Alternative nucleic acid sequencing methods

a technology of nucleic acid and composition, applied in the field of methods and compositions for nucleic acid sequencing, can solve problems such as uncertainty in the obtained sequen

Inactive Publication Date: 2009-03-05
APPL BIOSYSTEMS INC
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  • Abstract
  • Description
  • Claims
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Problems solved by technology

However, the various techniques often have problems t

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  • Alternative nucleic acid sequencing methods
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Examples

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example 1

[0093]This example illustrates possible methods for analysis of a polynucleotide sequence by sequencing two regions of the polynucleotide using sequencing by ligation and pyrosequencing.

[0094]The polynucleotide for sequencing is prepared by fragmenting the polynucleotide and clonally amplifying the fragments through emulsion PCR as described below. After amplification, a first region of the polynucleotide is subjected to sequencing using sequencing by ligation, as described below, to determine a first sequence (FIG. 3 at 70). Next, a second region of the polynucleotide is subjected to pyrosequencing, as described below, using to determine a second sequence (FIG. 3 at 80). The first sequence is compared to the second sequence (FIG. 3 at 90). Concordance or discordance between the data from the two different chemistries can be taken into account when making a final base call for a given position. Variations in the base and sequence biases of the different chemistries may be also be ta...

example 2

[0095]This example illustrates possible methods for analysis of a polynucleotide sequence by sequencing two regions of the polynucleotide using sequencing by ligation and pyrosequencing.

[0096]The polynucleotide for sequencing is prepared by fragmenting the polynucleotide and clonally amplifying the fragments through emulsion PCR as described below. After amplification, a first region of the polynucleotide is subjected to sequencing using sequencing by ligation, as described below, to determine a first sequence. Next, a second region of the polynucleotide is subjected to sequencing using reversible terminators to determine a second sequence. The first sequence is compared to the second sequence. Concordance or discordance between the data from the two different chemistries can be taken into account when making a final base call for a given position. Variations in the base and sequence biases of the different chemistries may be also be taken into account when making a final sequence d...

example 3

[0097]This example illustrates possible methods for analysis of a polynucleotide sequence by sequencing two regions of the polynucleotide using sequencing by ligation and pyrosequencing.

[0098]The polynucleotide for sequencing is prepared by fragmenting the polynucleotide and clonally amplifying the fragments through emulsion PCR as described below. After amplification, a first region of the polynucleotide is subjected to sequencing using reversible terminators to determine a first sequence. Next, a second region of the polynucleotide is subjected to pyrosequencing, as described below, using to determine a second sequence. The first sequence is compared to the second sequence. Concordance or discordance between the data from the two different chemistries can be taken into account when making a final base call for a given position. Variations in the base and sequence biases of the different chemistries may be also be taken into account when making a final sequence determination.

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Abstract

Embodiments are provided that provide for parallel sequencing of nucleic acid segments. In some embodiments, a single sequence is sequenced by at least two different sequencing techniques and the results compared, allowing for deficiencies or strengths of one technique to be complemented by the second technique.

Description

PRIORITY[0001]This Application claims priority under 35 U.S.C. §119(e) to U.S. Provisional Patent Application Ser. No. 60 / 968,834, filed Aug. 29, 2007, herein incorporated by reference in its entirety.FIELD[0002]The present disclosure relates to methods and compositions for nucleic acid sequencing.INTRODUCTION[0003]A large variety of techniques exist for sequencing nucleic acids. However, the various techniques often have problems that can lead to uncertainty in the obtained sequence.SUMMARY[0004]The present teachings provide, among other things, methods and apparatuses that facilitate more effective sequencing, such as through more accurate nucleic acid sequencing and / or greater read length.[0005]Various embodiments of a method of the present teachings comprise: determining a sequence of a first region of a polynucleotide using a first set of nucleic acid sequencing reagents; and determining a sequence of a second region of the polynucleotide using a second set of nucleic acid sequ...

Claims

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Application Information

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IPC IPC(8): C40B30/00C12Q1/68C40B40/06G16B30/20
CPCC12Q1/6869C12Q1/6874G06F19/22C12Q2565/301C12Q2537/149C12Q2535/101C12Q2565/627C12Q2533/107C40B20/08G16B30/00G16B30/20
Inventor BORNER, SCOTT R.
Owner APPL BIOSYSTEMS INC
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