Method for pair-wise sequencing a plurality of double stranded target polynucleotides
First Claim
1. A method for pairwise sequencing of first and second regions in individual double stranded target polynucleotides of a plurality of double stranded target polynucleotides, wherein each of the double stranded target polynucleotides comprises a first strand and a complementary strand, comprising(a) providing a solid support,wherein the first strands of the plurality of double stranded target polynucleotides are linked to the solid support,wherein a plurality of primers is linked to the solid support, andwherein the primers are capable of hybridizing to a first region of the first strands;
- (b) obtaining sequence data from the first region of the first strands by subjecting the first strand region of the first strands on the solid support to a sequencing method;
(c) hybridizing the primers to the first strands after obtaining the sequence data from the first region of the first strands and then extending the primers to copy the first strands, thereby generating the complementary strands of the double stranded target polynucleotides, whereby the complementary strands are linked to the solid support; and
(d) obtaining sequence data for a second region of the second strands, wherein the sequence data for the first region and the second region provides a pairwise sequence of the first and second regions of individual target polynucleotides in the plurality of double stranded polynucleotides.
2 Assignments
0 Petitions
Accused Products
Abstract
The invention relates to methods for pairwise sequencing of a double-stranded polynucleotide template, which methods result in the sequential determination of nucleotide sequences in two distinct and separate regions of the polynucleotide template. Using the methods of the invention it is possible to obtain two linked or paired reads of sequence information from each double-stranded template on a clustered array, rather than just a single sequencing read from one strand of the template.
-
Citations
17 Claims
-
1. A method for pairwise sequencing of first and second regions in individual double stranded target polynucleotides of a plurality of double stranded target polynucleotides, wherein each of the double stranded target polynucleotides comprises a first strand and a complementary strand, comprising
(a) providing a solid support, wherein the first strands of the plurality of double stranded target polynucleotides are linked to the solid support, wherein a plurality of primers is linked to the solid support, and wherein the primers are capable of hybridizing to a first region of the first strands; -
(b) obtaining sequence data from the first region of the first strands by subjecting the first strand region of the first strands on the solid support to a sequencing method; (c) hybridizing the primers to the first strands after obtaining the sequence data from the first region of the first strands and then extending the primers to copy the first strands, thereby generating the complementary strands of the double stranded target polynucleotides, whereby the complementary strands are linked to the solid support; and (d) obtaining sequence data for a second region of the second strands, wherein the sequence data for the first region and the second region provides a pairwise sequence of the first and second regions of individual target polynucleotides in the plurality of double stranded polynucleotides. - View Dependent Claims (2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17)
-
Specification