Systems and methods for determining structural variation and phasing using variant call data

Abstract

Systems and methods for determining structural variation and phasing using variant call data obtained from nucleic acid of a biological sample are provided. Sequence reads are obtained, each comprising a portion corresponding to a subset of the test nucleic acid and a portion encoding a barcode independent of the sequencing data. Bin information is obtained. Each bin represents a different portion of the sample nucleic acid. Each bin corresponds to a set of sequence reads in a plurality of sets of sequence reads formed from the sequence reads such that each sequence read in a respective set of sequence reads corresponds to a subset of the nucleic acid represented by the bin corresponding to the respective set. Binomial tests identify bin pairs having more sequence reads with the same barcode in common than expected by chance. Probabilistic models determine structural variation likelihood from the sequence reads of these bin pairs.

1 - 53 . (canceled) 
     
     
         54 . A method of phasing sequencing data of a test nucleic acid sample obtained from a biological sample from a single organism of a species, wherein the test nucleic acid sample comprises a first set of haplotypes (H O ) and a second set of haplotypes (H 1 ), the method comprising:
 at a computer system having one or more processors, and memory storing one or more programs for execution by the one or more processors:   (A) obtaining a plurality of variant calls A i,p  for the test nucleic acid sample, wherein
 i is an index to a position in a reference consensus sequence for all or a portion of a genome of the species, and 
 p∈{0, 1} in which label 0 assigns a respective variant call in A i,p  to H=0 and label 1 assigns the respective variant call to H=1; 
   (B) for each respective local block of variant calls in A i,p  that are localized to a corresponding subset of the reference consensus sequence, using a beam search over the haplotype assignments of local phasing vectors X k , X k+1 , . . . , X k+j  in the respective local block of variant calls, wherein
 k is the first variant in the respective local block of variant calls, 
 j is a number of variant calls in the respective local block of variant calls, 
 assignments of X k , X k+1 , . . . , X k+j  are found by computing an objective function in which the phasing vector of the objective function in respective computations is limited to X k , X k+1 , . . . , X k+j , and 
 the objective function is calculated by matching observed sequence reads of the test nucleic acid sample against the respective local block of variant calls in A i;p , thereby finding a phasing solution for each respective local block of variant calls in A i;p ; and 
   (C) greedily joining, upon completion of the beam search for each respective local block of variant calls in A i;p , neighboring local blocks of variant calls in A i;p  using the phasing solution for each respective local block of variant calls thereby obtaining a phasing configuration {circumflex over (X)} for the single organism of the species comprising a plurality of contigs having an N50 in excess of 10 kbp.   
     
     
         55 . A method of phasing sequencing data of a test nucleic acid sample obtained from a biological sample from a single organism of a species while accounting for error in variant call zygosity, wherein the test nucleic acid sample comprises a first set of haplotypes (H 0 ) and a second set of haplotypes (H 1 ), the method comprising:
 at a computer system having one or more processors, and memory storing one or more programs for execution by the one or more processors:   (A) obtaining a plurality of variant calls A i,p , wherein
 i is an index to a position in a reference consensus sequence for all or a portion of a genome of the species, and 
 p∈{0, 1, −1} in which label 0 assigns a respective variant call in A i,p  to H 0 , label 1 assigns the respective variant call to H 1 , and label −1 assigns the respective variant call to a zygosity error condition 
   (B) for each respective local block of variant calls in A i,p  that are localized to a corresponding subset of the reference consensus sequence, using a beam search over the haplotype assignments of local phasing vectors X k , X k+1 , . . . , X k+j , in the respective local block of variant calls, wherein
 k is the first variant in the respective local block of variant calls, 
 j is a number of variant calls in the respective local block of variant calls, 
 assignments of X k , X k+1 , . . . , X k+j  are found by computing an objective function in which the phasing vector of the objective function in respective computations is limited to X k , X k+1 , . . . , X k+j , and 
 the objective function is calculated by matching observed sequence reads of the test nucleic acid sample against the respective local block of variant calls in A i;p , thereby finding a phasing solution for each respective local block of variant calls in A i;p ; and 
   (C) greedily joining, upon completion of the beam search for each respective local block of variant calls in A i;p , neighboring local blocks of variant calls in A i;p  using the phasing solution for each respective local block of variant calls thereby obtaining a phasing configuration {circumflex over (X)} for the single organism of the species comprising a plurality of contigs having an N50 in excess of 10 kbp.   
     
     
         56 . The method of claim  225 , the method further comprising iteratively swapping the phase result of individual x i  in {circumflex over (X)} and recomputing the objective function, thereby obtaining {circumflex over (X)}. 
     
     
         57 . The method of claim  226 , the method further comprising iteratively swapping the phase result of individual x i  in {circumflex over (X)} and recomputing the objective function, thereby obtaining {circumflex over (X)}. 
     
     
         58 . The method of claim  225 , wherein a respective local block of variant calls consists of between 20 and 60 variants in A i;p . 
     
     
         59 . The method of claim  226 , wherein a respective local block of variant calls consists of between 20 and 60 variants in A i;p . 
     
     
         60 . The method of claim  225 , wherein an iteration of the beam search for the assignments of one of X k , X k+1 , . . . , X k+j  discards all but a predetermined number of solutions for {circumflex over (X)}. 
     
     
         61 . The method of claim  226 , wherein an iteration of the beam search for the assignments of one of X k , X k+1 , . . . , X k+j  discards all but a predetermined number of solutions for {circumflex over (X)}. 
     
     
         62 . The method of claim  225 , wherein the test nucleic acid sample is loaded onto a plurality of barcoded-oligo coated gel-beads from which a plurality of sequence reads is obtained in order to derive the plurality of variant calls A i;p  and wherein the test nucleic acid sample is 50 ng or less. 
     
     
         63 . The method of  claim 62 , wherein the plurality of barcoded-oligo coated gel-beads comprises 10,000 beads. 
     
     
         64 . The method of  claim 62 , wherein the test nucleic acid sample is 2.5 ng or less. 
     
     
         65 . The method of  claim 62 , wherein the plurality of sequencing reads is obtained within ten minutes of exposure to the plurality of barcodes. 
     
     
         66 . The method of claim  226 , wherein the test nucleic acid sample is loaded onto a plurality of barcoded-oligo coated gel-beads from which a plurality of sequence reads is obtained in order to derive the plurality of variant calls A i;p  and wherein the test nucleic acid sample is 50 ng or less. 
     
     
         67 . The method of  claim 66 , wherein the plurality of barcoded-oligo coated gel-beads comprises 10,000 beads. 
     
     
         68 . The method of  claim 66 , wherein the test nucleic acid sample is 2.5 ng or less. 
     
     
         69 . The method of  claim 66 , wherein the plurality of sequencing reads is obtained within ten minutes of exposure to the plurality of barcodes. 
     
     
         70 . A computing system, comprising:
 one or more processors;   memory storing one or more programs to be executed by the one or more processors;   the one or more programs comprising instructions for phasing sequencing data of a test nucleic acid sample obtained from a biological sample from a single organism of a species, wherein the test nucleic acid sample comprises a first set of haplotypes (H 0 ) and a second set of haplotypes (H 1 ) by executing a method comprising:   (A) obtaining a plurality of variant calls A i;p , wherein
 i is an index to a position in a reference consensus sequence for all or a portion of a genome of the species, and 
 p∈{0, 1, −1} in which label 0 assigns a respective variant call in A i;p  to H 0 , label 1 assigns the respective variant call to H 1 , and label −1 assigns the respective variant call to a zygosity error condition H −1 ; 
   (B) for each respective local block of variant calls in A i,p  that are localized to a corresponding subset of the reference consensus sequence, using a beam search over the haplotype assignments of local phasing vectors X k , X k+1 , . . . , X k+j  in the respective local block of variant calls, wherein
 k is the first variant in the respective local block of variant calls, 
 j is a number of variant calls in the respective local block of variant calls, 
 assignments of X k , X k+1 , . . . , X k+j  are found by computing an objective function in which the phasing vector of the objective function in respective computations is limited to X k , X k+1 , . . . , X k+j , and 
 the objective function is calculated by matching observed sequence reads of the test nucleic acid sample against the respective local block of variant calls in A i;p , 
   thereby finding a phasing solution for each respective local block of variant calls in A i;p ; and   (C) greedily joining, upon completion of the beam search for each respective local block of variant calls in A i;p , neighboring local blocks of variant calls in A i;p  using the phasing solution for each respective local block of variant calls thereby obtaining a phasing configuration {circumflex over (X)} for the single organism of the species comprising a plurality of contigs having an N50 in excess of 10 kbp.   
     
     
         71 . A computing system, comprising:
 one or more processors;   memory storing one or more programs to be executed by the one or more processors;   the one or more programs comprising instructions for phasing sequencing data of a test nucleic acid sample obtained from a biological sample from a single organism of a species while accounting for error in variant call zygosity, wherein the test nucleic acid sample comprises a first set of haplotypes (H 0 ) and a second set of haplotypes (H 1 ), the one or more programs executing a method comprising:   (A) obtaining a plurality of variant calls A i;p , wherein
 i is an index to a position in a reference consensus sequence for all or a portion of a genome of the species, and 
 p∈{0, 1} in which label 0 assigns a respective variant call in A i,p  to H O  and label 1 assigns the respective variant call to H 1 ; 
   (B) for each respective local block of variant calls in A i,p  that are localized to a corresponding subset of the reference consensus sequence, using a beam search over the haplotype assignments of local phasing vectors X k , X k+1 , . . . X k+j  in the respective local block of variant calls, wherein
 k is the first variant in the respective local block of variant calls, 
 j is a number of variant calls in the respective local block of variant calls, 
   assignments of X k , X k+1 , . . . , X k+j  are found by computing an objective function in which the phasing vector of the objective function in respective computations is limited to X k , X k+1 , . . . , X k+j , and
 the objective function is calculated by matching observed sequence reads of the test nucleic acid sample against the respective local block of variant calls in A i;p , thereby finding a phasing solution for each respective local block of variant calls in A i;p ; and 
   (C) greedily joining, upon completion of the beam search for each respective local block of variant calls in A i;p , neighboring local blocks of variant calls in A i;p  using the phasing solution for each respective local block of variant calls thereby obtaining a phasing configuration {circumflex over (X)} for the single organism of the species comprising a plurality of contigs having an N50 in excess of 10 kbp.   
     
     
         72 . A non-transitory computer readable storage medium storing one or more programs configured for execution by a computer, the one or more programs comprising instructions for phasing sequencing data of a test nucleic acid sample obtained from a biological sample from a single organism of a species, wherein the test nucleic acid sample comprises a first set of haplotypes (H 0 ) and a second set of haplotypes (H 1 ), the one or more programs collectively executing a method comprising:
 A) obtaining a plurality of variant calls A i;p , wherein
 i is an index to a position in a reference consensus sequence for all or a portion of a genome of the species, and 
 p∈{0, 1} in which label 0 assigns a respective variant call in A i,p  to H O  and label 1 assigns the respective variant call to H 1 ; 
   (B) for each respective local block of variant calls in A i,p  that are localized to a corresponding subset of the reference consensus sequence, using a beam search over the haplotype assignments of local phasing vectors X k , X k+1 , . . . , X k+j  in the respective local block of variant calls, wherein
 k is the first variant in the respective local block of variant calls, 
 j is a number of variant calls in the respective local block of variant calls, 
 assignments of X k , X k+1 , . . . , X k+j  are found by computing an objective function in which the phasing vector of the objective function in respective computations is limited to X k , X k+1 , . . . , X k+j , and 
 the objective function is calculated by matching observed sequence reads of the test nucleic acid sample against the respective local block of variant calls in A i;p , 
   thereby finding a phasing solution for each respective local block of variant calls in A i;p ; and   (C) greedily joining, upon completion of the beam search for each respective local block of variant calls in A i;p , neighboring local blocks of variant calls in A i;p  using the phasing solution for each respective local block of variant calls thereby obtaining a phasing configuration {circumflex over (X)} for the single organism of the species comprising a plurality of contigs having an N50 in excess of 10 kbp.   
     
     
         73 . A non-transitory computer readable storage medium storing one or more programs configured for execution by a computer, the one or more programs comprising instructions for phasing sequencing data of a test nucleic acid sample obtained from a biological sample from a single organism of a species while accounting for error in variant call zygosity, wherein the test nucleic acid sample comprises a first set of haplotypes (H 0 ) and a second set of haplotypes (H 1 ), the one or more programs collectively executing a method comprising:
 (A) obtaining a plurality of variant calls A i,p , wherein
 i is an index to a position in a reference consensus sequence for all or a portion of a genome of the species, and 
 p∈{0, 1} in which label 0 assigns a respective variant call in A i,p  to H=0 and label 1 assigns the respective variant call to H=1; 
   (B) for each respective local block of variant calls in A i,p  that are localized to a corresponding subset of the reference consensus sequence, using a beam search over the haplotype assignments of local phasing vectors X k , X k+1 , . . . , X k+j  in the respective local block of variant calls, wherein
 k is the first variant in the respective local block of variant calls, 
 j is a number of variant calls in the respective local block of variant calls, assignments of X k , X k+1 , . . . , X k+j  are found by computing an objective function in which the phasing vector of the objective function in respective computations is limited to X k , X k+1 , . . . , X k+j , and 
 the objective function is calculated by matching observed sequence reads of the test nucleic acid sample against the respective local block of variant calls in A i;p , thereby finding a phasing solution for each respective local block of variant calls in A i;p ; and 
   (C) greedily joining, upon completion of the beam search for each respective local block of variant calls in A i;p , neighboring local blocks of variant calls in A i;p  using the phasing solution for each respective local block of variant calls thereby obtaining a phasing configuration {circumflex over (X)} for the single organism of the species comprising a plurality of contigs having an N50 in excess of 10 kbp.