Methods and devices for multiplexing amplification reactions
The present invention provides a two-step multiplex amplification reaction wherein the first step truncates the standard initial multiplex amplification round to “boost” the sample copy number by only a 100–1000 fold increase in the target. Following the first step the product is divided into optimized secondary single amplification reactions, each containing one of the primer sets that were used previously in the first or multiplexed booster step. The booster step can occur using an aqueous target nucleic acid or using a solid phase archived nucleic acid. In particular, nucleic acid sequences that uniquely identify E. Coli were identified using the multiplex amplification method.
1. A method for discriminately identifying nucleic acid sequences of E. coli bacteria of the coliform species in a sample, comprising:
(a) contacting said sample with a primer comprising a sequence identified by SEQ ID. NO 85;
(b) amplifying said nucleic acid sequence; and
(c) detecting said amplified nucleic acid sequence wherein said amplified nucleic acid sequence discriminately identifies E. coli bacteria.