Skip to main content
Nucleic Acids Research logoLink to Nucleic Acids Research
. 1980 Apr 25;8(8):1823–1837. doi: 10.1093/nar/8.8.1823

Isolation and characterization of genomic DNA coding for alpha 2 type I collagen.

G Vogeli, E V Avvedimento, M Sullivan, J V Maizel Jr, G Lozano, S L Adams, I Pastan, B de Crombrugghe
PMCID: PMC324038  PMID: 7433131

Abstract

We have isolated and characterized a segment of the chick alpha 2 collagen gene by screening a library of chick genomic fragments using as hybridization probe an alpha 2 collagen cDNA clone. Several clones were isolated and one of them, lambda gCOL 204, was used for further studies. The DNA of lambda gCOL 204 hybridizes to a unique species of mRNA the size of alpha 2 collagen mRNA. This mRNA can be translated into a unique polypeptide which comigrates in SDS-gel electrophoresis with pro-alpha 2 collagen. Electron microscopic analysis by R-loop technique indicates that lambda gCOL 204 contains 7Kb of the alpha 2 collagen gene. This 7 Kb piece constitutes the 3' end of the gene. The same clone also contains 9 Kb of DNA that is immediately adjacent to the 3' end of the alpha 2 collagen gene. The cloned segment of the alpha 2 collagen gene is interrupted by 8 intervening sequences of various lengths. The coding sequences for collagen in this clone add up to approximately 1,800 bp, which correspond to about 1/3 of alpha 2 collagen mRNA. DNA sequence analysis of a small coding segment of lambda g COL 204 reveals a characteristic collagen type sequence which encodes for an amino acid sequence identical to a sequence found in calf alpha 2 collagen. The sequence of this region of the protein has not yet been determined for the chick alpha 2 collagen.

Full text

PDF
1824

Images in this article

Selected References

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

  1. Adams S. L., Alwine J. C., de Crombrugghe B., Pastan I. Use of recombinant plasmids to characterize collagen RNAs in normal and transformed chick embryo fibroblasts. J Biol Chem. 1979 Jun 25;254(12):4935–4938. [PubMed] [Google Scholar]
  2. Adams S. L., Sobel M. E., Howard B. H., Olden K., Yamada K. M., de Crombrugghe B., Pastan I. Levels of translatable mRNAs for cell surface protein, collagen precursors, and two membrane proteins are altered in Rous sarcoma virus-transformed chick embryo fibroblasts. Proc Natl Acad Sci U S A. 1977 Aug;74(8):3399–3403. doi: 10.1073/pnas.74.8.3399. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Alwine J. C., Kemp D. J., Stark G. R. Method for detection of specific RNAs in agarose gels by transfer to diazobenzyloxymethyl-paper and hybridization with DNA probes. Proc Natl Acad Sci U S A. 1977 Dec;74(12):5350–5354. doi: 10.1073/pnas.74.12.5350. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. 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]
  5. Cochet M., Gannon F., Hen R., Maroteaux L., Perrin F., Chambon P. Organization and sequence studies of the 17-piece chicken conalbumin gene. Nature. 1979 Dec 6;282(5739):567–574. doi: 10.1038/282567a0. [DOI] [PubMed] [Google Scholar]
  6. 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]
  7. Dodgson J. B., Strommer J., Engel J. D. Isolation of the chicken beta-globin gene and a linked embryonic beta-like globin gene from a chicken DNA recombinant library. Cell. 1979 Aug;17(4):879–887. doi: 10.1016/0092-8674(79)90328-3. [DOI] [PubMed] [Google Scholar]
  8. Gilbert W. Why genes in pieces? Nature. 1978 Feb 9;271(5645):501–501. doi: 10.1038/271501a0. [DOI] [PubMed] [Google Scholar]
  9. Howard B. H., Adams S. L., Sobel M. E., Pastan I., de Crombrugghe B. Decreased levels of collagen mRNA in rous sarcoma virus-transformed chick embryo fibroblasts. J Biol Chem. 1978 Aug 25;253(16):5869–5874. [PubMed] [Google Scholar]
  10. Laskey R. A., Mills A. D. Quantitative film detection of 3H and 14C in polyacrylamide gels by fluorography. Eur J Biochem. 1975 Aug 15;56(2):335–341. doi: 10.1111/j.1432-1033.1975.tb02238.x. [DOI] [PubMed] [Google Scholar]
  11. Lehrach H., Frischauf A. M., Hanahan D., Wozney J., Fuller F., Boedtker H. Construction and characterization of pro alpha 1 collagen complementary deoxyribonucleic acid clones. Biochemistry. 1979 Jul 10;18(14):3146–3152. doi: 10.2196/47873. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. 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]
  13. Levinson W., Bhatnagar R. S., Liu T. Z. Loss of ability to synthesize collagen in fibroblasts transformed by rous sarcoma virus. J Natl Cancer Inst. 1975 Oct;55(4):807–810. doi: 10.1093/jnci/55.4.807. [DOI] [PubMed] [Google Scholar]
  14. Loening U. E. The determination of the molecular weight of ribonucleic acid by polyacrylamide-gel electrophresis. The effects of changes in conformation. Biochem J. 1969 Jun;113(1):131–138. doi: 10.1042/bj1130131. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. Maniatis T., Hardison R. C., Lacy E., Lauer J., O'Connell C., Quon D., Sim G. K., Efstratiadis A. The isolation of structural genes from libraries of eucaryotic DNA. Cell. 1978 Oct;15(2):687–701. doi: 10.1016/0092-8674(78)90036-3. [DOI] [PubMed] [Google Scholar]
  16. 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]
  17. Max E. E., Seidman J. G., Leder P. Sequences of five potential recombination sites encoded close to an immunoglobulin kappa constant region gene. Proc Natl Acad Sci U S A. 1979 Jul;76(7):3450–3454. doi: 10.1073/pnas.76.7.3450. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. Maxam A. M., Gilbert W. A new method for sequencing DNA. Proc Natl Acad Sci U S A. 1977 Feb;74(2):560–564. doi: 10.1073/pnas.74.2.560. [DOI] [PMC free article] [PubMed] [Google Scholar]
  19. Mayne R., Vail M. S., Miller E. J. Analysis of changes in collagen biosynthesis that occur when chick chondrocytes are grown in 5-bromo-2'-deoxyuridine. Proc Natl Acad Sci U S A. 1975 Nov;72(11):4511–4515. doi: 10.1073/pnas.72.11.4511. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. Pero J., Hannett N. M., Talkington C. Restriction cleavage map of SP01 DNA: general location of early, middle, and late genes. J Virol. 1979 Jul;31(1):156–171. doi: 10.1128/jvi.31.1.156-171.1979. [DOI] [PMC free article] [PubMed] [Google Scholar]
  21. Royal A., Garapin A., Cami B., Perrin F., Mandel J. L., LeMeur M., Brégégègre F., Gannon F., LePennec J. P., Chambon P. The ovalbumin gene region: common features in the organisation of three genes expressed in chicken oviduct under hormonal control. Nature. 1979 May 10;279(5709):125–132. doi: 10.1038/279125a0. [DOI] [PubMed] [Google Scholar]
  22. Sobel M. E., Yamamoto T., Adams S. L., DiLauro R., Avvedimento V. E., de Crombrugghe B., Pastan I. Construction of a recombinant bacterial plasmid containing a chick pro-alpha2 collagen gene sequence. Proc Natl Acad Sci U S A. 1978 Dec;75(12):5846–5850. doi: 10.1073/pnas.75.12.5846. [DOI] [PMC free article] [PubMed] [Google Scholar]
  23. 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]
  24. Tilghman S. M., Curtis P. J., Tiemeier D. C., Leder P., Weissmann C. The intervening sequence of a mouse beta-globin gene is transcribed within the 15S beta-globin mRNA precursor. Proc Natl Acad Sci U S A. 1978 Mar;75(3):1309–1313. doi: 10.1073/pnas.75.3.1309. [DOI] [PMC free article] [PubMed] [Google Scholar]
  25. Tsujimoto Y., Suzuki Y. Structural analysis of the fibroin gene at the 5' end and its surrounding regions. Cell. 1979 Feb;16(2):425–436. doi: 10.1016/0092-8674(79)90018-7. [DOI] [PubMed] [Google Scholar]
  26. Tucker P. W., Marcu K. B., Newell N., Richards J., Blattner F. R. Sequence of the cloned gene for the constant region of murine gamma 2b immunoglobulin heavy chain. Science. 1979 Dec 14;206(4424):1303–1306. doi: 10.1126/science.117549. [DOI] [PubMed] [Google Scholar]

Articles from Nucleic Acids Research are provided here courtesy of Oxford University Press

RESOURCES