Patent ID: 7638310

Claim:
A method to determine single nucleotide polymorphism (SNP) in a target nucleic acid sequence, said method comprising the steps of: a) providing a template nucleic acid, consisting of a first and a second strand and having one unknown nucleotide pair; b) APEX-2 (Arrayed Primer Extension using Universal primer) oligonucleotide primer; said first APEX-2 primer consisting of a first specific oligonucleotide sequence in its 3′-end and a universal sequence in its 5′end, said universal sequence further having a modification in its 5′end, and said first specific oligonucleotide sequence being complementary to nucleotides before the unknown nucleotide in the first strand of the template nucleic acid; said second APEX-2 primer consisting of a second specific oligonucleotide sequence in its 3′-end and a universal sequence in its 5′end, said universal sequence optionally having a modification in its 5′end and, said second specific oligonucleotide sequence being complementary to nucleotides before the unknown nucleotide in the second strand of the template nucleic acid; c) amplifying the unknown nucleotide of the first and the second strand of the template nucleic acid by running a multiplex PCR primer extension reaction with the first and the second APEX-2 primers as templates, thereby producing a first amplification product, said first amplification product consisting of a first and a second strand, said first strand consisting of sequence of the first APEX-2 primer and a complementary sequence to the second APEX-2 primer and the unknown nucleotide between them, and the second strand consisting of complementary sequence to the APEX-2 first primer and the sequence of the second APEX-2 primer and the unknown nucleotide between them; d) amplifying the first amplification product in a PCR reaction with universal primer as template, thereby producing a second amplification product; e) providing a microarray of probes, where probes are identical to the first and the second APEX-2 primers of step b) and where the probes are immobilized on the array by attaching them to a solid surface from the modified 5′end; f) letting the second amplification products anneal with immobilized probes and providing labeled or modified terminator nucleotides for single base extension reaction; and g) identifying terminating nucleotide that has been added to the probes, said terminating nucleotides corresponding to the unknown nucleotides of the template nucleic acid.