Primase from Escherichia coli primes single-stranded templates in the absence of single-stranded DNA-binding protein or other auxiliary proteins. Template sequence requirements based on the bacteriophage G4 complementary strand origin and Okazaki fragment initiation sites.

摘要: Abstract We find that neither the hairpins nor bound single-stranded DNA-binding protein (SSB) are required for in vitro primase activity at G4 complementary strand origin. Primase has ability to prime synthetic DNA templates absence of any auxiliary proteins, including SSB, and secondary structure. does, however, initiate primer synthesis starting a specific nucleotide sequence present both origin system sites Okazaki fragment initiation. tested series consisted various portions support synthesis. Primer was determined using an assay which length quantity monitored by incorporation [gamma-32P]ATP once per products analyzed on denaturing polyacrylamide gel. requires only short template, 5'-(AC)7CTG CAA AGC-3', synthesize 17-nucleotide thymidine residue. also report primase's incorporate dideoxyribonucleotide triphosphates into primers, allowed us determine directly primers synthesized.