Single molecule sequencing of captured nucleic acids
View Patent ↗The invention provides methods and devices for detecting, enumerating or identifying target nucleic acid molecules using immobilized capture probes and single molecule sequencing techniques.
1. A method for obtaining sequence information from a selected nucleic acid, the method comprising the steps of:
capturing target nucleic acids with a sequence-specific capture probes to produce a plurality of target/probe duplexes, wherein the target nucleic acids comprise RNA;
melting the target/probe duplexes to release target nucleic acids;
annealing primers to released target nucleic acids to produce target/primer duplexes;
introducing a polymerase and at least one nucleotide species comprising an optically-detectable label under conditions sufficient for template-dependent nucleotide addition to said primer;
removing unincorporated nucleotides;
identifying nucleotide species incorporated into said primer, thereby obtaining sequence information from said target nucleic acids.
2. The method according to claim 1 , wherein the optically detectably labeled nucleotide is fluorescently labeled nucleotide.
3. The method according to claim 1 , wherein the target nucleic acids comprise adaptor sequences.
4. The method according to claim 3 , wherein different target nucleic acids have the same or different adaptor sequences.
5. The method of claim 1 wherein the probes are a plurality of different sequence-specific probes.