Patent ID: 6090552
Filing Date: 2000-07-18
Classification: C12Q

Abstract:
A method of detecting telomerase activity comprising:(a) contacting a sample suspected of having telomerase activity with at least two oligonucleotide primers comprising a first primer and a second primer, wherein said first primer comprises the following continuous sequences in 5' to 3' order:(i) a first nucleotide sequence of 6-30 nucleotides, wherein a nucleotide within said first nucleotide sequences is labeled with a first moiety selected from the group consisting of a donor moiety and an acceptor moiety of a molecular energy transfer pair, wherein the donor moiety emits energy of one or more particular wavelengths when excited, and the acceptormoiety absorbs energy at one or more particular wavelengths emitted by the donor moiety;(ii) a second, single-stranded nucleotide sequence of 3-20 or nucleotides;(iii) a third nucleotide sequence of 6-30 nucleotides, wherein a nucleotides within said third nucleotide sequence is labeled with a second moiety selected from the group consisting of said donor moiety and said acceptor moiety, and said second moiety is the member of said group not labeling said first nucleotide sequence, wherein said third nucleotide sequence is sufficiently complementary in reverse order to said first nucleotide sequence for a duplex to form between said first nucleotide sequence and said third nucleotide sequence such that said first moiety and second moiety are in sufficient proximity such that, when the donor moiety is excited and emits energy, the acceptor moiety absorbs energy emitted by the donor moiety; and(iv) at the 3' end of said first primer, a fourth, single-stranded nucleotide sequence of 8-40 nucleotides that comprises at its 3' end a sequence that is a substrate for a telomerase; wherein said second primer comprises at its 3' end a sequence sufficiently complementary so as to be able to hybridize to telomeric repeats that result from the activity of said telomerase;(b) subjecting the sample to conditions suitable for telomerase activity;(c) conducting a nucleic acid amplification reaction under conditions suitable for said first and second primers to prime DNA synthesis;(d) stimulating energy emission from said donor moiety; and(e) detecting or measuring energy emitted by said donor moiety or acceptor moiety, the presence or amount of said energy indicating the presence or amount of telomerase activity in the sample.