Patent ID: 6258533
Filing Date: 2001-07-10
Classification: C12Q

Abstract:
A sequencing method for identifying a first nucleotide n and a second nucleotide n+x in a double stranded nucleic acid segment, comprising:a) digesting said double stranded nucleic acid segment with a restriction enzyme whose cleavage site is separate from its recognition site to produce a double stranded molecule having a single stranded overhang sequence corresponding to an enzyme cut site;b) providing an adaptor having a cycle identification tag, a restriction enzyme recognition domain and a sequence identification region;c) hybridizing said adaptor to said double stranded nucleic acid having said single-stranded overhang sequence to form a ligated molecule;d) amplifying said ligated molecule from step (c) with a labeled primer specific for said cycle identification tag, restriction enzyme recognition domain, and a portion of said sequence identification region of said adaptor;e) identifying said nucleotide n by identifying said primer incorporated into the amplification product; andf) repeating steps (a) through (e) on said amplified molecule from step (e) to yield the identity of said nucleotide n+x, wherein x is less than or equal to the number of nucleotides between a recognition domain for a restriction enzyme and an enzyme cut site.