ONE ARMED PCR (OA-PCR) - AMPLIFICATION OF GENOMIC DNA FROM A SINGLE PRIMER DOMAIN

One-armed PCR (OA-PCR) is a novel technique that allows amplification of genomic DNA from a single region of known sequence. Previously described methods are either limited to cDNA or require extensive manipulation of template prior to amplification. In OA-PCR, DNA from a small insert phage Fugu genomic library is amplified using two sequence-specific primers and a tailed vector primer. Single, specific products of between 70 bp and 1.9 kb (mean 600 bp) have been obtained following two rounds of amplification and directly sequenced without cloning. The optimum parameters have been investigated: primer pairs of 18-24 bp, separated by between 0 and 500 bp, have successfully amplified a specific product. OA-PCR represents a rapid, simple method to extend genomic sequence into noncoding and regulatory sequences. In addition, degenerate OA-PCR primer pairs have allowed cross-species amplification of novel Fugu gene homologues. (C) 1994 Academic Press, Inc.