scholarly journals A resource of targeted mutant mouse lines for 5,061 genes

2019 ◽  
Author(s):  
Marie-Christine Birling ◽  
Atsushi Yoshiki ◽  
David J Adams ◽  
Shinya Ayabe ◽  
Arthur L Beaudet ◽  
...  
Keyword(s):  
Mutant Mouse ◽  
Embryonic Stem ◽  
Null Alleles ◽  
Mouse Mutant ◽  
International Mouse Phenotyping Consortium ◽  
Number Of Genes ◽  
Mutant Strains ◽  
Community Effort ◽  
Mutant Mouse Line ◽  
Public Repositories

AbstractThe International Mouse Phenotyping Consortium reports the generation of new mouse mutant strains for over 5,000 genes from targeted embryonic stem cells on the C57BL/6N genetic background. This includes 2,850 null alleles for which no equivalent mutant mouse line exists, 2,987 novel conditional-ready alleles, and 4,433 novel reporter alleles. This nearly triples the number of genes with reporter alleles and almost doubles the number of conditional alleles available to the scientific community. When combined with more than 30 years of community effort, the total mutant allele mouse resource covers more than half of the genome. The extensively validated collection is archived and distributed through public repositories, facilitating availability to the worldwide biomedical research community, and expanding our understanding of gene function and human disease.

scholarly journals The production of 4,182 mouse lines identifies experimental and biological variables impacting Cas9-mediated mutant mouse line production

2021 ◽  
Author(s):  
Hillary Elrick ◽  
Kevin A. Peterson ◽  
Joshua A. Wood ◽  
Denise G. Lanza ◽  
Elif F. Acar ◽  
...  
Keyword(s):  
Best Practice ◽  
Null Alleles ◽  
Mouse Line ◽  
Protein Coding ◽  
Line Production ◽  
International Mouse Phenotyping Consortium ◽  
Biological Variables ◽  
Mutant Mouse Line ◽  
Mouse Lines

AbstractThe International Mouse Phenotyping Consortium (IMPC) is generating and phenotyping null mutations for every protein-coding gene in the mouse1,2. The IMPC now uses Cas9, a programmable RNA-guided nuclease that has revolutionized mouse genome editing3 and increased capacity and flexibility to efficiently generate null alleles in the C57BL/6N strain. In addition to being a valuable novel and accessible research resource, the production of >3,300 knockout mouse lines using comparable protocols provides a rich dataset to analyze experimental and biological variables affecting in vivo null allele engineering with Cas9. Mouse line production has two critical steps – generation of founders with the desired allele and germline transmission (GLT) of that allele from founders to offspring. Our analysis identified that whether a gene is essential for viability was the primary factor influencing successful production of null alleles. Collectively, our findings provide best practice recommendations for generating null alleles in mice using Cas9; these recommendations may be applicable to other allele types and species.

scholarly journals Progress towards completing the mutant mouse null resource

2021 ◽  
Author(s):  
Kevin A. Peterson ◽  
Stephen A. Murray
Keyword(s):  
Large Scale ◽  
Null Allele ◽  
Mutant Mouse ◽  
Embryonic Stem ◽  
Pooled Analysis ◽  
Null Alleles ◽  
Scale Production ◽  
Protein Coding ◽  
Protein Coding Genes ◽  
Human Orthologs

AbstractThe generation of a comprehensive catalog of null alleles covering all protein-coding genes is the goal of the International Mouse Phenotyping Consortium. Over the past 20 years, significant progress has been made towards achieving this goal through the combined efforts of many large-scale programs that built an embryonic stem cell resource to generate knockout mice and more recently employed CRISPR/Cas9-based mutagenesis to delete critical regions predicted to result in frameshift mutations, thus, ablating gene function. The IMPC initiative builds on prior and ongoing work by individual research groups creating gene knockouts in the mouse. Here, we analyze the collective efforts focusing on the combined null allele resource resulting from strains developed by the research community and large-scale production programs. Based upon this pooled analysis, we examine the remaining fraction of protein-coding genes focusing on clearly defined mouse–human orthologs as the highest priority for completing the mutant mouse null resource. In summary, we find that there are less than 3400 mouse–human orthologs remaining in the genome without a targeted null allele that can be further prioritized to achieve our overall goal of the complete functional annotation of the protein-coding portion of a mammalian genome.

scholarly journals TspO as a Modulator of the Repressor/Antirepressor (PpsR/AppA) Regulatory System in Rhodobacter sphaeroides 2.4.1

2001 ◽  
Vol 183 (21) ◽  
pp. 6355-6364 ◽  
Author(s):  
Xiaohua Zeng ◽  
Samuel Kaplan
Keyword(s):  
Rhodobacter Sphaeroides ◽  
Sinorhizobium Meliloti ◽  
Target Gene ◽  
Regulatory System ◽  
Regulatory Proteins ◽  
Number Of Genes ◽  
Mutant Strains ◽  
Site Of Action ◽  
De Bruijn ◽  
Recent Extension

ABSTRACT The TspO outer membrane protein of Rhodobacter sphaeroides has been shown to be involved in controlling the transcription of a number of genes which encode enzymes involved in photopigment biosynthesis and the puc operon. The display of regulated genes appears identical to those genes encompassing the PpsR/AppA repressor/antirepressor regulon, although the effect of TspO is modest relative to that of PpsR/AppA. To directly address the hypothesis that TspO is effective through the PpsR/AppA system, we constructed mutant strains with mutations in bothtspO and appA. In all cases, the phenotypes examined resembled those of the appA lesion by itself, leading us to conclude that TspO works through or modulates the PpsR/AppA system and acts upstream of the site of action of these regulatory proteins. In earlier publications, we had suggested that TspO is involved in the efflux of a certain intermediate(s) of the porphyrin biosynthesis pathway and that transcriptional regulation of target gene expression could be explained by the accumulation of a coactivator of AppA function. Although the data reported here do not precisely identify this coactivator, they lend support to this hypothesis. We discuss the importance of this form of gene control as the result of the recent extension of the TspO system toSinorhizobium meliloti, as described by Davey and de Bruijn (M. E. Davey and F. J. de Bruijn, Appl. Environ. Microbiol. 66:5353–5359, 2000). It is therefore possible that this system constitutes a more widely, although not universally, demonstrated form of gene regulation.

scholarly journals An efficient method for generation of bi-allelic null mutant mouse embryonic stem cells and its application for investigating epigenetic modifiers

2017 ◽  
Vol 45 (21) ◽  
pp. e174-e174 ◽  
Author(s):  
Cynthia L. Fisher ◽  
Hendrik Marks ◽  
Lily Ting-yin Cho ◽  
Robert Andrews ◽  
Sam Wormald ◽  
...  
Keyword(s):  
Stem Cells ◽  
Embryonic Stem Cells ◽  
Efficient Method ◽  
Mutant Mouse ◽  
Embryonic Stem ◽  
Mouse Embryonic Stem Cells ◽  
Null Mutant ◽  
Epigenetic Modifiers ◽  
Null Mutant Mouse

Phenotypes associated with psychiatric disorders are sex-specific in a mutant mouse line

2016 ◽  
Vol 1652 ◽  
pp. 53-61 ◽  
Author(s):  
Mitsunari Nakajima ◽  
Takashi Watanabe ◽  
Rui Aoki ◽  
Risei Shimizu ◽  
Satoshi Okuyama ◽  
...  
Keyword(s):  
Psychiatric Disorders ◽  
Mutant Mouse ◽  
Mouse Line ◽  
Mutant Mouse Line

scholarly journals Inhibition of N-myristoyltransferase Promotes Naive Pluripotency in Mouse and Human Pluripotent Stem Cells

2021 ◽  
Author(s):  
Junko Yoshida ◽  
Hitomi Watanabe ◽  
Kaori Yamauchi ◽  
Takumi Nishikubo ◽  
Ayako Isotani ◽  
...  
Keyword(s):  
Stem Cells ◽  
Pluripotent Stem Cells ◽  
Mutant Mouse ◽  
Embryonic Stem ◽  
Epiblast Stem Cells ◽  
State Marker ◽  
Homozygous Mutant Mouse ◽  
Novel Target ◽  
Induced Pluripotent

Naive and primed states are distinct states of pluripotency during early embryonic development that can be captured and converted to each other in vitro. To elucidate the regulatory mechanism of pluripotency, we performed a recessive genetic screen of homozygous mutant mouse embryonic stem cells (mESCs) and found that suppression of N-myristoyltransferase (Nmt) promotes naive pluripotency. Disruption of Nmt1 in mESCs conferred resistance to differentiation. Suppression of Nmt in mouse epiblast stem cells (mEpiSCs) promoted the conversion from the primed to the naive state. This effect was independent of Src, which is a major substrate of Nmt and is known to promote differentiation of mESCs. Suppression of Nmt in naive-state human induced pluripotent stem cells (hiPSCs) increased the expression of the naive-state marker. These results indicate that Nmt is a novel target for the regulation of naive pluripotency conserved between mice and humans.

scholarly journals A Genomewide Screen for Tolerance to Cationic Drugs Reveals Genes Important for Potassium Homeostasis in Saccharomyces cerevisiae

2011 ◽  
Vol 10 (9) ◽  
pp. 1241-1250 ◽  
Author(s):  
Lina Barreto ◽  
David Canadell ◽  
Silvia Petrezsélyová ◽  
Clara Navarrete ◽  
Lydie Marešová ◽  
...  
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Potassium Concentration ◽  
Growth Defect ◽  
Electrochemical Gradient ◽  
Biochemical Tests ◽  
Content Type ◽  
Potassium Homeostasis ◽  
Moderate Growth ◽  
Number Of Genes ◽  
Mutant Strains

ABSTRACTPotassium homeostasis is crucial for living cells. In the yeastSaccharomyces cerevisiae, the uptake of potassium is driven by the electrochemical gradient generated by the Pma1 H+-ATPase, and this process represents a major consumer of the gradient. We considered that any mutation resulting in an alteration of the electrochemical gradient could give rise to anomalous sensitivity to any cationic drug independently of its toxicity mechanism. Here, we describe a genomewide screen for mutants that present altered tolerance to hygromycin B, spermine, and tetramethylammonium. Two hundred twenty-six mutant strains displayed altered tolerance to all three drugs (202 hypersensitive and 24 hypertolerant), and more than 50% presented a strong or moderate growth defect at a limiting potassium concentration (1 mM). Functional groups such as protein kinases and phosphatases, intracellular trafficking, transcription, or cell cycle and DNA processing were enriched. Essentially, our screen has identified a substantial number of genes that were not previously described to play a direct or indirect role in potassium homeostasis. A subset of 27 representative mutants were selected and subjected to diverse biochemical tests that, in some cases, allowed us to postulate the basis for the observed phenotypes.

scholarly journals Arginine methylation of METTL14 promotes RNA N6-methyladenosine modification and endoderm differentiation of mouse embryonic stem cells

2021 ◽  
Vol 12 (1) ◽  
Author(s):  
Xiaona Liu ◽  
Hailong Wang ◽  
Xueya Zhao ◽  
Qizhi Luo ◽  
Qingwen Wang ◽  
...  
Keyword(s):  
Stem Cells ◽  
Embryonic Stem Cells ◽  
Mutant Mouse ◽  
Embryonic Stem ◽  
Arginine Methylation ◽  
Mouse Embryonic Stem Cells ◽  
Protein Methylation ◽  
Dynamic Regulation ◽  
Endoderm Differentiation

AbstractRNA N6-methyladenosine (m6A), the most abundant internal modification of mRNAs, plays key roles in human development and health. Post-translational methylation of proteins is often critical for the dynamic regulation of enzymatic activity. However, the role of methylation of the core methyltransferase METTL3/METTL14 in m6A regulation remains elusive. We find by mass spectrometry that METTL14 arginine 255 (R255) is methylated (R255me). Global mRNA m6A levels are greatly decreased in METTL14 R255K mutant mouse embryonic stem cells (mESCs). We further find that R255me greatly enhances the interaction of METTL3/METTL14 with WTAP and promotes the binding of the complex to substrate RNA. We show that protein arginine N-methyltransferases 1 (PRMT1) interacts with and methylates METTL14 at R255, and consistent with this, loss of PRMT1 reduces mRNA m6A modification globally. Lastly, we find that loss of R255me preferentially affects endoderm differentiation in mESCs. Collectively, our findings show that arginine methylation of METTL14 stabilizes the binding of the m6A methyltransferase complex to its substrate RNA, thereby promoting global m6A modification and mESC endoderm differentiation. This work highlights the crosstalk between protein methylation and RNA methylation in gene expression.

Recombination of bacteriophage λ in recD mutants of Escherichia coli

Genome ◽  
1989 ◽  
Vol 31 (1) ◽  
pp. 53-67 ◽  
Author(s):  
David S. Thaler ◽  
Elizabeth Sampson ◽  
Imran Siddiqi ◽  
Susan M. Rosenberg ◽  
Lynn C. Thomason ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Dna Replication ◽  
Homologous Recombination ◽  
Restriction Enzyme ◽  
Null Alleles ◽  
Mutant Strains ◽  
Recbcd Enzyme ◽  
Mutant Cells ◽  
D Subunit

RecBCD enzyme is centrally important in homologous recombination in Escherichia coli and is the source of ExoV activity. Null alleles of either the recB or the recC genes, which encode the B and C subunits, respectively, manifest no recombination and none of the nuclease functions characteristic of the holoenzyme. Loss of the D subunit, by a recD mutation, likewise results in loss of ExoV activity. However, mutants lacking the D subunit are competent for homologous recombination. We report that the distribution of exchanges along the chromosome of Red−Gam−phage λ is strikingly altered by recD null mutations in the host. When λ DNA replication is blocked, recombination in recD mutant strains is high near λ's right end. In contrast, recombination in isogenic recD+ strains is approximately uniform along λ unless the λ chromosome contains a χ sequence. Recombination in recD mutant strains is focused toward the site of action of a type II restriction enzyme acting in vivo on λ. The distribution of exchanges in isogenic recD+ strains is scarcely altered by the restriction enzyme (unless the phage contains an otherwise silent χ). The distribution of exchanges in recD mutants is strongly affected by λ DNA replication. The distribution of exchanges on λ growing in rec+ cells is not influenced by DNA replication. The exchange distribution along λ in recD mutant cells is independent of χ in a variety of conditions. Recombination in rec+ cells is χ influenced. Recombination in recD mutants depends on recC function, occurs in strains deleted for rac prophage, and is independent of recJ, which is known to be required for λ recombination via the RecF pathway. We entertain two models for recombination in recD mutants: (i) recombination in recD mutants may proceed via double-chain break–repair, as it does in λ's Red pathway and E. coli's RecE pathway; (ii) the RecBC enzyme, missing its D subunit, is equivalent to the wild-type, RecBCD, enzyme after that enzyme has been activated by a χ sequence.Key words: χ sequence, RecBCD pathway, Red pathway, RecBC‡ pathway.

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