AMPLIFICATION OF TARGET NUCLEOTIDE SEQUENCE WITHOUT POLYMERASE CHAIN REACTION

摘要

The present disclosure relates to methods for generating single-stranded DNA molecules of defined sequence and length using ligase chain reaction (LCR). Specifically, a region of template containing target sequence is amplified by LCR, exogenous sequence is introduced by LCR primers or probes used in amplification, and LCR products may be used in further amplification steps involving rolling circle amplification (RCA) or polymerase chain reaction (PCR). LCR products may include sequence complementary to the backbone of a padlock probe, where the LCR product hybridizes to a padlock probe and after ligation of the padlock, serves as polymerazation primer. After amplification, single-stranded amplification products are trimmed to produce short single-stranded DNA molecules of defined sequence and length.