Parallel primer extension approach to nucleic acid sequence analysis

A method for determining nucleotide identity of at least one nucleotide position of a polynucleotide of interest, comprising the steps of;

• a) contacting said polynucleotide of interest with a population of single-stranded primers, wherein said single-stranded primers comprise an array of one or more sets of one or more oligonucleotides, wherein at least one set comprises at least two oligonucleotides that are substantially homologous but differ from each other by one base at their 3′ termini, wherein the oligonucleotides of the array have known sequence and wherein each oligonucleotide is attached to a solid support at a known location, to form the array, wherein at least one oligonucleotide of the array hybridizes to said polynucleotide of interest immediately adjacent to each nucleotide position to be identified, generating template-single-stranded primer complexes;
• b) subjecting said complexes to a single base extension reaction to extend each annealed primer by a terminating nucleotide, generating extended primers;
• c) identifying each terminating nucleotide that has been added to each primer;
• thereby determining the identity of at least one nucleotide position of a polynucleotide of interest;
• and
• d) removing the terminating nucleotides from the annealed primers after completed analysis to prepare the solid support for reuse.