Conditions for using DNA polymerase I as an RNA-dependent DNA polymerase.
Conditions are described for using Escherichia coli DNA polymerase I for synthesizing complementary DNA copies of natural RNA molecules, which are suitable for use in hybridization experiments. The molar ratio of enzyme to template is critical; below a certain level, synthesis is not observed. Hybrids formed with the complementary DNA are of comparable specificity and stability to those formed with complementary DNAs synthesized by viral RNA-directed DNA polymerase. Synthesis of dA-dT polymers, a common occurrence with this enzyme, can be eliminated by including distamycin in the reaction mixture.
D000818:Animals; D001354:Avian Leukosis Virus / Q000201:enzymology; D002499:Centrifugation, Density Gradient; D002845:Chromatography; D003597:Cytosine Nucleotides / Q000378:metabolism; D004247:DNA / Q000032:analysis / Q000096:biosynthesis; D003609:Dactinomycin; D004330:Drosophila; D004926:Escherichia coli / Q000201:enzymology*; D005914:Globins; D006160:Guanosine Triphosphate / Q000378:metabolism; D006358:Hot Temperature; D006882:Hydroxyapatites; D007700:Kinetics; D008970:Molecular Weight; D009693:Nucleic Acid Hybridization; D012333:RNA, Messenger / Q000378:metabolism; D012335:RNA, Ribosomal; D012367:RNA, Viral; D012194:RNA-Directed DNA Polymerase / Q000378:metabolism*; D011817:Rabbits; D013698:Templates, Genetic; D014316:Tritium
D003597:Cytosine Nucleotides; D006882:Hydroxyapatites; D012333:RNA, Messenger; D012335:RNA, Ribosomal; D012367:RNA, Viral; D014316:Tritium; D003609:Dactinomycin; D006160:Guanosine Triphosphate; D005914:Globins; D004247:DNA; D012194:RNA-Directed DNA Polymerase