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US 2013/0171648 A1Published application
Method for sequencing a polynucleotide template
AssigneeIllumina Cambridge Limited
InventorGEOFFREY PAUL SMITH
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Abstract
The invention relates to methods for pairwise sequencing of a double-stranded polynucleotide template, which permit the sequential determination of nucleotide sequences in two distinct and separate regions on complementary strands of the double-stranded polynucleotide template. The two regions for sequence determination may or may not be complementary to each other.
Claims18
1-14. (canceled)
A method for pairwise sequencing of first and second strands of target polynucleotides, wherein the first and second strands of each target polynucleotide have complementary sequences, the method comprising:
- (a) forming a plurality of nucleic acid colonies by solid phase nucleic acid amplification, wherein each of the nucleic acid colonies is formed by amplification of a single originating target polynucleotide;
- (b) hybridizing first sequencing primers to first strands of the colonies;
- (c) carrying out a first sequencing reaction by sequential addition of nucleotides to the first sequencing primers to determine sequences from the first strands;
- (d) cleaving the first strands;
- (e) removing at least a portion of the first strands from the colonies;
- (f) hybridizing second sequencing primers to second template strands of the colonies;
- (g) carrying out a second sequencing reaction by sequential addition of nucleotides to the second sequencing primers to determine sequences from the second template strands, wherein the first sequencing reaction and the second sequencing reaction permit pairwise sequencing of the target polynucleotides.
4 more elements
Dependencies are inferred from the claim text.