Large-Scale Screening for Targeted Knockouts in the Caenorhabditis elegans Genome

Abstract

The nematode Caenorhabditis elegans is a powerful model system to study contemporary biological problems. This system would be even more useful if we had mutations in all the genes of this multicellular metazoan. The combined efforts of the C. elegans Deletion Mutant Consortium and individuals within the worm community are moving us ever closer to this goal. At present, of the 20,377 protein-coding genes in this organism, 6764 genes with associated molecular lesions are either deletions or null mutations (WormBase WS220). Our three laboratories have contributed the majority of mutated genes, 6841 mutations in 6013 genes. The principal method we used to detect deletion mutations in the nematode utilizes polymerase chain reaction (PCR). More recently, we have used array comparative genome hybridization (aCGH) to detect deletions across the entire coding part of the genome and massively parallel short-read sequencing to identify nonsense, splicing, and missense defects in open reading frames. As deletion strains can be frozen and then thawed when needed, these strains will be an enduring community resource. Our combined molecular screening strategies have improved the overall throughput of our gene-knockout facilities and have broadened the types of mutations that we and others can identify. These multiple strategies should enable us to eventually identify a mutation in every gene in this multicellular organism. This knowledge will usher in a new age of metazoan genetics in which the contribution to any biological process can be assessed for all genes.

Shown for reading. Its publisher allows quoting it, not republishing it.

References 62

1. 1.

   Altun Z. F. Chen B. Wang Z. W. Hall D. H., 2009 High resolution map of Caenorhabditis elegans gap junction proteins. Dev. Dyn. 238: 1936–1950

   Not in our library10.1002/dvdy.22025

2. 2.

   Ambros V., 1997 Heterochronic genes, C. elegans II, Ed. 2, edited by Riddle D. L. Blumenthal T. Meyer B. J. Priess J. R. Cold Spring Harbor Laboratory Press, Cold Spring Harbor, NY

   Not in our library

3. 3.

   Ambros V., 2004 The functions of animal microRNAs. Nature 431: 350–355

   Not in our library10.1038/nature02871

4. 4.

   Antebi A., 2006 Nuclear hormone receptors in C. elegans (January 3, 2006), WormBook, ed. The C. elegans Research Community WormBook, /10.1895/wormbook.1.64.1, http://www.wormbook.org

   Not in our library10.1895/wormbook.1.64.1

5. 5.

   Ashrafi K. Chang F. Y. Watts J. L. Fraser A. G. Kamath R. S., 2003 Genome-wide RNAi analysis of Caenorhabditis elegans fat regulatory genes. Nature 421: 268–272

   Not in our library10.1038/nature01279

6. 6.

   Avery L., 1993 The genetics of feeding in Caenorhabditis elegans. Genetics 133: 897–917

   Not in our library10.1093/genetics/133.4.897

7. 7.

   Bargmann C. I., 1998 Neurobiology of the Caenorhabditis elegans genome. Science 282: 2028–2033

   Not in our library10.1126/science.282.5396.2028

8. 8.

   Barstead R., 2001 Genome-wide RNAi. Curr. Opin. Chem. Biol. 5: 63–66

   Not in our library10.1016/s1367-5931(00)00173-3

9. 9.

   Barstead R. J. Moerman D. G., 2006 C. elegans deletion mutant screening. Methods Mol. Biol. 351: 51–58

   Not in our library10.1385/1-59745-151-7:51

10. 10.

    Bessereau J.-L., 2006 Transposons in C. elegans (January 18, 2006), WormBook, ed. The C. elegans Research Community WormBook, /10.1895/wormbook.1.70.1, http://www.wormbook.org

    Not in our library

11. 11.

    Boch J., 2011 TALEs of genome targeting. Nat. Biotechnol. 29: 135–136

    Not in our library10.1038/nbt.1767

12. 12.

    Bogdanove A. J. Voytas D. F., 2011 TAL effectors: customizable proteins for DNA targeting. Science 333: 1843–1846

    Not in our library10.1126/science.1204094

13. 13.

    Brenner S., 1974 The genetics of Caenorhabditis elegans. Genetics 77: 71–94

    Not in our library10.1093/genetics/77.1.71

14. 14.

    C. elegans Sequencing Consortium, 1998 Genome sequence of the nematode C. elegans: a platform for investigating biology. Science 282: 2012–2018

    Not in our library10.1126/science.282.5396.2012

15. 15.

    Cermak T. Doyle E. L. Christian M. Wang L. Zhang Y., 2011 Efficient design and assembly of custom TALEN and other TAL effector-based constructs for DNA targeting. Nucleic Acids Res. 39: e82

    Not in our library10.1093/nar/gkr218

16. 16.

    Chervitz S. A. Aravind L. Sherlock G. Ball C. A. Koonin E. V., 1998 Comparison of the complete protein sets of worm and yeast: orthology and divergence. Science 282: 2022–2028

    Not in our library10.1126/science.282.5396.2022

17. 17.

    Edgley M. L. Baillie D. L. Riddle D. l. Rose A. M., 1995 Genetic Balancers. Methods Cell Biol. 48: 147–184

    Not in our library

18. 18.

    Edgley M. L. D’Souza A. Moulder G. McKay S. Shen B., 2002 Improved detection of small deletions in complex pools of DNA. Nucleic Acids Res. 30: e52

    Not in our library10.1093/nar/gnf051

19. 19.

    Fire A. Xu S. Montgomery M. K. Kostas S. A. Driver S. E., 1998 Potent and specific genetic interference by double-stranded RNA in Caenorhabditis elegans. Nature 391: 806–811

    Not in our library10.1038/35888

20. 20.

    Flibotte S. Edgley M. L. Chaudry I. Taylor J. Neil S. E., 2010 Whole-genome profiling of mutagenesis in Caenorhabditis elegans. Genetics 185: 431–441

    Not in our library10.1534/genetics.110.116616

21. 21.

    Fraser A. G. Kamath R. S. Zipperlen P. Martinez-Campos M. Sohrmann M., 2000 Functional genomic analysis of C. elegans chromosome I by systematic RNA interference. Nature 408: 325–330

    Not in our library10.1038/35042517

22. 22.

    Frøkjaer-Jensen C. Davis M. W. Hopkins C. E. Newman B. J. Thummel J. M., 2008 Single-copy insertion of transgenes in Caenorhabditis elegans. Nat. Genet. 40: 1375–1383

    Not in our library10.1038/ng.248

23. 23.

    Frøkjaer-Jensen C. Davis M. W. Hollopeter G. Taylor J. Harris T. W., 2010 Targeted gene deletions in C. elegans using transposon excision. Nat. Methods 7: 451–453

    Not in our library10.1038/nmeth.1454

24. 24.

    Gengyo-Ando K. Mitani S., 2000 Characterization of mutations induced by ethyl methanesulfonate, UV, and trimethylpsoralen in the nematode Caenorhabditis elegans. Biochem. Biophys. Res. Commun. 269: 64–69

    Not in our library10.1006/bbrc.2000.2260

25. 25.

    Giaever G. Chu A. M. Ni L. Connelly C. Riles L., 2002 Functional profiling of the Saccharomyces cerevisiae genome. Nature 418: 387–391

    Not in our library10.1038/nature00935

26. 26.

    Gönczy P. Echeverri C. Oegema K. Coulson A. Jones S. J., 2000 Functional genomic analysis of cell division in C. elegans using RNAi of genes on chromosome III. Nature 408: 331–336

    Not in our library10.1038/35042526

27. 27.

    Goodman M. B., 2006 Mechanosensation (January 6, 2006), WormBook, ed. The C. elegans Research Community WormBook, /10.1895/wormbook.1.62.1, http://www.wormbook.org

    Not in our library

28. 28.

    Hutter H. Ng M. P. Chen N., 2009 GExplore: a web server for integrated queries of protein domains, gene expression and mutant phenotypes. BMC Genomics 10: 529

    Not in our library10.1186/1471-2164-10-529

29. 29.

    Jansen G. Hazendonk E. Thijssen K. L. Plasterk R. H., 1997 Reverse genetics by chemical mutagenesis in Caenorhabditis elegans. Nat. Genet. 17: 119–121

    Not in our library10.1038/ng0997-119

30. 30.

    Kamath R. S. Fraser A. G. Dong Y. Poulin G. Durbin R., 2003 Systematic functional analysis of the Caenorhabditis elegans genome using RNAi. Nature 421: 231–237

    Not in our library10.1038/nature01278

31. 31.

    Kaufman E. J. Miska E. A., 2010 The microRNAs of Caenorhabditis elegans. Semin. Cell Dev. Biol. 21: 728–737

    Not in our library10.1016/j.semcdb.2010.07.001

32. 32.

    Knight S. W. Bass B. L., 2001 A role for the RNase III enzyme DCR-1 in RNA interference and germ line development in Caenorhabditis elegans. Science 293: 2269–2271

    Not in our library10.1126/science.1062039

33. 33.

    Krause M. Wild M. Rosenzweig B. Hirsh D., 1989 Wild-type and mutant actin genes in Caenorhabditis elegans. J. Mol. Biol. 208: 381–392

    Not in our library10.1016/0022-2836(89)90503-2

34. 34.

    Li C. Kim K., 2008 Neuropeptides (September 25, 2008), WormBook, ed. The C. elegans Research Community WormBook, /10.1895/wormbook.1.142.1, http://www.wormbook.org

    Not in our library

35. 35.

    Li T. Huang S. Jiang W. Z. Wright D. Spalding M. H., 2011 TAL nucleases (TALNs): hybrid proteins composed of TAL effectors and FokI DNA-cleavage domain. Nucleic Acids Res. 39: 359–372

    Not in our library10.1093/nar/gkq704

36. 36.

    Liu L. X. Spoerke J. M. Mulligan E. L. Chen J. Reardon B., 1999 High-throughput isolation of Caenorhabditis elegans deletion mutants. Genome Res. 9: 859–867

    Not in our library10.1101/gr.9.9.859

37. 37.

    MacQueen A. J. Baggett J. J. Perumov N. Bauer R. A. Januszewski T., 2005 ACT-5 is an essential Caenorhabditis elegans actin required for intestinal microvilli formation. Mol. Biol. Cell 16: 3247–3259

    Not in our library10.1091/mbc.e04-12-1061

38. 38.

    Manning G., 2005 Genomic overview of protein kinases (December 13, 2005), WormBook, ed. The C. elegans Research Community Wormbook, /10.1895/wormbook.1.60.1, http://www.wormbook.org

    Not in our library10.1895/wormbook.1.60.1

39. 39.

    Mano I. Driscoll M., 1999 DEG/ENaC channels: a touchy superfamily that watches its salt. Bioessays 21: 568–578

    Not in our library10.1002/(sici)1521-1878(199907)21:7<568::aid-bies5>3.0.co;2-l

40. 40.

    Maydan J. S. Flibotte S. Edgley M. L. Lau J. Selzer R. R., 2007 Efficient high-resolution deletion discovery in Caenorhabditis elegans by array comparative genomic hybridization. Genome Res. 17: 337–347

    Not in our library10.1101/gr.5690307

Cited by papers we hold 5

• C. elegans survival assays to discern global and transcription-coupled nucleotide excision repair · 2021
  Not checked
• Neuronal TORC1 modulates longevity via AMPK and cell nonautonomous regulation of mitochondrial dynamics in C. elegans · 2019
  No notice
• Ordered arrangement of dendrites within a C. elegans sensory nerve bundle · 2018
  Not checked
• An Efficient Genome Editing Strategy To Generate Putative Null Mutants in Caenorhabditis elegans Using CRISPR/Cas9 · 2018
  No notice
• Targeted mutagenesis in a human-parasitic nematode · 2017
  No notice

Reads

A person counts once a day per paper. We keep no cookie and no address: a reader is a hash that changes every day. Any month under five reads shows no number.