Method for accurate sequencing of DNA
View Patent ↗DNA is sequenced by (a) independently sequencing first and second strands of a dsDNA to obtain corresponding first and second sequences; and (b) combining the first and second sequences to generate a consensus sequence of the dsDNA. By independently sequencing first and second strands the error probability of the consensus sequence approximates a multiplication of those of the first and second sequences.
1. A method of sequencing DNA comprising:
a) independently sequencing first and second strands of a dsDNA by sequencing multiple copies of the first and second strands to obtain first and second copy sequences;
b) combining the copy sequences of each strand to obtain first and second strand consensus sequences; and
c) combining the first and second strand consensus sequences to generate a consensus sequence of the dsDNA,
further comprising the step of identifying one or more correspondences between the first and second copy sequences and a genomic DNA sequence by metagenomic analysis.
2. The method of claim 1 wherein prior to the sequencing step the method further comprises:
amplifying the dsDNA to obtain the multiple copies of the first and second strands.
3. A method of sequencing DNA comprising:
a) independently sequencing first and second strands of a dsDNA by sequencing multiple copies of the first and second strands to obtain first and second copy sequences;
b) combining the copy sequences of each strand to obtain first and second strand consensus sequences; and
c) combining the first and second strand consensus sequences to generate a consensus sequence of the dsDNA,
further comprising the steps of:
PCR amplifying a dsDNA fragment having a sequence flanked by Y-adapters to produce asymmetrical, amplified dsDNA;
denaturing the amplified dsDNA and attaching and then bridge amplifying resultant amplified ssDNA at discrete locations of a flow cell to produce polonies;
reading the sequences of the polonies; and
identifying same-sized complementary sequences, which provide the first and second copy sequences corresponding to the first and second strands of the dsDNA.
4. The method of claim 3 wherein the Y-adapters are randomized indexing adapters or are different adapters produced with two consecutive ligations, which ligations could be separated by fragmentation and amplification steps.
5. The method of claim 1 further comprising the step of identifying one or more correspondences between the first and second copy sequences and a genomic DNA sequence by mapping the first and second copy sequences to the genomic DNA sequence.
6. The method of claim 1 further comprising the step of identifying one or more correspondences between the first and second copy sequences and a genomic DNA sequence by relating one or more mutations of the genomic DNA sequence to the first and/or second copy sequence.
7. The method of claim 3 further comprising the step of identifying one or more correspondences between the first and second copy sequences and a genomic DNA sequence by mapping the first and second copy sequences to the genomic DNA sequence.
8. The method of claim 3 further comprising the step of identifying one or more correspondences between the first and second copy sequences and a genomic DNA sequence by metagenomic analysis.
9. The method of claim 3 further comprising the step of identifying one or more correspondences between the first and second copy sequences and a genomic DNA sequence by relating one or more mutations of the genomic DNA sequence to the first and/or second copy sequence.
10. The method of claim 1 wherein each of the copy sequences is a composite sequence, and the method further comprises generating each composite sequence from multiple reads of each copy.
11. The method of claim 2 wherein each of the copy sequences is a composite sequence, and the method further comprises generating each composite sequence from multiple reads of each copy.
12. The method of claim 3 wherein each of the copy sequences is a composite sequence, and the method further comprises generating each composite sequence from multiple reads of each copy.
13. The method of claim 4 wherein each of the copy sequences is a composite sequence, and the method further comprises generating each composite sequence from multiple reads of each copy.
14. The method of claim 5 wherein each of the copy sequences is a composite sequence, and the method further comprises generating each composite sequence from multiple reads of each copy.
15. The method of claim 6 wherein each of the copy sequences is a composite sequence, and the method further comprises generating each composite sequence from multiple reads of each copy.
16. The method of claim 7 wherein each of the copy sequences is a composite sequence, and the method further comprises generating each composite sequence from multiple reads of each copy.
17. The method of claim 8 wherein each of the copy sequences is a composite sequence, and the method further comprises generating each composite sequence from multiple reads of each copy.
18. The method of claim 9 wherein each of the copy sequences is a composite sequence, and the method further comprises generating each composite sequence from multiple reads of each copy.