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
. 1980 May;77(5):2450–2454. doi: 10.1073/pnas.77.5.2450

Formation of an RNA primer for initiation of replication of ColE1 DNA by ribonuclease H.

T Itoh, J Tomizawa
PMCID: PMC349417  PMID: 6156450

Abstract

A plasmid that consists of an 812-base-pair segment containing the replication origin of plasmid ColE1 and of a 1240-base-pair segment containing a beta-lactamase gene has been constructed. The plasmid DNA has three principal sites where transcription is initiated in vitro. One is located in the ColE1 segment 555 nucleotides upstream from the origin. Most transcription from this site extends past the origin; some of the transcripts form hybrids spontaneously with the template at their 3' portions. Cleavage of these transcripts by RNase H generates 3' termini at the origin region. When DNA polymerase I is included in the reaction along with RNA polymerase and RNase H, dAMP or dCMP is added directly onto the cleaved RNA molecules, most of which retain the intact 5' terminus. The addition of a deoxyribonucleotide to the cleaved RNA can be regarded as the first step of ColE1 DNA synthesis. Once it has served as a primer, the RNA is eliminated from the product by RNase H.

Full text

PDF
2451

Images in this article

2451Image
on p.2451
2451Image
on p.2451
2452Image
on p.2452
2452Image
on p.2452
2453Image
on p.2453
2453Image
on p.2453

Selected References

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

  1. Backman K., Betlach M., Boyer H. W., Yanofsky S. Genetic and physical studies on the replication of ColE1-type plasmids. Cold Spring Harb Symp Quant Biol. 1979;43(Pt 1):69–76. doi: 10.1101/sqb.1979.043.01.012. [DOI] [PubMed] [Google Scholar]
  2. Berkower I., Leis J., Hurwitz J. Isolation and characterization of an endonuclease from Escherichia coli specific for ribonucleic acid in ribonucleic acid-deoxyribonucleic acid hybrid structures. J Biol Chem. 1973 Sep 10;248(17):5914–5921. [PubMed] [Google Scholar]
  3. Bird R. E., Tomizawa J. Ribonucleotide-deoxyribonucleotide linkages at the origin of DNA replication of colicin E1 plasmid. J Mol Biol. 1978 Mar 25;120(1):137–143. doi: 10.1016/0022-2836(78)90300-5. [DOI] [PubMed] [Google Scholar]
  4. Bolivar F., Betlach M. C., Heyneker H. L., Shine J., Rodriguez R. L., Boyer H. W. Origin of replication of pBR345 plasmid DNA. Proc Natl Acad Sci U S A. 1977 Dec;74(12):5265–5269. doi: 10.1073/pnas.74.12.5265. [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. Brownlee G. G., Sanger F., Barrell B. G. The sequence of 5 s ribosomal ribonucleic acid. J Mol Biol. 1968 Jun 28;34(3):379–412. doi: 10.1016/0022-2836(68)90168-x. [DOI] [PubMed] [Google Scholar]
  7. Burgess R. R., Jendrisak J. J. A procedure for the rapid, large-scall purification of Escherichia coli DNA-dependent RNA polymerase involving Polymin P precipitation and DNA-cellulose chromatography. Biochemistry. 1975 Oct 21;14(21):4634–4638. doi: 10.1021/bi00692a011. [DOI] [PubMed] [Google Scholar]
  8. Carbon P., Ehresmann C., Ehresmann B., Ebel J. P. The complete nucleotide sequence of the ribosomal 16-S RNA from Excherichia coli. Experimental details and cistron heterogeneities. Eur J Biochem. 1979 Oct 15;100(2):399–410. doi: 10.1111/j.1432-1033.1979.tb04183.x. [DOI] [PubMed] [Google Scholar]
  9. England T. E., Uhlenbeck O. C. Enzymatic oligoribonucleotide synthesis with T4 RNA ligase. Biochemistry. 1978 May 30;17(11):2069–2076. doi: 10.1021/bi00604a008. [DOI] [PubMed] [Google Scholar]
  10. Gottesman M. E., Yarmolinsky M. B. Integration-negative mutants of bacteriophage lambda. J Mol Biol. 1968 Feb 14;31(3):487–505. doi: 10.1016/0022-2836(68)90423-3. [DOI] [PubMed] [Google Scholar]
  11. Horii T., Ogawa T., Ogawa H. Organization of the recA gene of Escherichia coli. Proc Natl Acad Sci U S A. 1980 Jan;77(1):313–317. doi: 10.1073/pnas.77.1.313. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Itoh T., Tomizawa J. Initiation of replication of plasmid ColE1 DNA by RNA polymerase, ribonuclease H, and DNA polymerase I. Cold Spring Harb Symp Quant Biol. 1979;43(Pt 1):409–417. doi: 10.1101/sqb.1979.043.01.047. [DOI] [PubMed] [Google Scholar]
  13. Jovin T. M., Englund P. T., Bertsch L. L. Enzymatic synthesis of deoxyribonucleic acid. XXVI. Physical and chemical studies of a homogeneous deoxyribonucleic acid polymerase. J Biol Chem. 1969 Jun 10;244(11):2996–3008. [PubMed] [Google Scholar]
  14. Kikuchi Y., Hishinuma F., Sakaguchi K. Addition of mononucleotides to oligoribonucleotide acceptors with T4 RNA ligase. Proc Natl Acad Sci U S A. 1978 Mar;75(3):1270–1273. doi: 10.1073/pnas.75.3.1270. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. Krupp G., Gross H. J. Rapid RNA sequencing: nucleases from Staphylococcus aureus and Neurospora crassa discriminate between uridine and cytidine. Nucleic Acids Res. 1979 Aug 10;6(11):3481–3490. doi: 10.1093/nar/6.11.3481. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Laemmli U. K. Cleavage of structural proteins during the assembly of the head of bacteriophage T4. Nature. 1970 Aug 15;227(5259):680–685. doi: 10.1038/227680a0. [DOI] [PubMed] [Google Scholar]
  17. Lebowitz P., Weissman S. M., Radding C. M. Nucleotide sequence of a ribonucleic acid transcribed in vitro from lambda phage deoxyribonucleic acid. J Biol Chem. 1971 Aug 25;246(16):5120–5139. [PubMed] [Google Scholar]
  18. Morita M., Oka A. The structure of a transcriptional unit on colicin E1 plasmid. Eur J Biochem. 1979 Jul;97(2):435–443. doi: 10.1111/j.1432-1033.1979.tb13131.x. [DOI] [PubMed] [Google Scholar]
  19. Ohmori H., Tomizawa J. Nucleotide sequence of the region required for maintenance of colicin E1 plasmid. Mol Gen Genet. 1979 Oct 3;176(2):161–170. doi: 10.1007/BF00273210. [DOI] [PubMed] [Google Scholar]
  20. Oka A., Nomura N., Morita M., Sugisaki H., Sugimoto K., Takanami M. Nucleotide sequence of small ColE1 derivatives: structure of the regions essential for autonomous replication and colicin E1 immunity. Mol Gen Genet. 1979 May 4;172(2):151–159. doi: 10.1007/BF00268276. [DOI] [PubMed] [Google Scholar]
  21. Sakakibara Y., Tomizawa J. I. Replication of colicin E1 plasmid DNA in cell extracts. Proc Natl Acad Sci U S A. 1974 Mar;71(3):802–806. doi: 10.1073/pnas.71.3.802. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. Schott H., Rudloff E., Schmidt P., Roychoudhury R., Kössel H. A dihydroxyboryl-substituted methacrylic polymer for the column chromatographic separation of mononucleotides, oligonucleotides, and transfer ribonucleic acid. Biochemistry. 1973 Feb 27;12(5):932–938. doi: 10.1021/bi00729a022. [DOI] [PubMed] [Google Scholar]
  23. Schwarz E., Scherer G., Hobom G., Kössel H. Nucleotide sequence of cro, cII and part of the O gene in phage lambda DNA. Nature. 1978 Mar 30;272(5652):410–414. doi: 10.1038/272410a0. [DOI] [PubMed] [Google Scholar]
  24. Simoncsits A., Brownlee G. G., Brown R. S., Rubin J. R., Guilley H. New rapid gel sequencing method for RNA. Nature. 1977 Oct 27;269(5631):833–836. doi: 10.1038/269833a0. [DOI] [PubMed] [Google Scholar]
  25. Staudenbauer W. L. Structure and replication of the colicin E1 plasmid. Curr Top Microbiol Immunol. 1978;83:93–156. doi: 10.1007/978-3-642-67087-9_3. [DOI] [PubMed] [Google Scholar]
  26. Sutcliffe J. G. Complete nucleotide sequence of the Escherichia coli plasmid pBR322. Cold Spring Harb Symp Quant Biol. 1979;43(Pt 1):77–90. doi: 10.1101/sqb.1979.043.01.013. [DOI] [PubMed] [Google Scholar]
  27. Tomizawa J. I., Ohmori H., Bird R. E. Origin of replication of colicin E1 plasmid DNA. Proc Natl Acad Sci U S A. 1977 May;74(5):1865–1869. doi: 10.1073/pnas.74.5.1865. [DOI] [PMC free article] [PubMed] [Google Scholar]
  28. Tomizawa J. I., Sakakibara Y., Kakefuda T. Replication of colicin E1 plasmid DNA added to cell extracts. Proc Natl Acad Sci U S A. 1975 Mar;72(3):1050–1054. doi: 10.1073/pnas.72.3.1050. [DOI] [PMC free article] [PubMed] [Google Scholar]
  29. Tomizawa J., Selzer G. Initiation of DNA synthesis in Escherichia coli. Annu Rev Biochem. 1979;48:999–1034. doi: 10.1146/annurev.bi.48.070179.005031. [DOI] [PubMed] [Google Scholar]
  30. Vicuna R., Hurwitz J., Wallace S., Girard M. Selective inhibition of in vitro DNA synthesis dependent on phiX174 compared with fd DNA. I. Protein requirements for selective inhibition. J Biol Chem. 1977 Apr 25;252(8):2524–2533. [PubMed] [Google Scholar]
  31. Vollenweider H. J., Stettler U., Kübler O., Koller T., Weber H. Refined molecular weights for phage, viral and ribosomal RNA. Gene. 1978 Jul;3(4):353–357. doi: 10.1016/0378-1119(78)90044-6. [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