Simple in vitro single-stranded linear and circular DNA preparation, functional selection, and validation using phosphor-derived modifications.

ABSTRACT: Interest in the preparation of circular ssDNA library has been increasing recently; therefore, developing a simple and an efficient method for circular DNA generation will be very useful for all procedures and techniques that are dependent on circular ssDNA preparation. In this study, a new simple method for in vitro preparation of circular ssDNA is proposed. We hypothesized that using a phosphorylated-phosphorothioated primer would not affect the efficiency of PCRs but, more importantly, would suppress the activity of the lambda exonuclease enzyme even if it is phosphorylated. The produced phosphorylated ssDNA is ready to be circularized via a ligation reaction using a bridging oligonucleotide. Several optimizations and enhancements have been conducted in the ligation reaction, notably by

SUBMITTER: Hamidi SV 

PROVIDER: S-EPMC12721166 | biostudies-literature | 2025 Oct

REPOSITORIES: biostudies-literature