Abstract: The invention comprises a two-step process for analysis of polynucleotides by chain extension of multiple polynucleotide primers attached to solid supports by first performing PCR of the samples in the presence of multiple oligonucleotides in solution, the oligonucleotides of both sets being similar or identical. This produces immobilized single-strand polynucleotides containing genetic sequence data derived from sample molecules. In a second step, support-bound polynucleotides are interrogated by hybridization with a single labeled oligonucleotide probe or by second-strand synthesis with a primer-dependent polymerase using an oligonucleotide primer and nucleotide monomers, in which either or both of the primer and nucleotide monomers are labeled. Incorporation of label demonstrates the presence of two separate defined-sequence primers within the sample polynucleotide.

Abstract: The invention comprises a two-step process for analysis of polynucleotides by chain extension of multiple polynucleotide primers attached to solid supports by first performing PCR of the samples in the presence of multiple oligonucleotides in solution, the oligonucleotides of both sets being similar or identical. This produces immobilized single-strand polynucleotides containing genetic sequence data derived from sample molecules. In a second step, support-bound polynucleotides are interrogated by hybridization with a single labeled oligonucleotide probe or by second-strand synthesis with a primer-dependent polymerase using an oligonucleotide primer and nucleotide monomers, in which either or both of the primer and nucleotide monomers are labeled. Incorporation of label demonstrates the presence of two separate defined-sequence primers within the sample polynucleotide.