Optimally labeled oligonucleotides
AssigneePoly Probe, Inc.
InventorThor W. Nilsen
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Abstract
A method for the preparation of optimally labeled oligonucleotides wherein label-conjugated nucleotide triphosphates are incorporated into a nucleic acid sequence in a defined repetitive manner which allows for the optimal specific detectability of the oligonucleotide. The oligonucleotides of the present invention are useful in the assay of a wide variety of nucleic acid sequences, specifically wherever labeled nucleic acid probes are desired.
Claims25
A method for the preparation of optimally labeled oligonucleotides, said method comprising the steps of:
- (a) preparing a primer;
- (b) preparing a template oligonucleotide, said template oligonucleotide containing a nucleotide sequence complementary to said primer, and a nucleotide repeat region downstream from said complementary region;
- (c) annealing the template and the primer in a suitable reaction medium, said reaction medium containing a polymerase, nucleotide triphosphates and label-conjugated nucleotide triphosphates;
- (d) initiating synthesis of a complementary strand on the template;
- (e) attaching said oligonucleotide containing a target sequence adjacent to said complementary strand;
- and
- (f) purifying said optimally labeled oligonucleotide by any appropriate method.
A method for the preparation of optimally labeled oligonucleotides, said method comprising the steps of:
- (a) preparing a template, said template comprising a primer binding region, a 5′ extension region for the subsequent incorporation of label-conjugated nucleotide triphosphates, and a 3′ overhang region of 6-200 nucleotides;
- and
- (b) labeling an oligonucleotide target sequence by denaturing the target sequence, adding excess template, the appropriate nucleotide triphosphates and polymerase in a suitable reaction medium.
An oligonucleotide comprising a nucleotide sequence complementary to a primer, and a nucleotide repeat region downstream from said complementary region, wherein said nucleotide repeat region comprises Nt, where Nt is any nucleotide which can form a base pair with the labeled-conjugated nucleotide triphosphate, said repeat region having the formula:
- Nt Nt)nNt where n is an integer from 20 to 1000.
Dependencies are inferred from the claim text.