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. 1978 Oct;75(10):4709–4713. doi: 10.1073/pnas.75.10.4709

DNA clones containing mouse immunoglobulin kappa chain genes isolated by in vitro packaging into phage lambda coats.

R Lenhard-Schuller, B Hohn, C Brack, M Hirama, S Tonegawa
PMCID: PMC336189  PMID: 105355

Abstract

Endonuclease EcoRI-digested DNAs from BALB/c mouse embryos and MOPC 321 (a kappa chain secretor) myeloma were fractionated by agarose gel electrophoresis, and the DNA fragments containing part or all of the MOPC 321 kappa chain structural gene sequences were visualized by the Southern gel blotting technique using as the hybridization probes pCRI plasmids containing all or part of the enzymatically synthesized cDNA transcripts of the MOPC 321 kappa chain mRNA. The clear differences observed in the hybridization patterns of the two DNAs are in agreement with our previously reported results obtained with endonuclease BamHI and confirms that the sequence arrangement of kappa chain genes is different in the embryo and myeloma cells. We have cloned most of the kappa-sequence-positive EcoRI DNA fragments in Charon 4A phage by using the highly efficient in vitro phage lambda DNA packaging method, and we have characterized the cloned mouse DNA sequences by agarose gel blotting and R-loop mapping in electron microscopy. These studies identified, among others, one EcoRI DNA fragment which contains both variable and constant immunoglobulin kappa-gene sequences and is present only in the myeloma DNA. The two sequences are separated by a 2.8-kbase intron. We tentatively conclude that the kappa gene sequences on this DNA fragment underwent somatic rearrangement.

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Selected References

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

  1. Benton W. D., Davis R. W. Screening lambdagt recombinant clones by hybridization to single plaques in situ. Science. 1977 Apr 8;196(4286):180–182. doi: 10.1126/science.322279. [DOI] [PubMed] [Google Scholar]
  2. Blattner F. R., Williams B. G., Blechl A. E., Denniston-Thompson K., Faber H. E., Furlong L., Grunwald D. J., Kiefer D. O., Moore D. D., Schumm J. W. Charon phages: safer derivatives of bacteriophage lambda for DNA cloning. Science. 1977 Apr 8;196(4286):161–169. doi: 10.1126/science.847462. [DOI] [PubMed] [Google Scholar]
  3. Brack C., Tonegawa S. Variable and constant parts of the immunoglobulin light chain gene of a mouse myeloma cell are 1250 nontranslated bases apart. Proc Natl Acad Sci U S A. 1977 Dec;74(12):5652–5656. doi: 10.1073/pnas.74.12.5652. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Cameron J. R., Panasenko S. M., Lehman I. R., Davis R. W. In vitro construction of bacteriophage lambda carrying segments of the Escherichia coli chromosome: selection of hybrids containing the gene for DNA ligase. Proc Natl Acad Sci U S A. 1975 Sep;72(9):3416–3420. doi: 10.1073/pnas.72.9.3416. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. Hozumi N., Brack C., Pirrotta V., Lenhard-Schuller R., Tonegawa S. Characterization of a mouse DNA clone containing an immunoglobulin variable region gene. Nucleic Acids Res. 1978 Jun;5(6):1779–1799. doi: 10.1093/nar/5.6.1779. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. Hozumi N., Tonegawa S. Evidence for somatic rearrangement of immunoglobulin genes coding for variable and constant regions. Proc Natl Acad Sci U S A. 1976 Oct;73(10):3628–3632. doi: 10.1073/pnas.73.10.3628. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. Leder P., Tiemeier D., Enquist L. EK2 derivatives of bacteriophage lambda useful in the cloning of DNA from higher organisms: the lambdagtWES system. Science. 1977 Apr 8;196(4286):175–177. doi: 10.1126/science.322278. [DOI] [PubMed] [Google Scholar]
  8. 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]
  9. 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]
  10. Sternberg N., Tiemeier D., Enquist L. In vitro packaging of a lambda Dam vector containing EcoRI DNA fragments of Escherichia coli and phage P1. Gene. 1977 May;1(3-4):255–280. doi: 10.1016/0378-1119(77)90049-x. [DOI] [PubMed] [Google Scholar]
  11. Tonegawa S., Brack C., Hozumi N., Matthyssens G., Schuller R. Dynamics of immunoglobulin genes. Immunol Rev. 1977;36:73–94. doi: 10.1111/j.1600-065x.1977.tb00383.x. [DOI] [PubMed] [Google Scholar]
  12. Tonegawa S., Brack C., Hozumi N., Schuller R. Cloning of an immunoglobulin variable region gene from mouse embryo. Proc Natl Acad Sci U S A. 1977 Aug;74(8):3518–3522. doi: 10.1073/pnas.74.8.3518. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. Tonegawa S., Hozumi N., Matthyssens G., Schuller R. Somatic changes in the content and context of immunoglobulin genes. Cold Spring Harb Symp Quant Biol. 1977;41(Pt 2):877–889. doi: 10.1101/sqb.1977.041.01.097. [DOI] [PubMed] [Google Scholar]
  14. Tonegawa S., Maxam A. M., Tizard R., Bernard O., Gilbert W. Sequence of a mouse germ-line gene for a variable region of an immunoglobulin light chain. Proc Natl Acad Sci U S A. 1978 Mar;75(3):1485–1489. doi: 10.1073/pnas.75.3.1485. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. Tonegawa S. Reiteration frequency of immunoglobulin light chain genes: further evidence for somatic generation of antibody diversity. Proc Natl Acad Sci U S A. 1976 Jan;73(1):203–207. doi: 10.1073/pnas.73.1.203. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Tonegawa S., Steinberg C., Dube S., Bernardini A. Evidence for somatic generation of antibody diversity. Proc Natl Acad Sci U S A. 1974 Oct;71(10):4027–4031. doi: 10.1073/pnas.71.10.4027. [DOI] [PMC free article] [PubMed] [Google Scholar]
  17. Weigert M., Riblet R. Genetic control of antibody variable regions. Cold Spring Harb Symp Quant Biol. 1977;41(Pt 2):837–846. doi: 10.1101/sqb.1977.041.01.093. [DOI] [PubMed] [Google Scholar]

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