Skip to main content
Molecular and Cellular Biology logoLink to Molecular and Cellular Biology
. 1985 Dec;5(12):3484–3496. doi: 10.1128/mcb.5.12.3484

Extrachromosomal DNA transformation of Caenorhabditis elegans.

D T Stinchcomb, J E Shaw, S H Carr, D Hirsh
PMCID: PMC369179  PMID: 3837845

Abstract

DNA was introduced into the germ line of the nematode Caenorhabditis elegans by microinjection. Approximately 10% of the injected worms gave rise to transformed progeny. Upon injection, supercoiled molecules formed a high-molecular-weight array predominantly composed of tandem repeats of the injected sequence. Injected linear molecules formed both tandem and inverted repeats as if they had ligated to each other. No worm DNA sequences were required in the injected plasmid for the formation of these high-molecular-weight arrays. Surprisingly, these high-molecular-weight arrays were extrachromosomal and heritable. On average 50% of the progeny of a transformed hermaphrodite still carried the exogenous sequences. In situ hybridization experiments demonstrated that approximately half of the transformed animals carried foreign DNA in all of their cells; the remainder were mosaic animals in which some cells contained the exogenous sequences while others carried no detectable foreign DNA. The presence of mosaic and nonmosaic nematodes in transformed populations may permit detailed analysis of the expression and function of C. elegans genes.

Full text

PDF
3484

Images in this article

Selected References

These references are in PubMed. This may not be the complete list of references from this article.

  1. Albertson D. G. Localization of the ribosomal genes in Caenorhabditis elegans chromosomes by in situ hybridization using biotin-labeled probes. EMBO J. 1984 Jun;3(6):1227–1234. doi: 10.1002/j.1460-2075.1984.tb01957.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Albertson D. G., Thomson J. N. The kinetochores of Caenorhabditis elegans. Chromosoma. 1982;86(3):409–428. doi: 10.1007/BF00292267. [DOI] [PubMed] [Google Scholar]
  3. Bishop J. M. Cellular oncogenes and retroviruses. Annu Rev Biochem. 1983;52:301–354. doi: 10.1146/annurev.bi.52.070183.001505. [DOI] [PubMed] [Google Scholar]
  4. Bolivar F., Rodriguez R. L., Greene P. J., Betlach M. C., Heyneker H. L., Boyer H. W., Crosa J. H., Falkow S. Construction and characterization of new cloning vehicles. II. A multipurpose cloning system. Gene. 1977;2(2):95–113. [PubMed] [Google Scholar]
  5. Brenner S. The genetics of Caenorhabditis elegans. Genetics. 1974 May;77(1):71–94. doi: 10.1093/genetics/77.1.71. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. Brinster R. L., Chen H. Y., Trumbauer M., Senear A. W., Warren R., Palmiter R. D. Somatic expression of herpes thymidine kinase in mice following injection of a fusion gene into eggs. Cell. 1981 Nov;27(1 Pt 2):223–231. doi: 10.1016/0092-8674(81)90376-7. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. Brinster R. L., Ritchie K. A., Hammer R. E., O'Brien R. L., Arp B., Storb U. Expression of a microinjected immunoglobulin gene in the spleen of transgenic mice. Nature. 1983 Nov 24;306(5941):332–336. doi: 10.1038/306332a0. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Capecchi M. R. High efficiency transformation by direct microinjection of DNA into cultured mammalian cells. Cell. 1980 Nov;22(2 Pt 2):479–488. doi: 10.1016/0092-8674(80)90358-x. [DOI] [PubMed] [Google Scholar]
  9. Clarke L., Carbon J. Genomic substitutions of centromeres in Saccharomyces cerevisiae. Nature. 1983 Sep 1;305(5929):23–28. doi: 10.1038/305023a0. [DOI] [PubMed] [Google Scholar]
  10. Costantini F., Lacy E. Introduction of a rabbit beta-globin gene into the mouse germ line. Nature. 1981 Nov 5;294(5836):92–94. doi: 10.1038/294092a0. [DOI] [PubMed] [Google Scholar]
  11. Emmons S. W., Klass M. R., Hirsh D. Analysis of the constancy of DNA sequences during development and evolution of the nematode Caenorhabditis elegans. Proc Natl Acad Sci U S A. 1979 Mar;76(3):1333–1337. doi: 10.1073/pnas.76.3.1333. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Emmons S. W., Yesner L., Ruan K. S., Katzenberg D. Evidence for a transposon in Caenorhabditis elegans. Cell. 1983 Jan;32(1):55–65. doi: 10.1016/0092-8674(83)90496-8. [DOI] [PubMed] [Google Scholar]
  13. Etkin L. D., Roberts M. Transmission of integrated sea urchin histone genes by nuclear transplantation in Xenopus laevis. Science. 1983 Jul 1;221(4605):67–69. doi: 10.1126/science.6857265. [DOI] [PubMed] [Google Scholar]
  14. Files J. G., Carr S., Hirsh D. Actin gene family of Caenorhabditis elegans. J Mol Biol. 1983 Mar 5;164(3):355–375. doi: 10.1016/0022-2836(83)90056-6. [DOI] [PubMed] [Google Scholar]
  15. Goldberg D. A., Posakony J. W., Maniatis T. Correct developmental expression of a cloned alcohol dehydrogenase gene transduced into the Drosophila germ line. Cell. 1983 Aug;34(1):59–73. doi: 10.1016/0092-8674(83)90136-8. [DOI] [PubMed] [Google Scholar]
  16. Gordon J. W., Scangos G. A., Plotkin D. J., Barbosa J. A., Ruddle F. H. Genetic transformation of mouse embryos by microinjection of purified DNA. Proc Natl Acad Sci U S A. 1980 Dec;77(12):7380–7384. doi: 10.1073/pnas.77.12.7380. [DOI] [PMC free article] [PubMed] [Google Scholar]
  17. Grosschedl R., Weaver D., Baltimore D., Costantini F. Introduction of a mu immunoglobulin gene into the mouse germ line: specific expression in lymphoid cells and synthesis of functional antibody. Cell. 1984 Oct;38(3):647–658. doi: 10.1016/0092-8674(84)90259-9. [DOI] [PubMed] [Google Scholar]
  18. Herman R. K. Analysis of genetic mosaics of the nematode Caneorhabditis elegans. Genetics. 1984 Sep;108(1):165–180. doi: 10.1093/genetics/108.1.165. [DOI] [PMC free article] [PubMed] [Google Scholar]
  19. Herman R. K., Madl J. E., Kari C. K. Duplications in Caenorhabditis elegans. Genetics. 1979 Jun;92(2):419–435. doi: 10.1093/genetics/92.2.419. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. Hirsh D., Oppenheim D., Klass M. Development of the reproductive system of Caenorhabditis elegans. Dev Biol. 1976 Mar;49(1):200–219. doi: 10.1016/0012-1606(76)90267-0. [DOI] [PubMed] [Google Scholar]
  21. Karn J., Brenner S., Barnett L. Protein structural domains in the Caenorhabditis elegans unc-54 myosin heavy chain gene are not separated by introns. Proc Natl Acad Sci U S A. 1983 Jul;80(14):4253–4257. doi: 10.1073/pnas.80.14.4253. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. Kimble J., Hirsh D. The postembryonic cell lineages of the hermaphrodite and male gonads in Caenorhabditis elegans. Dev Biol. 1979 Jun;70(2):396–417. doi: 10.1016/0012-1606(79)90035-6. [DOI] [PubMed] [Google Scholar]
  23. Kimble J., Hodgkin J., Smith T., Smith J. Suppression of an amber mutation by microinjection of suppressor tRNA in C. elegans. Nature. 1982 Sep 30;299(5882):456–458. doi: 10.1038/299456a0. [DOI] [PubMed] [Google Scholar]
  24. Liao L. W., Rosenzweig B., Hirsh D. Analysis of a transposable element in Caenorhabditis elegans. Proc Natl Acad Sci U S A. 1983 Jun;80(12):3585–3589. doi: 10.1073/pnas.80.12.3585. [DOI] [PMC free article] [PubMed] [Google Scholar]
  25. MacLeod A. R., Karn J., Brenner S. Molecular analysis of the unc-54 myosin heavy-chain gene of Caenorhabditis elegans. Nature. 1981 Jun 4;291(5814):386–390. doi: 10.1038/291386a0. [DOI] [PubMed] [Google Scholar]
  26. Mandel M., Higa A. Calcium-dependent bacteriophage DNA infection. J Mol Biol. 1970 Oct 14;53(1):159–162. doi: 10.1016/0022-2836(70)90051-3. [DOI] [PubMed] [Google Scholar]
  27. McDonell M. W., Simon M. N., Studier F. W. Analysis of restriction fragments of T7 DNA and determination of molecular weights by electrophoresis in neutral and alkaline gels. J Mol Biol. 1977 Feb 15;110(1):119–146. doi: 10.1016/s0022-2836(77)80102-2. [DOI] [PubMed] [Google Scholar]
  28. McKnight G. S., Hammer R. E., Kuenzel E. A., Brinster R. L. Expression of the chicken transferrin gene in transgenic mice. Cell. 1983 Sep;34(2):335–341. doi: 10.1016/0092-8674(83)90368-9. [DOI] [PubMed] [Google Scholar]
  29. NIGON V., BRUN J. L'évolution des structures nucléaires dans l'ovogenèse de Caenorhabditis elegans Maupas 1900. Chromosoma. 1955;7(2-3):129–169. doi: 10.1007/BF00329722. [DOI] [PubMed] [Google Scholar]
  30. Nasmyth K. A. Molecular genetics of yeast mating type. Annu Rev Genet. 1982;16:439–500. doi: 10.1146/annurev.ge.16.120182.002255. [DOI] [PubMed] [Google Scholar]
  31. Orr-Weaver T. L., Szostak J. W., Rothstein R. J. Yeast transformation: a model system for the study of recombination. Proc Natl Acad Sci U S A. 1981 Oct;78(10):6354–6358. doi: 10.1073/pnas.78.10.6354. [DOI] [PMC free article] [PubMed] [Google Scholar]
  32. Pardue M. L., Gall J. G. Nucleic acid hybridization to the DNA of cytological preparations. Methods Cell Biol. 1975;10:1–16. doi: 10.1016/s0091-679x(08)60727-x. [DOI] [PubMed] [Google Scholar]
  33. Rigby P. W., Dieckmann M., Rhodes C., Berg P. Labeling deoxyribonucleic acid to high specific activity in vitro by nick translation with DNA polymerase I. J Mol Biol. 1977 Jun 15;113(1):237–251. doi: 10.1016/0022-2836(77)90052-3. [DOI] [PubMed] [Google Scholar]
  34. Rubin G. M., Spradling A. C. Genetic transformation of Drosophila with transposable element vectors. Science. 1982 Oct 22;218(4570):348–353. doi: 10.1126/science.6289436. [DOI] [PubMed] [Google Scholar]
  35. Sakaguchi J., Yamamoto M. Cloned ural locus of Schizosaccharomyces pombe propagates autonomously in this yeast assuming a polymeric form. Proc Natl Acad Sci U S A. 1982 Dec;79(24):7819–7823. doi: 10.1073/pnas.79.24.7819. [DOI] [PMC free article] [PubMed] [Google Scholar]
  36. Scangos G., Ruddle F. H. Mechanisms and applications of DNA-mediated gene transfer in mammalian cells - a review. Gene. 1981 Jun-Jul;14(1-2):1–10. doi: 10.1016/0378-1119(81)90143-8. [DOI] [PubMed] [Google Scholar]
  37. Scholnick S. B., Morgan B. A., Hirsh J. The cloned dopa decarboxylase gene is developmentally regulated when reintegrated into the Drosophila genome. Cell. 1983 Aug;34(1):37–45. doi: 10.1016/0092-8674(83)90134-4. [DOI] [PubMed] [Google Scholar]
  38. Southern E. M. Detection of specific sequences among DNA fragments separated by gel electrophoresis. J Mol Biol. 1975 Nov 5;98(3):503–517. doi: 10.1016/s0022-2836(75)80083-0. [DOI] [PubMed] [Google Scholar]
  39. Southern P. J., Berg P. Transformation of mammalian cells to antibiotic resistance with a bacterial gene under control of the SV40 early region promoter. J Mol Appl Genet. 1982;1(4):327–341. [PubMed] [Google Scholar]
  40. Spradling A. C., Rubin G. M. The effect of chromosomal position on the expression of the Drosophila xanthine dehydrogenase gene. Cell. 1983 Aug;34(1):47–57. doi: 10.1016/0092-8674(83)90135-6. [DOI] [PubMed] [Google Scholar]
  41. Stinchcomb D. T., Mann C., Davis R. W. Centromeric DNA from Saccharomyces cerevisiae. J Mol Biol. 1982 Jun 25;158(2):157–190. doi: 10.1016/0022-2836(82)90427-2. [DOI] [PubMed] [Google Scholar]
  42. Sulston J. E., Horvitz H. R. Post-embryonic cell lineages of the nematode, Caenorhabditis elegans. Dev Biol. 1977 Mar;56(1):110–156. doi: 10.1016/0012-1606(77)90158-0. [DOI] [PubMed] [Google Scholar]
  43. Sulston J. E. Post-embryonic development in the ventral cord of Caenorhabditis elegans. Philos Trans R Soc Lond B Biol Sci. 1976 Aug 10;275(938):287–297. doi: 10.1098/rstb.1976.0084. [DOI] [PubMed] [Google Scholar]
  44. Swift G. H., Hammer R. E., MacDonald R. J., Brinster R. L. Tissue-specific expression of the rat pancreatic elastase I gene in transgenic mice. Cell. 1984 Oct;38(3):639–646. doi: 10.1016/0092-8674(84)90258-7. [DOI] [PubMed] [Google Scholar]
  45. Szostak J. W., Blackburn E. H. Cloning yeast telomeres on linear plasmid vectors. Cell. 1982 May;29(1):245–255. doi: 10.1016/0092-8674(82)90109-x. [DOI] [PubMed] [Google Scholar]
  46. Vieira J., Messing J. The pUC plasmids, an M13mp7-derived system for insertion mutagenesis and sequencing with synthetic universal primers. Gene. 1982 Oct;19(3):259–268. doi: 10.1016/0378-1119(82)90015-4. [DOI] [PubMed] [Google Scholar]
  47. Wagner E. F., Stewart T. A., Mintz B. The human beta-globin gene and a functional viral thymidine kinase gene in developing mice. Proc Natl Acad Sci U S A. 1981 Aug;78(8):5016–5020. doi: 10.1073/pnas.78.8.5016. [DOI] [PMC free article] [PubMed] [Google Scholar]

Articles from Molecular and Cellular Biology are provided here courtesy of Taylor & Francis

RESOURCES