Abstract
The nucleoprotein filament formed by the RecA protein of Escherichia coli on single-stranded DNA catalyzes the hybridization of RNA transcripts with single-stranded DNA sequences at 37 degrees C, in vitro. RecA protein rapidly promotes hybridization, even when noncomplementary RNA is in a millionfold nucleotide excess over hybridizing RNA, and in a thousandfold nucleotide excess over hybridizing single-stranded DNA. Heterologous double-stranded DNA and RecA-coated noncomplementary single-stranded DNA are also poor competitors of RNA transcripts produced in vitro. Since large excesses of noncomplementary RNA fail to inhibit sharply the hybridization reaction by RecA protein under mild, non-degradative conditions, the reaction may be useful in the identification and isolation of transcripts produced in vivo.
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- Birnstiel M. L., Sells B. H., Purdom I. F. Kinetic complexity of RNA molecules. J Mol Biol. 1972 Jan 14;63(1):21–39. doi: 10.1016/0022-2836(72)90519-0. [DOI] [PubMed] [Google Scholar]
- Bryant F. R., Taylor A. R., Lehman I. R. Interaction of the recA protein of Escherichia coli with single-stranded DNA. J Biol Chem. 1985 Jan 25;260(2):1196–1202. [PubMed] [Google Scholar]
- Cunningham R. P., DasGupta C., Shibata T., Radding C. M. Homologous pairing in genetic recombination: recA protein makes joint molecules of gapped circular DNA and closed circular DNA. Cell. 1980 May;20(1):223–235. doi: 10.1016/0092-8674(80)90250-0. [DOI] [PubMed] [Google Scholar]
- Gonda D. K., Radding C. M. The mechanism of the search for homology promoted by recA protein. Facilitated diffusion within nucleoprotein networks. J Biol Chem. 1986 Oct 5;261(28):13087–13096. [PubMed] [Google Scholar]
- Honigberg S. M., Gonda D. K., Flory J., Radding C. M. The pairing activity of stable nucleoprotein filaments made from recA protein, single-stranded DNA, and adenosine 5'-(gamma-thio)triphosphate. J Biol Chem. 1985 Sep 25;260(21):11845–11851. [PubMed] [Google Scholar]
- Honigberg S. M., Rao B. J., Radding C. M. Ability of RecA protein to promote a search for rare sequences in duplex DNA. Proc Natl Acad Sci U S A. 1986 Dec;83(24):9586–9590. doi: 10.1073/pnas.83.24.9586. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Kirkpatrick D. P., Rao B. J., Radding C. M. RNA-DNA hybridization promoted by E. coli RecA protein. Nucleic Acids Res. 1992 Aug 25;20(16):4339–4346. doi: 10.1093/nar/20.16.4339. [DOI] [PMC free article] [PubMed] [Google Scholar]
- McEntee K., Weinstock G. M., Lehman I. R. Binding of the recA protein of Escherichia coli to single- and double-stranded DNA. J Biol Chem. 1981 Aug 25;256(16):8835–8844. [PubMed] [Google Scholar]
- Melton D. A., Krieg P. A., Rebagliati M. R., Maniatis T., Zinn K., Green M. R. Efficient in vitro synthesis of biologically active RNA and RNA hybridization probes from plasmids containing a bacteriophage SP6 promoter. Nucleic Acids Res. 1984 Sep 25;12(18):7035–7056. doi: 10.1093/nar/12.18.7035. [DOI] [PMC free article] [PubMed] [Google Scholar]
- NYGAARD A. P., HALL B. D. FORMATION AND PROPERTIES OF RNA-DNA COMPLEXES. J Mol Biol. 1964 Jul;9:125–142. doi: 10.1016/s0022-2836(64)80095-4. [DOI] [PubMed] [Google Scholar]
- Shibata T., Cunningham R. P., Radding C. M. Homologous pairing in genetic recombination. Purification and characterization of Escherichia coli recA protein. J Biol Chem. 1981 Jul 25;256(14):7557–7564. [PubMed] [Google Scholar]
- Vary C. P. A homogeneous nucleic acid hybridization assay based on strand displacement. Nucleic Acids Res. 1987 Sep 11;15(17):6883–6897. doi: 10.1093/nar/15.17.6883. [DOI] [PMC free article] [PubMed] [Google Scholar]