined. These data have led to numerous online tools that facilitate the selection of guide RNAs in target sequences. It appears that for most basic research applications, cleavage activity can be maximized and off-targets minimized by carefully choosing guide RNAs based on computational predictions. Moreover, recent studies of Cas proteins have further improved the flexibility and precision of the CRISPR-Cas toolkit for genome editing. Inspired by the crystal structure of the complex of sgRNA-SpCas9 bound to target DNA, several variants of SpCas9 have recently been engineered, either with novel protospacer adjacent motifs (PAMs) or with drastically reduced off-targets. Novel Cas9 and Cas9-like proteins called Cpf1 have also been characterized from other bacteria and will benefit from the insights obtained from SpCas9. Genome editing with CRISPR-Cas9 may also progress with better understanding and control of cellular DNA repair pathways activated after Cas9-induced DNA cleavage. PMID:27210042

  3. The CRISPR/Cas9 system for gene editing and its potential application in pain research

    Science.gov (United States)

    Sun, Linlin; Lutz, Brianna Marie; Tao, Yuan-Xiang

    2016-01-01

    The CRISPR/Cas9 system is a research hotspot in genome editing and regulation. Currently, it is used in genomic silencing and knock-in experiments as well as transcriptional activation and repression. This versatile system consists of two components: a guide RNA (gRNA) and a Cas9 nuclease. Recognition of a genomic DNA target is mediated through base pairing with a 20-base gRNA. The latter further recruits the Cas9 endonuclease protein to the target site and creates double-stranded breaks in the target DNA. Compared with traditional genome editing directed by DNA-binding protein domains, this short RNA-directed Cas9 endonuclease system is simple and easily programmable. Although this system may have off-target effects and in vivo delivery and immune challenges, researchers have employed this system in vivo to establish disease models, study specific gene functions under certain disease conditions, and correct genomic information for disease treatment. In regards to pain research, the CRISPR/Cas9 system may act as a novel tool in gene correction therapy for pain-associated hereditary diseases and may be a new approach for RNA-guided transcriptional activation or repression of pain-related genes. In addition, this system is also applied to loss-of-function mutations in pain-related genes and knockin of reporter genes or loxP tags at pain-related genomic loci. The CRISPR/Cas9 system will likely be carried out widely in both bench work and clinical settings in the pain field. PMID:27500183

  4. Functional characterization of two paralogs that are novel RNA binding proteins influencing mitochondrial transcripts of Trypanosoma brucei

    Czech Academy of Sciences Publication Activity Database

    Kafková, Lucie; Ammerman, M. L.; Faktorová, D.; Fisk, J. C.; Zimmer, S.L.; Sobotka, Roman; Read, L. K.; Lukeš, Julius; Hashimi, Hassan

    2012-01-01

    Roč. 18, č. 10 (2012), s. 1846-1861. ISSN 1355-8382 R&D Projects: GA ČR GA204/09/1667 Institutional support: RVO:60077344 ; RVO:61388971 Keywords : RNA editing * RNA binding protein * ribonuclear protein (RNP) * mitochondria * trypanosome Subject RIV: EB - Genetics ; Molecular Biology Impact factor: 5.088, year: 2012 http:// rna journal.cshlp.org/content/18/10/1846

  5. Extracellular RNA Communication (ExRNA)

    Data.gov (United States)

    Federal Laboratory Consortium — Until recently, scientists believed RNA worked mostly inside the cell that produced it. Some types of RNA help translate genes into proteins that are necessary for...

  6. RNA metabolism in ALS: when normal processes become pathological.

    Science.gov (United States)

    Droppelmann, Cristian A; Campos-Melo, Danae; Ishtiaq, Muhammad; Volkening, Kathryn; Strong, Michael J

    2014-09-01

    Amyotrophic lateral sclerosis (ALS) is a fatal neurodegenerative disease caused by the death of motor neurons. While the exact molecular and cellular basis for motor neuron death is not yet fully understood, the current conceptualization is that multiple aberrant biological processes contribute. Among these, one of the most compelling is based on alterations of RNA metabolism. In this review, we examine how the normal process of cellular response to stress leading to RNA stress granule formation might become pathological, resulting in the formation of stable protein aggregates. We discuss the emerging roles of post-translational modifications of RNA binding proteins in the genesis of these aggregates. We also review the contemporary literature regarding the potential role for more widespread alterations in RNA metabolism in ALS, including alterations in miRNA biogenesis, spliceosome integrity and RNA editing. A hypothesis is presented in which aberrant RNA processing, modulated through pathological stress granule formation as a reflection of either mutations within intrinsically disordered or prion-like domains of critical RNA binding proteins, or the post-translational modification of RNA binding proteins, contributes directly to motor neuron death. PMID:24555412

  7. Transposable B2 SINE elements can provide mobile RNA polymerase II promoters.

    Science.gov (United States)

    Ferrigno, O; Virolle, T; Djabari, Z; Ortonne, J P; White, R J; Aberdam, D

    2001-05-01

    Short interspersed elements (SINEs) are highly abundant components of mammalian genomes that are propagated by retrotransposition. SINEs are recognized as a causal agent of human disease and must also have had a profound influence in shaping eukaryotic genomes. The B2 SINE family constitutes approximately 0.7% of total mouse genomic DNA (ref. 2) and is also found at low abundance in humans. It resembles the Alu family in several respects, such as its mechanism of propagation. B2 SINEs are derived from tRNA and are transcribed by RNA polymerase (pol) III to generate short transcripts that are not translated. We find here, however, that one B2 SINE also carries an active pol II promoter located outside the tRNA region. Indeed, a B2 element is responsible for the production of a mouse Lama3 transcript. The B2 pol II promoters can be bound and stimulated by the transcription factor USF (for upstream stimulatory factor), as shown by transient transfection experiments. Moreover, this pol II activity does not preclude the pol III transcription necessary for retrotransposition. Dispersal of B2 SINEs by retrotransposition may therefore have provided numerous opportunities for creating regulated pol II transcription at novel genomic sites. This mechanism may have allowed the evolution of new transcription units and new genes. PMID:11326281

  8. Combinatorics of RNA-RNA interaction

    DEFF Research Database (Denmark)

    Li, Thomas J X; Reidys, Christian

    2012-01-01

    RNA-RNA binding is an important phenomenon observed for many classes of non-coding RNAs and plays a crucial role in a number of regulatory processes. Recently several MFE folding algorithms for predicting the joint structure of two interacting RNA molecules have been proposed. Here joint structure...

  9. Building java programs (3rd edition)

    CERN Document Server

    Reges, Stuart

    2013-01-01

    Building Java Programs: A Back to Basics Approach, Third Edition, introduces novice programmers to basic constructs and common pitfalls by emphasizing the essentials of procedural programming, problem solving, and algorithmic reasoning. By using objects early to solve interesting problems and defining objects later in the course, Building Java Programs develops programming knowledge for a broad audience. NEW! This edition is available with MyProgrammingLab, an innovative online homework and assessment tool. Through the power of practice and immediate personalized feedback, MyProgrammingLab helps students fully grasp the logic, semantics, and syntax of programming. Note: If you are purchasing the standalone text or electronic version, MyProgrammingLab does not come automatically packaged with the text. To purchase MyProgrammingLab, please visit: myprogramminglab.com or you can purchase a package of the physical text + MyProgrammingLab by searching the Pearson Higher Education web site. MyProgrammi...

  10. Texture Editing Using Frequency Swap Strategy

    Czech Academy of Sciences Publication Activity Database

    Haindl, Michal; Havlíček, Vojtěch

    Heidelberg : Springer, 2009 - (X. Jiang, N.Petkov), s. 1146-1153 ISBN 978-3-642-03766-5. ISSN 0302-9743. - (Lecture Notes in Computer Science. 5702). [13th International Conference on Computer Analysis of Images and Patterns. Muenster (DE), 02.09.2009-04.09.2009] R&D Projects: GA MŠk 1M0572; GA ČR GA102/08/0593 Grant ostatní: GA MŠk(CZ) 2C06019 Institutional research plan: CEZ:AV0Z10750506 Keywords : texture editing * Markov random fields Subject RIV: BD - Theory of Information http://library.utia.cas.cz/separaty/2009/RO/haindl-texture editing using frequency swap strategy.pdf

  11. Transportation Energy Data Book, Edition 18

    Energy Technology Data Exchange (ETDEWEB)

    Davis, Stacy C.

    1998-09-01

    The Transportation Energy Data Book: Edition 18 is a statistical compendium prepared and published by Oak Ridge National Laboratory (ORNL) under contract with the Office of Transportation Technologies in the Department of Energy (DOE). Designed for use as a desk-top reference, the data book represents an assembly and display of statistics and information that characterize transportation activity, and presents data on other factors that influence transportation energy use. The purpose of this document is to present relevant statistical data in the form of tables and graphs. This edition of the Data Book has 11 chapters which focus on various aspects of the transportation industry. Chapter 1 focuses on petroleum; Chapter 2 - energy Chapter 3 - emissions; Chapter 4 - transportation and the economy; Chapter 5 - highway vehicles; Chapter 6 - Light vehicles; Chapter 7 - heavy vehicles; Chapter 8 - alternative fuel vehicles; Chapter 9 - fleet vehicles; Chapter 10 - household vehicles; and Chapter 11 - nonhighway modes. The sources used represent the latest available data.

  12. Motion Editing for Time-Varying Mesh

    Science.gov (United States)

    Xu, Jianfeng; Yamasaki, Toshihiko; Aizawa, Kiyoharu

    2008-12-01

    Recently, time-varying mesh (TVM), which is composed of a sequence of mesh models, has received considerable interest due to its new and attractive functions such as free viewpoint and interactivity. TVM captures the dynamic scene of the real world from multiple synchronized cameras. However, it is expensive and time consuming to generate a TVM sequence. In this paper, an editing system is presented to reuse the original data, which reorganizes the motions to obtain a new sequence based on the user requirements. Hierarchical motion structure is observed and parsed in TVM sequences. Then, the representative motions are chosen into a motion database, where a motion graph is constructed to connect those motions with smooth transitions. After the user selects some desired motions from the motion database, the best paths are searched by a modified Dijkstra algorithm to achieve a new sequence. Our experimental results demonstrate that the edited sequences are natural and smooth.

  13. Motion Editing for Time-Varying Mesh

    Directory of Open Access Journals (Sweden)

    Kiyoharu Aizawa

    2008-08-01

    Full Text Available Recently, time-varying mesh (TVM, which is composed of a sequence of mesh models, has received considerable interest due to its new and attractive functions such as free viewpoint and interactivity. TVM captures the dynamic scene of the real world from multiple synchronized cameras. However, it is expensive and time consuming to generate a TVM sequence. In this paper, an editing system is presented to reuse the original data, which reorganizes the motions to obtain a new sequence based on the user requirements. Hierarchical motion structure is observed and parsed in TVM sequences. Then, the representative motions are chosen into a motion database, where a motion graph is constructed to connect those motions with smooth transitions. After the user selects some desired motions from the motion database, the best paths are searched by a modified Dijkstra algorithm to achieve a new sequence. Our experimental results demonstrate that the edited sequences are natural and smooth.

  14. Development and potential applications of CRISPR-Cas9 genome editing technology in sarcoma.

    Science.gov (United States)

    Liu, Tang; Shen, Jacson K; Li, Zhihong; Choy, Edwin; Hornicek, Francis J; Duan, Zhenfeng

    2016-04-01

    Sarcomas include some of the most aggressive tumors and typically respond poorly to chemotherapy. In recent years, specific gene fusion/mutations and gene over-expression/activation have been shown to drive sarcoma pathogenesis and development. These emerging genomic alterations may provide targets for novel therapeutic strategies and have the potential to transform sarcoma patient care. The RNA-guided nuclease CRISPR-Cas9 (Clustered Regularly Interspaced Short Palindromic Repeats (CRISPR)-associated protein-9 nuclease) is a convenient and versatile platform for site-specific genome editing and epigenome targeted modulation. Given that sarcoma is believed to develop as a result of genetic alterations in mesenchymal progenitor/stem cells, CRISPR-Cas9 genome editing technologies hold extensive application potentials in sarcoma models and therapies. We review the development and mechanisms of the CRISPR-Cas9 system in genome editing and introduce its application in sarcoma research and potential therapy in clinic. Additionally, we propose future directions and discuss the challenges faced with these applications, providing concise and enlightening information for readers interested in this area. PMID:26806808

  15. Marine mammals: evolutionary biology (second edition)

    OpenAIRE

    Berta, Annalisa; Sumach, James L.; Kit M. Kovacs

    2007-01-01

    The ever-popular field of biology that focuses on mammals that evolved to exploit the marine environment has spawned a suite of texts in the last decade. These have been dedicated to the evolution, systematics, morphology, anatomy, physiology, behaviour, sociobiology, ecology and conservation of this well-studied group. Berta et al.’s latest addition to this literature is an impressive advancement of their first edition (2003), and is certainly the only major authored work of its kind [the co...

  16. Learning Good Edit Similarities with Generalization Guarantees

    OpenAIRE

    Bellet, Aurélien; Habrard, Amaury; Sebban, Marc

    2011-01-01

    Similarity and distance functions are essential to many learning algorithms, thus training them has attracted a lot of interest. When it comes to dealing with structured data (e.g., strings or trees), edit similarities are widely used, and there exists a few methods for learning them. However, these methods offer no theoretical guarantee as to the generalization performance and discriminative power of the resulting similarities. Recently, a theory of learning with good similarity functions wa...

  17. Migration and Remittances Factbook 2016, Third Edition

    OpenAIRE

    World Bank Group

    2016-01-01

    The Migration and Remittances Factbook 2016 attempts to present numbers and facts behind the stories of international migration and remittances, drawing on authoritative, publicly available data. It provides a snapshot of statistics on immigration, emigration, skilled emigration, and remittance flows for 210 countries and 15 regional and income groups. The Migration and Remittances Factbook 2016 updates the 2011 edition of the Factbook with additional data on bilateral migration and remittanc...

  18. Selective access and editing in a database

    Science.gov (United States)

    Maluf, David A. (Inventor); Gawdiak, Yuri O. (Inventor)

    2010-01-01

    Method and system for providing selective access to different portions of a database by different subgroups of database users. Where N users are involved, up to 2.sup.N-1 distinguishable access subgroups in a group space can be formed, where no two access subgroups have the same members. Two or more members of a given access subgroup can edit, substantially simultaneously, a document accessible to each member.

  19. The Challenge of Editing Einstein's Scientific Manuscripts

    OpenAIRE

    Sauer, Tilman

    2004-01-01

    Einstein's research manuscripts provide important insights into his exceptional creativity. At the same time, they can present difficulties for a publication in the documentary edition of the Collected Papers of Albert Einstein (CPAE). The problems are illustrated by discussing how some important examples of Einstein's research manuscripts have been included in previous volumes of the CPAE series: his Scratch Notebook from the years 1910-1914, his so-called Zurich Notebook from 1912, document...

  20. ELECTRONIC EDITIONS OF MATHEMATICAL WORKS IN SERBIA

    OpenAIRE

    D. Blagojevic; Ž. Mijajlovic; Z. Ognjanovic

    2005-01-01

    The paper describes two ongoing projects carried out by the Mathematical Institute of Serbian Academy of Science and Arts, Belgrade and the Faculty of Mathematics, Belgrade. The projects concern building of electronic resources and presentations of electronic editions of mathematical works in Serbia. The first project is related to retro digitization of old books, articles and the other mathematical works and development of the corresponding virtual library. In the second project a database a...

  1. Parallel OCR for Ancient Greek Critical Editions

    OpenAIRE

    Del Grosso, Angelo; Boschetti, Federico

    2012-01-01

    This project is focused on the parallelization of OCR processes applied to Ancient Greek critical editions. Two experiments have been performed. The first experiment is related to parameters differently tuned on the nodes of the grid, in order to identify the best combination that improves the accuracy of the recognition. The second experiment concerns the application of OCR with the best parameters on sample pages by a divide et impera strategy. Results related to the performances of the par...

  2. Efficient Communication Protocols for Deciding Edit Distance

    DEFF Research Database (Denmark)

    Jowhari, Hossein

    2012-01-01

    In this paper we present two communication protocols on computing edit distance. In our first result, we give a one-way protocol for the following Document Exchange problem. Namely given x ∈ Σn to Alice and y ∈ Σn to Bob and integer k to both, Alice sends a message to Bob so that he learns x or...

  3. Medical Imaging Physics, 4th Edition

    Science.gov (United States)

    Hendee, William R.; Ritenour, E. Russell

    2002-05-01

    This comprehensive publication covers all aspects of image formation in modern medical imaging modalities, from radiography, fluoroscopy, and computed tomography, to magnetic resonance imaging and ultrasound. It addresses the techniques and instrumentation used in the rapidly changing field of medical imaging. Now in its fourth edition, this text provides the reader with the tools necessary to be comfortable with the physical principles, equipment, and procedures used in diagnostic imaging, as well as appreciate the capabilities and limitations of the technologies.

  4. Technology Roadmap: Wind Energy. 2013 edition

    Energy Technology Data Exchange (ETDEWEB)

    NONE

    2013-07-01

    The IEA Wind Power Technology Roadmap 2013 Edition recognises the very significant progress made since the first edition was published in 2009. The technology continues to improve rapidly, and costs of generation from land-based wind installations continue to fall. Wind power is now being deployed in countries with good resources without any dedicated financial incentives. The 2013 Edition targets an increased share (15% to 18%) of global electricity to be provided by wind power in 2050, compared to 12% in the original roadmap of 2009. However, increasing levels of low-cost wind still require predictable, supportive regulatory environments and appropriate market designs. The challenges of integrating higher levels of variable wind power into the grid need to be addressed. For offshore wind, much remains to be done to develop appropriate large-scale systems and to reduce costs. The 2013 Wind Power Roadmap also provides updated analysis on the barriers that exist for the technology and suggests ways to address them, including legal and regulatory recommendations.

  5. Transportation Energy Data Book: Edition 21; TOPICAL

    International Nuclear Information System (INIS)

    The Transportation Energy Data Book: Edition 21 is a statistical compendium prepared and published by Oak Ridge National Laboratory (ORNL) under contract with the Office of Transportation Technologies in the Department of Energy (DOE). Designed for use as a desk-top reference, the data book represents an assembly and display of statistics and information that characterize transportation activity, and presents data on other factors that influence transportation energy use. The purpose of this document is to present relevant statistical data in the form of tables and graphs. The latest editions of the Data Book are available to a larger audience via the Internet (www-cta.ornl.gov/data/tedb.htm). This edition of the Data Book has 12 chapters which focus on various aspects of the transportation industry. Chapter 1 focuses on petroleum; Chapter 2-energy; Chapter 3-greenhouse gas emissions; Chapter 4-criteria pollutant emissions; Chapter 5-transportation and the economy; Chapter 6-highway vehicles; Chapter 7-light vehicles; Chapter 8-heavy vehicles; Chapter 9-alternative fuel vehicles; Chapter 10-fleet vehicles; Chapter 11-household vehicles; and Chapter 12-nonhighway modes. The sources used represent the latest available data. There are also three appendices which include detailed source information for some tables, measures of conversion, and the definition of Census divisions and regions. A glossary of terms and a title index are also included for the readers convenience

  6. Edit Distance to Monotonicity in Sliding Windows

    DEFF Research Database (Denmark)

    Chan, Ho-Leung; Lam, Tak-Wah; Lee, Lap Kei;

    2011-01-01

    algorithm which can return an estimate within a factor of (4 + ε) using O(21log2(w)) space. We also extend the study in two directions. First, we consider a stream where each item is associated with a value from a partial ordered set. We give a randomized (4 + ε)-approximate algorithm using O(21log2wlogw......Given a stream of items each associated with a numerical value, its edit distance to monotonicity is the minimum number of items to remove so that the remaining items are non-decreasing with respect to the numerical value. The space complexity of estimating the edit distance to monotonicity...... of a data stream is becoming well-understood over the past few years. Motivated by applications on network quality monitoring, we extend the study to estimating the edit distance to monotonicity of a sliding window covering the w most recent items in the stream for any w ≥ 1. We give a deterministic...

  7. Transportation Energy Data Book (Edition 20)

    Energy Technology Data Exchange (ETDEWEB)

    Davis, S.C.

    2000-10-09

    The ''Transportation Energy Data Book: Edition 20'' is a statistical compendium prepared and published by Oak Ridge National Laboratory (ORNL) under contract with the Office of Transportation Technologies in the Department of Energy (DOE). Designed for use as a desk-top reference, the data book represents an assembly and display of statistics and information that characterize transportation activity, and presents data on other factors that influence transportation energy use. The purpose of this document is to present relevant statistical data in the form of tables and graphs. The latest editions of the Data Book are available to a larger audience via the Internet (www-cta.ornl.gov/data/tedb.htm). This edition of the Data Book has 12 chapters which focus on various aspects of the transportation industry. Chapter 1 focuses on petroleum; Chapter 2--energy; Chapter 3--greenhouse gas emissions; Chapter 4--criteria pollutant emissions; Chapter 5--transportation and the economy; Chapter 6--highway vehicles; Chapter 7--light vehicles; Chapter 8--heavy vehicles; Chapter 9--alternative fuel vehicles; Chapter 10--fleet vehicles; Chapter 11--household vehicles; and Chapter 12--nonhighway modes. The sources used represent the latest available data. There are also three appendices which include detailed source information for some tables, measures of conversion, and the definition of Census divisions and regions. A glossary of terms and a title index are also included for the readers convenience.

  8. Transportation Energy Data Book: Edition 26

    Energy Technology Data Exchange (ETDEWEB)

    Davis, Stacy Cagle [ORNL; Diegel, Susan W [ORNL

    2007-07-01

    The Transportation Energy Data Book: Edition 26 is a statistical compendium prepared and published by Oak Ridge National Laboratory (ORNL) under contract with the Office of Planning, Budget Formulation, and Analysis, under the Energy Efficiency and Renewable Energy (EERE) program in the Department of Energy (DOE). Designed for use as a desk-top reference, the data book represents an assembly and display of statistics and information that characterize transportation activity, and presents data on other factors that influence transportation energy use. The purpose of this document is to present relevant statistical data in the form of tables and graphs. The latest editions of the Data Book are available to a larger audience via the Internet (cta.ornl.gov/data). This edition of the Data Book has 12 chapters which focus on various aspects of the transportation industry. Chapter 1 focuses on petroleum; Chapter 2 - energy; Chapter 3 - highway vehicles; Chapter 4 - light vehicles; Chapter 5 - heavy vehicles; Chapter 6 - alternative fuel vehicles; Chapter 7 - fleet vehicles; Chapter 8 - household vehicles; and Chapter 9- nonhighway modes; Chapter 10 - transportation and the economy; Chapter 11 - greenhouse gas emissions; and Chapter 12 - criteria pollutant emissions. The sources used represent the latest available data. There are also three appendices which include detailed source information for some tables, measures of conversion, and the definition of Census divisions and regions. A glossary of terms and a title index are also included for the readers convenience.

  9. Transportation Energy Data Book: Edition 27

    Energy Technology Data Exchange (ETDEWEB)

    Davis, Stacy Cagle [ORNL; Diegel, Susan W [ORNL; Boundy, Robert Gary [ORNL

    2008-06-01

    The Transportation Energy Data Book: Edition 27 is a statistical compendium prepared and published by Oak Ridge National Laboratory (ORNL) under contract with the Office of Planning, Budget Formulation, and Analysis, under the Energy Efficiency and Renewable Energy (EERE) program in the Department of Energy (DOE). Designed for use as a desk-top reference, the data book represents an assembly and display of statistics and information that characterize transportation activity, and presents data on other factors that influence transportation energy use. The purpose of this document is to present relevant statistical data in the form of tables and graphs. The latest editions of the Data Book are available to a larger audience via the Internet (cta.ornl.gov/data). This edition of the Data Book has 12 chapters which focus on various aspects of the transportation industry. Chapter 1 focuses on petroleum; Chapter 2 energy; Chapter 3 highway vehicles; Chapter 4 light vehicles; Chapter 5 heavy vehicles; Chapter 6 alternative fuel vehicles; Chapter 7 fleet vehicles; Chapter 8 household vehicles; and Chapter 9 nonhighway modes; Chapter 10 transportation and the economy; Chapter 11 greenhouse gas emissions; and Chapter 12 criteria pollutant emissions. The sources used represent the latest available data. There are also three appendices which include detailed source information for some tables, measures of conversion, and the definition of Census divisions and regions. A glossary of terms and a title index are also included for the readers convenience.

  10. Transportation Energy Data Book: Edition 25

    Energy Technology Data Exchange (ETDEWEB)

    Davis, Stacy Cagle [ORNL; Diegel, Susan W [ORNL

    2006-06-01

    The Transportation Energy Data Book: Edition 25 is a statistical compendium prepared and published by Oak Ridge National Laboratory (ORNL) under contract with the Office of Planning, Budget Formulation, and Analysis, under the Energy Efficiency and Renewable Energy (EERE) program in the Department of Energy (DOE). Designed for use as a desk-top reference, the data book represents an assembly and display of statistics and information that characterize transportation activity, and presents data on other factors that influence transportation energy use. The purpose of this document is to present relevant statistical data in the form of tables and graphs. The latest editions of the Data Book are available to a larger audience via the Internet (cta.ornl.gov/data). This edition of the Data Book has 12 chapters which focus on various aspects of the transportation industry. Chapter 1 focuses on petroleum; Chapter 2 - energy; Chapter 3 - highway vehicles; Chapter 4 - light vehicles; Chapter 5 - heavy vehicles; Chapter 6 - alternative fuel vehicles; Chapter 7 - fleet vehicles; Chapter 8 - household vehicles; and Chapter 9- nonhighway modes; Chapter 10 - transportation and the economy; Chapter 11 - greenhouse gas emissions; and Chapter 12 - criteria pollutant emissions. The sources used represent the latest available data. There are also three appendices which include detailed source information for some tables, measures of conversion, and the definition of Census divisions and regions. A glossary of terms and a title index are also included for the readers convenience.

  11. Transportation Energy Data Book: Edition 28

    Energy Technology Data Exchange (ETDEWEB)

    Davis, Stacy Cagle [ORNL; Diegel, Susan W [ORNL; Boundy, Robert Gary [ORNL

    2009-06-01

    The Transportation Energy Data Book: Edition 28 is a statistical compendium prepared and published by Oak Ridge National Laboratory (ORNL) under contract with U.S Department of Energy, Office of Energy Efficiency and Renewable Energy, Vehicle Technologies Program and the Hydrogen, Fuel Cells, and Infrastructure Technologies Program. Designed for use as a desk-top reference, the data book represents an assembly and display of statistics and information that characterize transportation activity, and presents data on other factors that influence transportation energy use. The purpose of this document is to present relevant statistical data in the form of tables and graphs. The latest edition of the Data Book are available to a larger audience via the Internet (cta.ornl.gov/data). This edition of the Data Book has 12 chapters which focus on various aspects of the transportation industry. Chapter 1 focuses on petroleum; Chapter 2 energy; Chapter 3 highway vehicles; Chapter 4 light vehicles; Chapter 5 heavy vehicles; Chapter 6 alternative fuel vehicles; Chapter 7 fleet vehicles; Chapter 8 household vehicles; and Chapter 9 nonhighway modes; Chapter 10 transportation and the economy; Chapter 11 greenhouse gas emissions; and Chapter 12 criteria pollutant emissions. The sources used represent the latest available data. There are also three appendices which include detailed source information for some tables, measures of conversion, and the definition of Census divisions and regions. A glossary of terms and a title index are also included for the readers convenience.

  12. Transportation Energy Data Book: Edition 29

    Energy Technology Data Exchange (ETDEWEB)

    Davis, Stacy Cagle [ORNL; Diegel, Susan W [ORNL; Boundy, Robert Gary [ORNL

    2010-07-01

    The Transportation Energy Data Book: Edition 29 is a statistical compendium prepared and published by Oak Ridge National Laboratory (ORNL) under contract with the U.S. Department of Energy, Office of Energy Efficiency and Renewable Energy, Vehicle Technologies Program. Designed for use as a desk-top reference, the Data Book represents an assembly and display of statistics and information that characterize transportation activity, and presents data on other factors that influence transportation energy use. The purpose of this document is to present relevant statistical data in the form of tables and graphs. The latest edition of the Data Book is available to a larger audience via the Internet (cta.ornl.gov/data). This edition of the Data Book has 12 chapters which focus on various aspects of the transportation industry. Chapter 1 focuses on petroleum; Chapter 2 energy; Chapter 3 highway vehicles; Chapter 4 light vehicles; Chapter 5 heavy vehicles; Chapter 6 alternative fuel vehicles; Chapter 7 fleet vehicles; Chapter 8 household vehicles; Chapter 9 nonhighway modes; Chapter 10 transportation and the economy; Chapter 11 greenhouse gas emissions; and Chapter 12 criteria pollutant emissions. The sources used represent the latest available data. There are also three appendices which include detailed source information for some tables, measures of conversion, and the definition of Census divisions and regions. A glossary of terms and a title index are also included for the reader s convenience.

  13. Transportation Energy Data Book: Edition 21

    Energy Technology Data Exchange (ETDEWEB)

    Davis, S.C.

    2001-09-13

    The ''Transportation Energy Data Book: Edition 21'' is a statistical compendium prepared and published by Oak Ridge National Laboratory (ORNL) under contract with the Office of Transportation Technologies in the Department of Energy (DOE). Designed for use as a desk-top reference, the data book represents an assembly and display of statistics and information that characterize transportation activity, and presents data on other factors that influence transportation energy use. The purpose of this document is to present relevant statistical data in the form of tables and graphs. The latest editions of the Data Book are available to a larger audience via the Internet (www-cta.ornl.gov/data/tedb.htm). This edition of the Data Book has 12 chapters which focus on various aspects of the transportation industry. Chapter 1 focuses on petroleum; Chapter 2--energy; Chapter 3--greenhouse gas emissions; Chapter 4--criteria pollutant emissions; Chapter 5--transportation and the economy; Chapter 6--highway vehicles; Chapter 7--light vehicles; Chapter 8--heavy vehicles; Chapter 9--alternative fuel vehicles; Chapter 10--fleet vehicles; Chapter 11--household vehicles; and Chapter 12--nonhighway modes. The sources used represent the latest available data. There are also three appendices which include detailed source information for some tables, measures of conversion, and the definition of Census divisions and regions. A glossary of terms and a title index are also included for the readers convenience.

  14. Transportation Energy Data Book. Edition 33

    Energy Technology Data Exchange (ETDEWEB)

    Davis, Stacy Cagle [Oak Ridge National Lab. (ORNL), Oak Ridge, TN (United States); Williams, Susan E. [Oak Ridge National Lab. (ORNL), Oak Ridge, TN (United States); Boundy, Robert Gary [Oak Ridge National Lab. (ORNL), Oak Ridge, TN (United States)

    2014-07-01

    The Transportation Energy Data Book: Edition 33 is a statistical compendium prepared and published by Oak Ridge National Laboratory (ORNL) under contract with the U.S. Department of Energy, Office of Energy Efficiency and Renewable Energy, Vehicle Technologies Office. Designed for use as a desk-top reference, the Data Book represents an assembly and display of statistics and information that characterize transportation activity, and presents data on other factors that influence transportation energy use. The purpose of this document is to present relevant statistical data in the form of tables and graphs. The latest edition of the Data Book is available to a larger audience via the Internet (cta.ornl.gov/data). This edition of the Data Book has 12 chapters which focus on various aspects of the transportation industry. Chapter 1 focuses on petroleum; Chapter 2 energy; Chapter 3 highway vehicles; Chapter 4 light vehicles; Chapter 5 heavy vehicles; Chapter 6 alternative fuel vehicles; Chapter 7 fleet vehicles; Chapter 8 household vehicles; Chapter 9 nonhighway modes; Chapter 10 transportation and the economy; Chapter 11 greenhouse gas emissions; and Chapter 12 criteria pollutant emissions. The sources used represent the latest available data. There are also three appendices which include detailed source information for some tables, measures of conversion, and the definition of Census divisions and regions. A glossary of terms and a title index are also included for the reader s convenience.

  15. Transportation Energy Data Book: Edition 31

    Energy Technology Data Exchange (ETDEWEB)

    Davis, Stacy Cagle [ORNL; Diegel, Susan W [ORNL; Boundy, Robert Gary [ORNL

    2012-08-01

    The Transportation Energy Data Book: Edition 31 is a statistical compendium prepared and published by Oak Ridge National Laboratory (ORNL) under contract with the U.S. Department of Energy, Office of Energy Efficiency and Renewable Energy, Vehicle Technologies Program. Designed for use as a desk-top reference, the Data Book represents an assembly and display of statistics and information that characterize transportation activity, and presents data on other factors that influence transportation energy use. The purpose of this document is to present relevant statistical data in the form of tables and graphs. The latest edition of the Data Book is available to a larger audience via the Internet (cta.ornl.gov/data). This edition of the Data Book has 12 chapters which focus on various aspects of the transportation industry. Chapter 1 focuses on petroleum; Chapter 2 energy; Chapter 3 highway vehicles; Chapter 4 light vehicles; Chapter 5 heavy vehicles; Chapter 6 alternative fuel vehicles; Chapter 7 fleet vehicles; Chapter 8 household vehicles; Chapter 9 nonhighway modes; Chapter 10 transportation and the economy; Chapter 11 greenhouse gas emissions; and Chapter 12 criteria pollutant emissions. The sources used represent the latest available data. There are also three appendices which include detailed source information for some tables, measures of conversion, and the definition of Census divisions and regions. A glossary of terms and a title index are also included for the reader s convenience.

  16. Transportation Energy Data Book: Edition 32

    Energy Technology Data Exchange (ETDEWEB)

    Davis, Stacy Cagle [ORNL; Diegel, Susan W [ORNL; Boundy, Robert Gary [ORNL

    2013-08-01

    The Transportation Energy Data Book: Edition 32 is a statistical compendium prepared and published by Oak Ridge National Laboratory (ORNL) under contract with the U.S. Department of Energy, Office of Energy Efficiency and Renewable Energy, Vehicle Technologies Office. Designed for use as a desk-top reference, the Data Book represents an assembly and display of statistics and information that characterize transportation activity, and presents data on other factors that influence transportation energy use. The purpose of this document is to present relevant statistical data in the form of tables and graphs. The latest edition of the Data Book is available to a larger audience via the Internet (cta.ornl.gov/data). This edition of the Data Book has 12 chapters which focus on various aspects of the transportation industry. Chapter 1 focuses on petroleum; Chapter 2 energy; Chapter 3 highway vehicles; Chapter 4 light vehicles; Chapter 5 heavy vehicles; Chapter 6 alternative fuel vehicles; Chapter 7 fleet vehicles; Chapter 8 household vehicles; Chapter 9 nonhighway modes; Chapter 10 transportation and the economy; Chapter 11 greenhouse gas emissions; and Chapter 12 criteria pollutant emissions. The sources used represent the latest available data. There are also three appendices which include detailed source information for some tables, measures of conversion, and the definition of Census divisions and regions. A glossary of terms and a title index are also included for the reader s convenience.

  17. Transportation Energy Data Book: Edition 30

    Energy Technology Data Exchange (ETDEWEB)

    Davis, Stacy Cagle [ORNL; Diegel, Susan W [ORNL; Boundy, Robert Gary [ORNL

    2011-07-01

    The Transportation Energy Data Book: Edition 30 is a statistical compendium prepared and published by Oak Ridge National Laboratory (ORNL) under contract with the U.S. Department of Energy, Office of Energy Efficiency and Renewable Energy, Vehicle Technologies Program. Designed for use as a desk-top reference, the Data Book represents an assembly and display of statistics and information that characterize transportation activity, and presents data on other factors that influence transportation energy use. The purpose of this document is to present relevant statistical data in the form of tables and graphs. The latest edition of the Data Book is available to a larger audience via the Internet (cta.ornl.gov/data). This edition of the Data Book has 12 chapters which focus on various aspects of the transportation industry. Chapter 1 focuses on petroleum; Chapter 2 energy; Chapter 3 highway vehicles; Chapter 4 light vehicles; Chapter 5 heavy vehicles; Chapter 6 alternative fuel vehicles; Chapter 7 fleet vehicles; Chapter 8 household vehicles; Chapter 9 nonhighway modes; Chapter 10 transportation and the economy; Chapter 11 greenhouse gas emissions; and Chapter 12 criteria pollutant emissions. The sources used represent the latest available data. There are also three appendices which include detailed source information for some tables, measures of conversion, and the definition of Census divisions and regions. A glossary of terms and a title index are also included for the reader s convenience.

  18. Spacecraft Systems Engineering, 3rd Edition

    Science.gov (United States)

    Fortescue, Peter; Stark, John; Swinerd, Graham

    2003-03-01

    Following on from the hugely successful previous editions, the third edition of Spacecraft Systems Engineering incorporates the most recent technological advances in spacecraft and satellite engineering. With emphasis on recent developments in space activities, this new edition has been completely revised. Every chapter has been updated and rewritten by an expert engineer in the field, with emphasis on the bus rather than the payload. Encompassing the fundamentals of spacecraft engineering, the book begins with front-end system-level issues, such as environment, mission analysis and system engineering, and progresses to a detailed examination of subsystem elements which represent the core of spacecraft design - mechanical, electrical, propulsion, thermal, control etc. This quantitative treatment is supplemented by an appreciation of the interactions between the elements, which deeply influence the process of spacecraft systems design. In particular the revised text includes * A new chapter on small satellites engineering and applications which has been contributed by two internationally-recognised experts, with insights into small satellite systems engineering. * Additions to the mission analysis chapter, treating issues of aero-manouevring, constellation design and small body missions. In summary, this is an outstanding textbook for aerospace engineering and design students, and offers essential reading for spacecraft engineers, designers and research scientists. The comprehensive approach provides an invaluable resource to spacecraft manufacturers and agencies across the world.

  19. Transportation Energy Data Book: Edition 24

    Energy Technology Data Exchange (ETDEWEB)

    Davis, S.C.

    2005-03-08

    The ''Transportation Energy Data Book: Edition 24'' is a statistical compendium prepared and published by Oak Ridge National Laboratory (ORNL) under contract with the Office of Planning, Budget Formulation, and Analysis, under the Energy Efficiency and Renewable Energy (EERE) program in the Department of Energy (DOE). Designed for use as a desk-top reference, the data book represents an assembly and display of statistics and information that characterize transportation activity, and presents data on other factors that influence transportation energy use. The purpose of this document is to present relevant statistical data in the form of tables and graphs. The latest editions of the Data Book are available to a larger audience via the Internet (cta.ornl.gov/data). This edition of the Data Book has 12 chapters which focus on various aspects of the transportation industry. Chapter 1 focuses on petroleum; Chapter 2--energy; Chapter 3--highway vehicles; Chapter 4--light vehicles; Chapter 5--heavy vehicles; Chapter 6--alternative fuel vehicles; Chapter 7--fleet vehicles; Chapter 8--household vehicles; and Chapter 9--nonhighway modes; Chapter 10--transportation and the economy; Chapter 11--greenhouse gas emissions; and Chapter 12--criteria pollutant emissions. The sources used represent the latest available data. There are also three appendices which include detailed source information for some tables, measures of conversion, and the definition of Census divisions and regions. A glossary of terms and a title index are also included for the readers convenience.

  20. Transportation Energy Data Book: Edition 23

    Energy Technology Data Exchange (ETDEWEB)

    Davis, S.C.

    2003-10-24

    The ''Transportation Energy Data Book: Edition 23'' is a statistical compendium prepared and published by Oak Ridge National Laboratory (ORNL) under contract with the Office of Planning, Budget Formulation, and Analysis, under the Energy Efficiency and Renewable Energy (EERE) program in the Department of Energy (DOE). Designed for use as a desk-top reference, the data book represents an assembly and display of statistics and information that characterize transportation activity, and presents data on other factors that influence transportation energy use. The purpose of this document is to present relevant statistical data in the form of tables and graphs. The latest editions of the Data Book are available to a larger audience via the Internet (www-cta.ornl.gov/data). This edition of the Data Book has 12 chapters which focus on various aspects of the transportation industry. Chapter 1 focuses on petroleum; Chapter 2--energy; Chapter 3--highway vehicles; Chapter 4--light vehicles; Chapter 5--heavy vehicles; Chapter 6--alternative fuel vehicles; Chapter 7--fleet vehicles; Chapter 8--household vehicles; and Chapter 9--nonhighway modes; Chapter 10--transportation and the economy; Chapter 11--greenhouse gas emissions; and Chapter 12--criteria pollutant emissions. The sources used represent the latest available data. There are also three appendices which include detailed source information for some tables, measures of conversion, and the definition of Census divisions and regions. A glossary of terms and a title index are also included for the readers convenience.

  1. Transportation Energy Data Book: Edition 34

    Energy Technology Data Exchange (ETDEWEB)

    Davis, Stacy Cagle [Oak Ridge National Lab. (ORNL), Oak Ridge, TN (United States); Williams, Susan E [Oak Ridge National Lab. (ORNL), Oak Ridge, TN (United States); Boundy, Robert Gary [Roltek, Inc., Clinton, TN (United States)

    2015-08-01

    The Transportation Energy Data Book: Edition 34 is a statistical compendium prepared and published by Oak Ridge National Laboratory (ORNL) under contract with the U.S. Department of Energy, Office of Energy Efficiency and Renewable Energy, Vehicle Technologies Office. Designed for use as a desk-top reference, the Data Book represents an assembly and display of statistics and information that characterize transportation activity, and presents data on other factors that influence transportation energy use. The purpose of this document is to present relevant statistical data in the form of tables and graphs. The latest edition of the Data Book is available to a larger audience via the Internet (cta.ornl.gov/data). This edition of the Data Book has 12 chapters which focus on various aspects of the transportation industry. Chapter 1 focuses on petroleum; Chapter 2 energy; Chapter 3 highway vehicles; Chapter 4 light vehicles; Chapter 5 heavy vehicles; Chapter 6 alternative fuel vehicles; Chapter 7 fleet vehicles; Chapter 8 household vehicles; Chapter 9 nonhighway modes; Chapter 10 transportation and the economy; Chapter 11 greenhouse gas emissions; and Chapter 12 criteria pollutant emissions. The sources used represent the latest available data. There are also three appendices which include detailed source information for some tables, measures of conversion, and the definition of Census divisions and regions. A glossary of terms and a title index are also included for the reader s convenience.

  2. Next-Generation Sequencing and Genome Editing in Plant Virology

    Science.gov (United States)

    Hadidi, Ahmed; Flores, Ricardo; Candresse, Thierry; Barba, Marina

    2016-01-01

    Next-generation sequencing (NGS) has been applied to plant virology since 2009. NGS provides highly efficient, rapid, low cost DNA, or RNA high-throughput sequencing of the genomes of plant viruses and viroids and of the specific small RNAs generated during the infection process. These small RNAs, which cover frequently the whole genome of the infectious agent, are 21–24 nt long and are known as vsRNAs for viruses and vd-sRNAs for viroids. NGS has been used in a number of studies in plant virology including, but not limited to, discovery of novel viruses and viroids as well as detection and identification of those pathogens already known, analysis of genome diversity and evolution, and study of pathogen epidemiology. The genome engineering editing method, clustered regularly interspaced short palindromic repeats (CRISPR)-Cas9 system has been successfully used recently to engineer resistance to DNA geminiviruses (family, Geminiviridae) by targeting different viral genome sequences in infected Nicotiana benthamiana or Arabidopsis plants. The DNA viruses targeted include tomato yellow leaf curl virus and merremia mosaic virus (begomovirus); beet curly top virus and beet severe curly top virus (curtovirus); and bean yellow dwarf virus (mastrevirus). The technique has also been used against the RNA viruses zucchini yellow mosaic virus, papaya ringspot virus and turnip mosaic virus (potyvirus) and cucumber vein yellowing virus (ipomovirus, family, Potyviridae) by targeting the translation initiation genes eIF4E in cucumber or Arabidopsis plants. From these recent advances of major importance, it is expected that NGS and CRISPR-Cas technologies will play a significant role in the very near future in advancing the field of plant virology and connecting it with other related fields of biology.

  3. Next-Generation Sequencing and Genome Editing in Plant Virology.

    Science.gov (United States)

    Hadidi, Ahmed; Flores, Ricardo; Candresse, Thierry; Barba, Marina

    2016-01-01

    Next-generation sequencing (NGS) has been applied to plant virology since 2009. NGS provides highly efficient, rapid, low cost DNA, or RNA high-throughput sequencing of the genomes of plant viruses and viroids and of the specific small RNAs generated during the infection process. These small RNAs, which cover frequently the whole genome of the infectious agent, are 21-24 nt long and are known as vsRNAs for viruses and vd-sRNAs for viroids. NGS has been used in a number of studies in plant virology including, but not limited to, discovery of novel viruses and viroids as well as detection and identification of those pathogens already known, analysis of genome diversity and evolution, and study of pathogen epidemiology. The genome engineering editing method, clustered regularly interspaced short palindromic repeats (CRISPR)-Cas9 system has been successfully used recently to engineer resistance to DNA geminiviruses (family, Geminiviridae) by targeting different viral genome sequences in infected Nicotiana benthamiana or Arabidopsis plants. The DNA viruses targeted include tomato yellow leaf curl virus and merremia mosaic virus (begomovirus); beet curly top virus and beet severe curly top virus (curtovirus); and bean yellow dwarf virus (mastrevirus). The technique has also been used against the RNA viruses zucchini yellow mosaic virus, papaya ringspot virus and turnip mosaic virus (potyvirus) and cucumber vein yellowing virus (ipomovirus, family, Potyviridae) by targeting the translation initiation genes eIF4E in cucumber or Arabidopsis plants. From these recent advances of major importance, it is expected that NGS and CRISPR-Cas technologies will play a significant role in the very near future in advancing the field of plant virology and connecting it with other related fields of biology. PMID:27617007

  4. Efficient generation of gene-modified pigs via injection of zygote with Cas9/sgRNA

    OpenAIRE

    Yong Wang; Yinan Du; Bin Shen; Xiaoyang Zhou; Jian Li; Yu Liu; Jianying Wang; Jiankui Zhou; Bian Hu; Nannan Kang; Jimin Gao; Liqing Yu; Xingxu Huang; Hong Wei

    2015-01-01

    Co-injection of zygotes with Cas9 mRNA and sgRNA has been proven to be an efficient gene-editing strategy for genome modification of different species. Genetic engineering in pigs holds a great promise in biomedical research. By co-injection of one-cell stage embryos with Cas9 mRNA and Npc1l1 sgRNA, we achieved precise Npc1l1 targeting in Chinese Bama miniature pigs at the efficiency as high as 100%. Meanwhile, we carefully analyzed the Npc1l1 sgRNA:Cas9-mediated on- and off-target mutations ...

  5. Successful Transient Expression of Cas9 and Single Guide RNA Genes in Chlamydomonas reinhardtii

    OpenAIRE

    Jiang, Wenzhi; Brueggeman, Andrew J.; Horken, Kempton M.; Plucinak, Thomas M.; Weeks, Donald P.

    2014-01-01

    The clustered regularly interspaced short palindromic repeat (CRISPR)/Cas9 system has become a powerful and precise tool for targeted gene modification (e.g., gene knockout and gene replacement) in numerous eukaryotic organisms. Initial attempts to apply this technology to a model, the single-cell alga, Chlamydomonas reinhardtii, failed to yield cells containing edited genes. To determine if the Cas9 and single guide RNA (sgRNA) genes were functional in C. reinhardtii, we tested the ability o...

  6. Genome editing in Ustilago maydis using the CRISPR-Cas system.

    Science.gov (United States)

    Schuster, Mariana; Schweizer, Gabriel; Reissmann, Stefanie; Kahmann, Regine

    2016-04-01

    This communication describes the establishment of the type II bacterial CRISPR-Cas9 system to efficiently disrupt target genes in the fungal maize pathogen Ustilago maydis. A single step transformation of a self-replicating plasmid constitutively expressing the U. maydis codon-optimized cas9 gene and a suitable sgRNA under control of the U. maydis U6 snRNA promoter was sufficient to induce genome editing. On average 70% of the progeny of a single transformant were disrupted within the respective b gene. Without selection the self-replicating plasmid was lost rapidly allowing transient expression of the CRISPR-Cas9 system to minimize potential long-term negative effects of Cas9. This technology will be an important advance for the simultaneous disruption of functionally redundant genes and gene families to investigate their contribution to virulence of U. maydis. PMID:26365384

  7. ADAR2 editing enzyme is a novel human immunodeficiency virus-1 proviral factor.

    Science.gov (United States)

    Doria, Margherita; Tomaselli, Sara; Neri, Francesca; Ciafrè, Silvia Anna; Farace, Maria Giulia; Michienzi, Alessandro; Gallo, Angela

    2011-05-01

    The adenosine deaminases acting on RNA (ADAR) enzymes catalyse conversion of adenosine to inosine in dsRNA. A positive effect of ADAR1 on human immunodeficiency virus type 1 (HIV-1) replication has recently been reported. Here, we show that another ADAR enzyme, ADAR2, positively affects the replication process of HIV-1. We found that, analogously to ADAR1, ADAR2 enhances the release of progeny virions by an editing-dependent mechanism. However, differently from the ADAR1 enzyme, ADAR2 does not increase the infectious potential of the virus. Importantly, downregulation of ADAR2 in Jurkat cells significantly impairs viral replication. Therefore, ADAR2 shares some but not all proviral functions of ADAR1. These results suggest a novel role of ADAR2 as a viral regulator. PMID:21289159

  8. Single primer-mediated circular polymerase chain reaction for hairpin DNA cloning and plasmid editing.

    Science.gov (United States)

    Huang, Jiansheng; Khan, Inamullah; Liu, Rui; Yang, Yan; Zhu, Naishuo

    2016-05-01

    We developed and validated a universal polymerase chain reaction (PCR) method, single primer circular (SPC)-PCR, using single primer to simultaneously insert and amplify a short hairpin sequence into a vector with a high success rate. In this method, the hairpin structure is divided into two parts and fused into a vector by PCR. Then, a single primer is used to cyclize the chimera into a mature short hairpin RNA (shRNA) expression vector. It is not biased by loop length or palindromic structures. Six hairpin DNAs with short 4-nucleotide loops were successfully cloned. Moreover, SPC-PCR was also applied to plasmid editing within 3 h with a success rate higher than 95%. PMID:26792375

  9. [Application of CRISPR-Cas9 genome editing for constructing animal models of human diseases].

    Science.gov (United States)

    Ou, Zhanhui; Sun, Xiaofang

    2016-08-10

    The CRISPR-Cas9 system is a new targeted nuclease for genome editing, which can directly introduce modifications at the targeted genomic locus. The system utilizes a short single guide RNA (sgRNA) to direct the endonuclease Cas9 in the genome. Upon targeting, Cas9 can generate DNA double-strand breaks (DSBs). As such DSBs are repaired by non-homologous end joining (NHEJ) or homology directed repair (HDR), therefore facilitates introduction of random or specific mutations, repair of endogenous mutations, or insertion of DNA elements. The system has been successfully used to generate gene targeted cell lines including those of human, animals and plants. This article reviews recent advances made in this rapidly evolving technique for the generation of animal models for human diseases. PMID:27455021

  10. The levels of edit. [technical writing in science

    Science.gov (United States)

    Vanburen, R.; Buehler, M. F.; Wallenbrock, D. (Editor)

    1976-01-01

    The editorial process is analyzed, and five levels of edit are identified. These levels represent cumulative combinations of nine types of edit: (1) coordination, (2) policy, (3) integrity, (4) screening, (5) copy clarification, (6) Mechanical Style, (7) Language, and (9) substantive. The levels and types of edit, although developed for specific use with external reports at the Jet Propulsion Laboratory, cover the general range of technical editing, especially as it applies to an in-house technical publications organization. Each type of edit is set forth in terms of groups of actions to be performed by the editor. The edit-level concept has enhanced understanding and communication among editors, authors, and publications managers concerning the specific editorial work to be done on each manuscript. It has also proved useful as a management tool for estimating and monitoring cost.

  11. Long Distance Revisions in Drafting and Post-editing

    OpenAIRE

    Carl, Michael; Kay, Martin; Jensen, Kristian T. H.

    2010-01-01

    This paper investigates properties of translation processes, as observed in the translation behaviour of student and professional translators. The translation process can be divided into a gisting, drafting and post-editing phase. We find that student translators have longer gisting phases whereas professional translators have longer post-editing phases. Long-distance revisions, which would typically be expected during post-editing, occur to the same extent during drafting a...

  12. Fact book on aging in British Columbia, 5th Edition

    OpenAIRE

    Wister, Andrew V; Sixsmith, Andrew; Adams, Raymond G.; Sinden, Danielle

    2009-01-01

    This fifth edition of the Fact Book updates the figures with data from the 2006 census and supplemental sources. Like the first four editions of the Fact Book, this edition references all source documents and presents data primarily in numeric tabular form. Topics covered in this document are listed in the Table of Contents and include: the size and historical rate of growth of the elderly population of British Columbia; trends in life expectancy, mortality rates and causes of death; the mari...

  13. Fact book on aging in British Columbia, 4th Edition

    OpenAIRE

    Wister, Andrew V; Gutman, Gloria M.; Adams, Raymond G.; Chou, Pak Hei Benedito

    2006-01-01

    This fourth edition of the Fact Book updates the figures with data from the 2001 census and supplemental sources. Like the first three editions of the Fact Book, the fourth edition references all source documents and presents data primarily in numeric tabular form. Topics covered in this document are listed in the Table of Contents and include: the size and historical rate of growth of the elderly population of British Columbia; trends in life expectancy, mortality rates and causes of death; ...

  14. Tree edit distance as a baseline approach for paraphrase representation

    OpenAIRE

    Vila Rigat, Marta; Dras, Mark

    2012-01-01

    Finding an adequate paraphrase representation formalism is a challenging issue in Natural Language Processing. In this paper, we analyse the performance of Tree Edit Distance as a paraphrase representation baseline. Our experiments using Edit Distance Textual Entailment Suite show that, as Tree Edit Distance consists of a purely syntactic approach, paraphrase alternations not based on structural reorganizations do not find an adequate representation. They also show that there is much scope fo...

  15. Learning Stochastic Edit Distance: application in handwritten character recognition

    OpenAIRE

    Oncina, Jose; Sebban, Marc

    2006-01-01

    Many pattern recognition algorithms are based on the nearest neighbour search and use the well known edit distance, for which the primitive edit costs are usually fixed in advance. In this article, we aim at learning an unbiased stochastic edit distance in the form of a finite-state transducer from a corpus of (input,output) pairs of strings. Contrary to the other standard methods, which generally use the Expectation Maximisation algorithm, our algorithm learns a transducer independently on t...

  16. 5. Defining Electronic Editions: A Historical and Functional Perspective

    OpenAIRE

    Vanhoutte, Edward

    2013-01-01

    1. Introduction Since Peter Robinson published the first electronic edition in a series designed to accommodate all of Chaucer’s Canterbury Tales, he has been theorizing and writing about the nature and definition of electronic editions. Along with Jerome McGann and others, he has been at the centre of the debate on electronic textual editing since he began work on the collation and textual criticism of Icelandic manuscripts, for which he developed programs collectively called Collate. Becaus...

  17. On the graph edit distance cost: Properties and applications

    OpenAIRE

    Solé Ribalta, Albert; Serratosa Casanelles, Francesc; Sanfeliu Cortés, Alberto

    2012-01-01

    We model the edit distance as a function in a labelling space. A labelling space is an Euclidean space where coordinates are the edit costs. Through this model, we define a class of cost. A class of cost is a region in the labelling space that all the edit costs have the same optimal labelling. Moreover, we characterise the distance value through the labelling space. This new point of view of the edit distance gives as the opportunity of defining some interesting properties that are useful fo...

  18. Bibliography of integral charged particle nuclear data. Archival edition

    International Nuclear Information System (INIS)

    This is the fourth annual edition of the National Nuclear Data Center charged-particle bibliography. This edition is cumulative and supersedes the previous editions. The bibliography's primary aims are to satisfy the need for a concise and comprehensive index of integral charged-particle cross section data and to provide an index of charged-particle data compiled in the international exchange format, EXFOR. This part of the publication deals with isotope production; references are ordered by mass of the nuclide produced. The present publication is an archival volume; future publications will be cumulative supplements to this edition

  19. Bibliography of integral charged particle nuclear data. Archival edition

    International Nuclear Information System (INIS)

    This is the fourth annual edition of the National Nuclear Data Center charged-particle bibliography. This edition is cumulative and supersedes the previous editions. The bibliography's primary aims are to satisfy the need for a concise and comprehensive index of integral charged-particle cross section data and to provide an index of charged-particle data compiled in the international exchange format. References in this Part are by target for the various incident charged particles (in order of increasing A). The present publication is an archival volume; future publications will be cumulative supplements to this edition

  20. Analyzing MiRNA-LncRNA Interactions.

    Science.gov (United States)

    Paraskevopoulou, Maria D; Hatzigeorgiou, Artemis G

    2016-01-01

    Long noncoding RNAs (lncRNAs) are noncoding transcripts usually longer than 200 nts that have recently emerged as one of the largest and significantly diverse RNA families. The biological role and functions of lncRNAs are still mostly uncharacterized. Their target-mimetic, sponge/decoy function on microRNAs was recently uncovered. miRNAs are a class of noncoding RNA species (~22 nts) that play a central role in posttranscriptional regulation of protein coding genes by mRNA cleavage, direct translational repression and/or mRNA destabilization. LncRNAs can act as miRNA sponges, reducing their regulatory effect on mRNAs. This function introduces an extra layer of complexity in the miRNA-target interaction network. This chapter focuses on the study of miRNA-lncRNA interactions with either in silico or experimentally supported analyses. The proposed methodologies can be appropriately adapted in order to become the backbone of advanced multistep functional miRNA analyses. PMID:26721498