title: Multiplex PCR: use of heat-stable Thermus thermophilus RecA protein to minimize non-specific PCR products.
abstract: In this paper we report that the inclusion of heat-resistant RecA protein from a thermophilic bacteria, Thermus thermophilus, and its cofactor (ATP) in PCR effectively eliminates non-specific PCR products. The effect of RecA protein, which catalyzes pairing between homologous DNA molecules with great fidelity in genetic recombination, is due to its promotion of precise priming in PCR (i.e. priming at sites where the primer sequence is completely complementary to that of the target sequence). In addition, the RecA protein substantially reduces the primer concentration required for PCR. These experimental results have led to the realization of multiplex PCR, which involves PCR for multiple sites in the same reaction mixture. We were able to successfully perform multiplex PCR with over a dozen reactions without affecting the amplification pattern of the PCR products.
authors.affiliation: Kazusa DNA Research Institute 2-6-7 Kazusa-kamatari, Kisarazu, Chiba 292-0818, Japan.
authors.forename: Yasushi
authors.initials: Y
authors.lastname: Shigemori
authors.forename: Tsutomu
authors.initials: T
authors.lastname: Mikawa
authors.forename: Takehiko
authors.initials: T
authors.lastname: Shibata
authors.forename: Michio
authors.initials: M
authors.lastname: Oishi
mesh_terms: D020137:Base Pair Mismatch; D004247:DNA / Q000096:biosynthesis; D017931:DNA Primers / Q000737:chemistry; D006358:Hot Temperature; D006801:Humans; D016133:Polymerase Chain Reaction / Q000379:methods*; D011938:Rec A Recombinases / Q000378:metabolism*; D016963:Thermus thermophilus
chemical_list: D017931:DNA Primers; D004247:DNA; D011938:Rec A Recombinases