All Title Author
Keywords Abstract

PLOS ONE  2008 

High-Throughput Isolation and Mapping of C. elegans Mutants Susceptible to Pathogen Infection

DOI: 10.1371/journal.pone.0002882

Full-Text   Cite this paper   Add to My Lib


We present a novel strategy that uses high-throughput methods of isolating and mapping C. elegans mutants susceptible to pathogen infection. We show that C. elegans mutants that exhibit an enhanced pathogen accumulation (epa) phenotype can be rapidly identified and isolated using a sorting system that allows automation of the analysis, sorting, and dispensing of C. elegans by measuring fluorescent bacteria inside the animals. Furthermore, we validate the use of Amplifluor? as a new single nucleotide polymorphism (SNP) mapping technique in C. elegans. We show that a set of 9 SNPs allows the linkage of C. elegans mutants to a 5–8 megabase sub-chromosomal region.


[1]  Mylonakis E, Aballay A (2005) Worms and flies as genetically tractable animal models to study host-pathogen interactions. Infect Immun 73: 3833–3841.
[2]  Kim DH, Ausubel FM (2005) Evolutionary perspectives on innate immunity from the study of Caenorhabditis elegans. Curr Opin Immunol 17: 4–10.
[3]  Sifri CD, Begun J, Ausubel FM (2005) The worm has turned - microbial virulence modeled in Caenorhabditis elegans. Trends Microbiol 13: 119–127.
[4]  Gravato-Nobre MJ, Hodgkin J (2005) Caenorhabditis elegans as a model for innate immunity to pathogens. Cell Microbiol 7: 741–751.
[5]  Kurz CL, Ewbank JJ (2003) Caenorhabditis elegans: an emerging genetic model for the study of innate immunity. Nat Rev Genet 4: 380–390.
[6]  Kim DH, Feinbaum R, Alloing G, Emerson FE, Garsin DA, et al. (2002) A conserved p38 MAP kinase pathway in Caenorhabditis elegans innate immunity. Science 297: 623–626.
[7]  Aballay A, Yorgey P, Ausubel FM (2000) Salmonella typhimurium proliferates and establishes a persistent infection in the intestine of Caenorhabditis elegans. Curr Biol 10: 1539–1542.
[8]  Wicks SR, Yeh RT, Gish WR, Waterston RH, Plasterk RH (2001) Rapid gene mapping in Caenorhabditis elegans using a high density polymorphism map. Nat Genet 28: 160–164.
[9]  Davis MW, Hammarlund M, Harrach T, Hullett P, Olsen S, et al. (2005) Rapid single nucleotide polymorphism mapping in C. elegans. BMC Genomics 6: 118.
[10]  Zipperlen P, Nairz K, Rimann I, Basler K, Hafen E, et al. (2005) A universal method for automated gene mapping. Genome Biol 6: R19.
[11]  Nairz K, Zipperlen P, Schneider M (2007) FLP-Mapping: a universal, cost-effective, and automatable method for gene mapping. Methods Mol Biol 396: 419–432.
[12]  Swan KA, Curtis DE, McKusick KB, Voinov AV, Mapa FA, et al. (2002) High-throughput gene mapping in Caenorhabditis elegans. Genome Res 12: 1100–1105.
[13]  Myakishev MV, Khripin Y, Hu S, Hamer DH (2001) High-throughput SNP genotyping by allele-specific PCR with universal energy-transfer-labeled primers. Genome Res 11: 163–169.
[14]  Petkov PM, Ding Y, Cassell MA, Zhang W, Wagner G, et al. (2004) An efficient SNP system for mouse genome scanning and elucidating strain relationships. Genome Res 14: 1806–1811.
[15]  Garsin DA, Sifri CD, Mylonakis E, Qin X, Singh KV, et al. (2001) A simple model host for identifying Gram-positive virulence factors. Proc Natl Acad Sci U S A 98: 10892–10897.
[16]  Kurz CL, Chauvet S, Andres E, Aurouze M, Vallet I, et al. (2003) Virulence factors of the human opportunistic pathogen Serratia marcescens identified by in vivo screening. Embo J 22: 1451–1460.
[17]  Styer KL, Hopkins GW, Bartra SS, Plano GV, Frothingham R, et al. (2005) Yersinia pestis kills Caenorhabditis elegans by a biofilm-independent process that involves novel virulence factors. EMBO Rep 6: 992–997.
[18]  Tenor JL, Aballay A (2008) A conserved Toll-like receptor is required for Caenorhabditis elegans innate immunity. EMBO Rep 9: 103–109.
[19]  Aballay A, Drenkard E, Hilbun LR, Ausubel FM (2003) Caenorhabditis elegans innate immune response triggered by Salmonella enterica requires intact LPS and is mediated by a MAPK signaling pathway. Curr Biol 13: 47–52.
[20]  Alegado RA, Tan MW (2008) Resistance to antimicrobial peptides contributes to persistence of Salmonella typhimurium in the C. elegans intestine. Cell Microbiol.
[21]  Davies EK, Peters AD, Keightley PD (1999) High frequency of cryptic deleterious mutations in Caenorhabditis elegans. Science 285: 1748–1751.
[22]  Anderson P (1995) Mutagenesis. Methods Cell Biol 48: 31–58.
[23]  Brenner S (1974) The genetics of Caenorhabditis elegans. Genetics 77: 71–94.
[24]  Vazquez-Torres A, Jones-Carson J, Baumler AJ, Falkow S, Valdivia R, et al. (1999) Extraintestinal dissemination of Salmonella by CD18-expressing phagocytes. Nature 401: 804–808.
[25]  Wray C, Sojka WJ (1978) Experimental Salmonella typhimurium infection in calves. Res Vet Sci 25: 139–143.
[26]  Tan MW, Mahajan-Miklos S, Ausubel FM (1999) Killing of Caenorhabditis elegans by Pseudomonas aeruginosa used to model mammalian bacterial pathogenesis. Proc Natl Acad Sci U S A 96: 715–720.
[27]  Murray BE, Singh KV, Ross RP, Heath JD, Dunny GM, et al. (1993) Generation of restriction map of Enterococcus faecalis OG1 and investigation of growth requirements and regions encoding biosynthetic function. J Bacteriol 175: 5216–5223.
[28]  Meneely PM, Farago AF, Kauffman TM (2002) Crossover distribution and high interference for both the X chromosome and an autosome during oogenesis and spermatogenesis in Caenorhabditis elegans. Genetics 162: 1169–1177.
[29]  Bengra C, Mifflin TE, Khripin Y, Manunta P, Williams SM, et al. (2002) Genotyping of essential hypertension single-nucleotide polymorphisms by a homogeneous PCR method with universal energy transfer primers. Clin Chem 48: 2131–2140.


comments powered by Disqus

Contact Us


微信:OALib Journal