Skip to main content
PMC home page
Proceedings of the National Academy of Sciences of the United States of America logoLink to Proceedings of the National Academy of Sciences of the United States of America
. 1982 Aug;79(16):5038–5041. doi: 10.1073/pnas.79.16.5038

Isolation of a genomic clone partially encoding human hypoxanthine phosphoribosyltransferase.

D J Jolly, A C Esty, H U Bernard, T Friedmann
PMCID: PMC346822  PMID: 6956912

Abstract

Mouse cells deficient in the enzyme hypoxanthine phosphoribosyltransferase (HPRT; EC 2.4.2.8) have been transfected with total human DNA, and cells producing human enzyme were isolated by growth in selective medium. DNA from several such cell lines has been used to generate secondary transfectants that make human HPRT. Blots of the DNA of these secondary cells have been hybridized with total human DNA probes or with cloned human Alu sequences, and one of several common bands has been cloned in pBR322. Colonies of transformed Escherichia coli containing human sequences were detected by their homology with human DNA, and subclones of resulting recombinant plasmids were prepared. Two subclones free of Alu sequences were found to contain human sequences that hybridized to human X chromosome DNA. One of these, pBR1.5, also hybridized to a single RNA band on gel blots of human and secondary transfectant cytoplasmic poly(A)+RNA but not to RNA from the parent mouse cell line. These results indicate that these clones represent human HPRT gene fragments. This has been confirmed by using pBR1.5 as a probe to isolate an authentic and expressible human HPRT cDNA clone from a library prepared by H. Okayama and P. Berg.

Full text

PDF
5038

Images in this article

5039Image
on p.5039
5040Image
on p.5040
5040Image
on p.5040

Selected References

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

  1. Aviv H., Leder P. Purification of biologically active globin messenger RNA by chromatography on oligothymidylic acid-cellulose. Proc Natl Acad Sci U S A. 1972 Jun;69(6):1408–1412. doi: 10.1073/pnas.69.6.1408. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Becker M. A., Seegmiller J. E. Recent advances in the identification of enzyme abnormalities underlying excessive purine synthesis in man. Arthritis Rheum. 1975 Nov-Dec;18(6 Suppl):687–694. doi: 10.1002/art.1780180708. [DOI] [PubMed] [Google Scholar]
  3. Becker M. A., Yen R. C., Itkin P., Goss S. J., Seegmiller J. E., Bakay B. Regional localization of the gene for human phosphoribosylpyrophosphate synthetase on the X chromosome. Science. 1979 Mar 9;203(4384):1016–1019. doi: 10.1126/science.218284. [DOI] [PubMed] [Google Scholar]
  4. Bell G. I., Pictet R., Rutter W. J. Analysis of the regions flanking the human insulin gene and sequence of an Alu family member. Nucleic Acids Res. 1980 Sep 25;8(18):4091–4109. doi: 10.1093/nar/8.18.4091. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. 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]
  6. 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]
  7. Caskey C. T., Kruh G. D. The HPRT locus. Cell. 1979 Jan;16(1):1–9. doi: 10.1016/0092-8674(79)90182-x. [DOI] [PubMed] [Google Scholar]
  8. Chang A. C., Cohen S. N. Construction and characterization of amplifiable multicopy DNA cloning vehicles derived from the P15A cryptic miniplasmid. J Bacteriol. 1978 Jun;134(3):1141–1156. doi: 10.1128/jb.134.3.1141-1156.1978. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Clewell D. B. Nature of Col E 1 plasmid replication in Escherichia coli in the presence of the chloramphenicol. J Bacteriol. 1972 May;110(2):667–676. doi: 10.1128/jb.110.2.667-676.1972. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. Cory S., Adams J. M. A very large repeating unit of mouse DNA containing the 18S, 28S and 5.8S rRNA genes. Cell. 1977 Aug;11(4):795–805. doi: 10.1016/0092-8674(77)90292-6. [DOI] [PubMed] [Google Scholar]
  11. Deininger P. L., Jolly D. J., Rubin C. M., Friedmann T., Schmid C. W. Base sequence studies of 300 nucleotide renatured repeated human DNA clones. J Mol Biol. 1981 Sep 5;151(1):17–33. doi: 10.1016/0022-2836(81)90219-9. [DOI] [PubMed] [Google Scholar]
  12. Deininger P. L., Schmid C. W. An electron microscope study of the DNA sequence organization of the human genome. J Mol Biol. 1976 Sep 25;106(3):773–790. doi: 10.1016/0022-2836(76)90264-3. [DOI] [PubMed] [Google Scholar]
  13. Denhardt D. T. A membrane-filter technique for the detection of complementary DNA. Biochem Biophys Res Commun. 1966 Jun 13;23(5):641–646. doi: 10.1016/0006-291x(66)90447-5. [DOI] [PubMed] [Google Scholar]
  14. Duncan C., Biro P. A., Choudary P. V., Elder J. T., Wang R. R., Forget B. G., de Riel J. K., Weissman S. M. RNA polymerase III transcriptional units are interspersed among human non-alpha-globin genes. Proc Natl Acad Sci U S A. 1979 Oct;76(10):5095–5099. doi: 10.1073/pnas.76.10.5095. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. Fritsch E. F., Lawn R. M., Maniatis T. Molecular cloning and characterization of the human beta-like globin gene cluster. Cell. 1980 Apr;19(4):959–972. doi: 10.1016/0092-8674(80)90087-2. [DOI] [PubMed] [Google Scholar]
  16. Fuhrman S. A., Deininger P. L., LaPorte P., Friedmann T., Geiduschek E. P. Analysis of transcription of the human Alu family ubiquitous repeating element by eukaryotic RNA polymerase III. Nucleic Acids Res. 1981 Dec 11;9(23):6439–6456. doi: 10.1093/nar/9.23.6439. [DOI] [PMC free article] [PubMed] [Google Scholar]
  17. Gergen J. P., Stern R. H., Wensink P. C. Filter replicas and permanent collections of recombinant DNA plasmids. Nucleic Acids Res. 1979 Dec 20;7(8):2115–2136. doi: 10.1093/nar/7.8.2115. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. Graf L. H., Jr, Urlaub G., Chasin L. A. Transformation of the gene for hypoxanthine phosphoribosyltransferase. Somatic Cell Genet. 1979 Nov;5(6):1031–1044. doi: 10.1007/BF01542658. [DOI] [PubMed] [Google Scholar]
  19. Graham D. E. The isolation of high molecular weight DNA from whole organisms or large tissue masses. Anal Biochem. 1978 Apr;85(2):609–613. doi: 10.1016/0003-2697(78)90262-2. [DOI] [PubMed] [Google Scholar]
  20. Gusella J. F., Keys C., VarsanyiBreiner A., Kao F. T., Jones C., Puck T. T., Housman D. Isolation and localization of DNA segments from specific human chromosomes. Proc Natl Acad Sci U S A. 1980 May;77(5):2829–2833. doi: 10.1073/pnas.77.5.2829. [DOI] [PMC free article] [PubMed] [Google Scholar]
  21. Holmes D. S., Quigley M. A rapid boiling method for the preparation of bacterial plasmids. Anal Biochem. 1981 Jun;114(1):193–197. doi: 10.1016/0003-2697(81)90473-5. [DOI] [PubMed] [Google Scholar]
  22. Johnson G. G., Eisenberg L. R., Migeon B. R. Human and mouse hypoxanthine-guanine phosphoribosyltransferase: dimers and tetramers. Science. 1979 Jan 12;203(4376):174–176. doi: 10.1126/science.569362. [DOI] [PubMed] [Google Scholar]
  23. Lawn R. M., Gross M., Houck C. M., Franke A. E., Gray P. V., Goeddel D. V. DNA sequence of a major human leukocyte interferon gene. Proc Natl Acad Sci U S A. 1981 Sep;78(9):5435–5439. doi: 10.1073/pnas.78.9.5435. [DOI] [PMC free article] [PubMed] [Google Scholar]
  24. Lehrach H., Frischauf A. M., Hanahan D., Wozney J., Fuller F., Crkvenjakov R., Boedtker H., Doty P. Construction and characterization of a 2.5-kilobase procollagen clone. Proc Natl Acad Sci U S A. 1978 Nov;75(11):5417–5421. doi: 10.1073/pnas.75.11.5417. [DOI] [PMC free article] [PubMed] [Google Scholar]
  25. Lester S. C., LeVan S. K., Steglich C., DeMars R. Expression of human genes for adenine phosphoribosyltransferase and hypoxanthine-guanine phosphoribosyltransferase after genetic transformation of mouse cells with purified human DNA. Somatic Cell Genet. 1980 Mar;6(2):241–259. doi: 10.1007/BF01538799. [DOI] [PubMed] [Google Scholar]
  26. Levis R., Penman S. Processing steps and methylation in the formation of the ribosomal RNA of cultured Drosophila cells. J Mol Biol. 1978 May 15;121(2):219–238. doi: 10.1016/s0022-2836(78)80006-0. [DOI] [PubMed] [Google Scholar]
  27. Littlefield J. W. The use of drug-resistant markers to study the hybridization of mouse fibroblasts. Exp Cell Res. 1966 Jan;41(1):190–196. doi: 10.1016/0014-4827(66)90558-1. [DOI] [PubMed] [Google Scholar]
  28. Maniatis T., Jeffrey A., Kleid D. G. Nucleotide sequence of the rightward operator of phage lambda. Proc Natl Acad Sci U S A. 1975 Mar;72(3):1184–1188. doi: 10.1073/pnas.72.3.1184. [DOI] [PMC free article] [PubMed] [Google Scholar]
  29. Murray M. J., Shilo B. Z., Shih C., Cowing D., Hsu H. W., Weinberg R. A. Three different human tumor cell lines contain different oncogenes. Cell. 1981 Aug;25(2):355–361. doi: 10.1016/0092-8674(81)90054-4. [DOI] [PubMed] [Google Scholar]
  30. Pellicer A., Wigler M., Axel R., Silverstein S. The transfer and stable integration of the HSV thymidine kinase gene into mouse cells. Cell. 1978 May;14(1):133–141. doi: 10.1016/0092-8674(78)90308-2. [DOI] [PubMed] [Google Scholar]
  31. Perucho M., Goldfarb M., Shimizu K., Lama C., Fogh J., Wigler M. Human-tumor-derived cell lines contain common and different transforming genes. Cell. 1981 Dec;27(3 Pt 2):467–476. doi: 10.1016/0092-8674(81)90388-3. [DOI] [PubMed] [Google Scholar]
  32. Peterson J. L., McBride O. W. Cotransfer of linked eukaryotic genes and efficient transfer of hypoxanthine phosphoribosyltransferase by DNA-mediated gene transfer. Proc Natl Acad Sci U S A. 1980 Mar;77(3):1583–1587. doi: 10.1073/pnas.77.3.1583. [DOI] [PMC free article] [PubMed] [Google Scholar]
  33. Philippsen P., Kramer R. A., Davis R. W. Cloning of the yeast ribosomal DNA repeat unit in SstI and HindIII lambda vectors using genetic and physical size selections. J Mol Biol. 1978 Aug 15;123(3):371–386. doi: 10.1016/0022-2836(78)90085-2. [DOI] [PubMed] [Google Scholar]
  34. Seegmiller J. E., Rosenbloom F. M., Kelley W. N. Enzyme defect associated with a sex-linked human neurological disorder and excessive purine synthesis. Science. 1967 Mar 31;155(3770):1682–1684. doi: 10.1126/science.155.3770.1682. [DOI] [PubMed] [Google Scholar]
  35. Smith A. J. The use of exonuclease III for preparing single stranded DNA for use as a template in the chain terminator sequencing method. Nucleic Acids Res. 1979 Mar;6(3):831–848. doi: 10.1093/nar/6.3.831. [DOI] [PMC free article] [PubMed] [Google Scholar]
  36. 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]
  37. Thomas P. S. Hybridization of denatured RNA and small DNA fragments transferred to nitrocellulose. Proc Natl Acad Sci U S A. 1980 Sep;77(9):5201–5205. doi: 10.1073/pnas.77.9.5201. [DOI] [PMC free article] [PubMed] [Google Scholar]
  38. Ullrich A., Shine J., Chirgwin J., Pictet R., Tischer E., Rutter W. J., Goodman H. M. Rat insulin genes: construction of plasmids containing the coding sequences. Science. 1977 Jun 17;196(4296):1313–1319. doi: 10.1126/science.325648. [DOI] [PubMed] [Google Scholar]
  39. Wahl G. M., Stern M., Stark G. R. Efficient transfer of large DNA fragments from agarose gels to diazobenzyloxymethyl-paper and rapid hybridization by using dextran sulfate. Proc Natl Acad Sci U S A. 1979 Aug;76(8):3683–3687. doi: 10.1073/pnas.76.8.3683. [DOI] [PMC free article] [PubMed] [Google Scholar]
  40. Wigler M., Pellicer A., Silverstein S., Axel R. Biochemical transfer of single-copy eucaryotic genes using total cellular DNA as donor. Cell. 1978 Jul;14(3):725–731. doi: 10.1016/0092-8674(78)90254-4. [DOI] [PubMed] [Google Scholar]

Articles from Proceedings of the National Academy of Sciences of the United States of America are provided here courtesy of National Academy of Sciences

RESOURCES