US 2014/0235461 A1 · ALGORITHMS FOR SEQUENCE DETERMINATIONS

A computer-implemented method of developing a consensus sequence from a plurality of sequencing reads of a nucleic acid target, comprising

(a) receiving a population of raw target sequencing reads of a nucleic acid target comprising an anchor segment and an adjacent segment;
the anchor segment being of known sequence and the adjacent segment being of unknown sequence;
and at least some of the raw target sequencing reads containing sequencing errors;
(b) evaluating the accuracy of sequencing of the anchor segment in different raw target sequencing reads by comparing raw target sequencing reads of the anchor segment with the known sequence of the anchor segment;
(c) assigning a subset of the raw target sequencing reads into an accepted class based on reaching at least a threshold level of accuracy of the sequencing of the anchor segment;
(d) polling nucleobases at a position equidistant to the anchor segment sequence in raw target sequencing reads in the accepted class to determine a consensus nucleobase, which consensus nucleobase is assigned as the first nucleobase of a nascent sequence of the adjacent segment;
(e) assigning raw target sequencing reads having the consensus nucleobase determined in the prior polling step to remain in an accepted class and assigning raw target sequencing reads lacking the consensus nucleobase determined in the prior polling step to the rejected class;
(f) optionally reassigning a raw target sequencing read from the rejected class to the accepted class by scoring similarity of the raw target sequencing read to the nascent sequence and reintroducing the raw target sequencing read if the sequence similarity reaches at least a threshold level of similarity;
and
(g) repeating steps (d), (e) and optionally (f), except that a repetition polls a position adjacent the position poled in the previous polling step for raw target sequencing reads having the consensus nucleobase polled in the previous step or in the case of a raw target sequencing read reassigned from the rejected class to the accepted class and not polled in the previous polling step or if polled not having the consensus nucleobase in the previous polling step, the polling polls a position adjacent the position aligned with the last nucleobase of the nascent sequence to determine a consensus nucleobase, and the consensus nucleobases determined in successive repetitions are assigned as successive nucleobases in the nascent sequence of the adjacent segment.

7 more elements

A computer program product for analyzing a nucleic acid target, comprising

(a) code for receiving a population of raw target sequencing reads of a nucleic acid target comprising an anchor segment and an adjacent segment;
the anchor segment being of known sequence and the adjacent segment being of unknown sequence;
and at least some of the raw target sequencing reads containing sequencing errors;
(b) code for evaluating the accuracy of sequencing of the anchor segment in different raw target sequencing reads by comparing the raw target sequencing reads of the anchor segment with the known sequence of the anchor segment;
(c) code for assigning a subset of the raw target sequencing reads into an accepted class based on reaching at least a threshold level of accuracy of the sequencing of the anchor segment;
(d) code for polling nucleobases at a position equidistant to the anchor segment sequence in raw target sequencing reads in the accepted class to determine a consensus nucleobase, which consensus nucleobase is assigned as the first nucleobase of a nascent sequence of the adjacent segment;
(e) code for assigning raw target sequencing reads having the consensus nucleobase determined in the prior polling step to remain in an accepted class and assigning raw target sequencing reads lacking the consensus nucleobase determined in the prior polling step to the rejected class;
(f) code for optionally reassigning a raw target sequencing read from the rejected class to the accepted class by scoring similarity of the raw target sequencing read to the nascent sequence and reintroducing the raw target sequencing read if the sequence similarity reaches at least a threshold level of similarity;
and
(g) code for repeating steps coded in (d), (e) and optionally (f), except that a repetition polls a position adjacent the position poled in the previous polling step for raw target sequencing reads having the consensus nucleobase polled in the previous step or in the case of a raw target sequencing read reassigned from the rejected class to the accepted class and not polled in the previous polling step or if polled not having the consensus nucleobase in the previous polling step, the polling polls a position adjacent the position aligned with the last nucleobase of the nascent sequence to determine a consensus nucleobase, and the consensus nucleobases determined in successive repetitions are assigned as successive nucleobases in the nascent sequence of the adjacent segment.

7 more elements

A system for analyzing a nucleic acid target, comprising:

(1) a system bus;
(2) a memory coupled to the system bus;
and (3) a processor coupled to the system bus operatively disposed to:
(a) receive a population of raw target sequencing reads of a nucleic acid target comprising an anchor segment and an adjacent segment;
the anchor segment being of known sequence and the adjacent segment being of unknown sequence;
and at least some of the raw target sequencing reads containing sequencing errors;
(b) evaluate the accuracy of sequencing of the anchor segment in different raw target sequencing reads by comparing raw target sequencing reads of the anchor segment with the known sequence of the anchor segment;
(c) assign a subset of the raw target sequencing reads into an accepted class based on reaching at least a threshold level of accuracy of the sequencing of the anchor segment;
(d) poll nucleobases at a position equidistant to the anchor segment sequence in raw target sequencing reads in the accepted class to determine a consensus nucleobase, which consensus nucleobase is assigned as the first nucleobase of a nascent sequence of the adjacent segment;
(e) assign raw target sequencing reads having the consensus nucleobase determined in the prior polling step to remain in an accepted class and assigning raw target sequencing reads lacking the consensus nucleobase determined in the prior polling step to the rejected class;
(f) optionally reassign a raw target sequencing read from the rejected class to the accepted class by scoring similarity of the raw target sequencing read to the nascent sequence and reintroducing the raw target sequencing read if the sequence similarity reaches at least a threshold level of similarity;
and
(g) repeat steps (d), (e) and optionally (f), except that a repetition polls a position adjacent the position poled in the previous polling step for raw target sequencing reads having the consensus nucleobase polled in the previous step or in the case of a raw target sequencing read reassigned from the rejected class to the accepted class and not polled in the previous polling step or if polled not having the consensus nucleobase in the previous polling step, the polling polls a position adjacent the position aligned with the last nucleobase of the nascent sequence to determine a consensus nucleobase, and the consensus nucleobases determined in successive repetitions are assigned as successive nucleobases in the nascent sequence of the adjacent segment.

10 more elements

A method of differentially treating a patient population, comprising;

sequencing samples from members of the patient population;
wherein for each sample the sequencing comprises
(a) receiving a population of raw target sequencing reads of a nucleic acid target comprising an anchor segment and an adjacent segment;
the anchor segment being of known sequence and the adjacent segment being of unknown sequence;
and at least some of the raw target sequencing reads containing sequencing errors;
(b) evaluating the accuracy of sequencing of the anchor segment in different raw target sequencing reads by comparing raw target sequencing read of the anchor segment with the known sequence of the anchor segment;
(c) assigning a subset of the raw target sequencing reads into an accepted class based on reaching at least a threshold level of accuracy of the sequencing of the anchor segment;
(d) polling nucleobases at a position adjacent the anchor segment sequence in raw target sequencing reads in the accepted class to determine a consensus nucleobase, which consensus nucleobase is assigned as the first nucleobase of a nascent sequence of the adjacent segment;
(e) assigning raw target sequencing reads having the consensus nucleobase determined in the prior polling step to remain in an accepted class and assigning raw target sequencing reads lacking the consensus nucleobase determined in the prior polling step to the rejected class;
(f) optionally reassigning a raw target sequencing read from the rejected class to the accepted class by scoring similarity of the raw target sequencing read to the nascent sequence and reintroducing the raw target sequencing read if the sequence similarity reaches at least a threshold level of similarity;
and
(g) repeating steps (d), (e) and optionally (f), except that a repetition polls a position adjacent the position poled in the previous polling step for raw target sequencing reads having the consensus nucleobase polled in the previous step or in the case of a raw target sequencing read reassigned from the rejected class to the accepted class and not polled in the previous polling step or if polled not having the consensus nucleobase in the previous polling step, the polling polls a position adjacent the position aligned with the last nucleobase of the nascent sequence to determine a consensus nucleobase, and the consensus nucleobases determined in successive repetitions are assigned as successive nucleobases in the nascent sequence of the adjacent segment;
wherein different members of the patient population receive different treatment regimes depending on the determined sequence for the sample from each member.

10 more elements

Algorithms for sequence determinations

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Algorithms for sequence determinations

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Abstract

Claims88