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US 2006/0040282 A1Published application
Oligonucleotide sizing using cleavable primers
AssigneeSequenom, Inc.
InventorsJoseph Monforte, Christopher Becker, Thomas Shaler, Daniel Pollart
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Abstract
The present invention provides modified oligonucleotide primers designed to incorporate a cleavable moiety so that a 3′ portion of the primer (linked to an extension product) can be released from an upstream 5′ portion of the primer. Upon selective cleavage of the cleavable site, primer extension products that contain about five or fewer base pairs of the primer sequence are released, to provide more useful sizing and sequence information per fragment than extension products containing the entire primer.
Claims19
A method for determining presence of a polymorphism, comprising:
- (a) hybridizing a primer, having a 5′ end and a 3′ end, with a target nucleic acid suspected of containing a polymorphism, wherein said primer has a first region containing the 5′ end of the primer and a second region containing the 3′ end of the primer and a cleavable site;
- (b) extending the 3′ end of the primer with a polymerase in the presence of a nucleotide to generate an extension product;
- (c) cleaving said extension product at the cleavable site to release an extension segment;
- sizing the extension segment by mass spectrometry, whereby said cleaving is effective to increase the read length of the extension segment relative to the read length of the product of step (b);
- and
- identifying any added nucleotides.
3 more elements
Dependencies are inferred from the claim text.