Detecting mutations and ploidy in chromosomal segments

Abstract

The invention provides methods, systems, and computer readable medium for detecting ploidy of chromosome segments or entire chromosomes, for detecting single nucleotide variants and for detecting both ploidy of chromosome segments and single nucleotide variants. In some aspects, the invention provides methods, systems, and computer readable medium for detecting cancer or a chromosomal abnormality in a gestating fetus.

What is claimed is: 
     
         1 . A method for preparing a sample of a subject having cancer or suspected of having cancer useful for identifying one or more tumor-specific variants in the blood, plasma, serum, or urine sample, the method comprising:
 (a) performing whole exome sequencing or whole genome sequencing on nucleic acids derived from a tumor sample of the subject and identifying a plurality of tumor-specific variants;   (b) selectively enriching or amplifying a plurality of target loci from cell-free DNA derived from a blood, plasma, serum, or urine sample of the subject to obtain selectively enriched or amplified DNA, wherein 10 to 2,000 of the target loci each encompasses a different tumor-specific variant identified in the tumor sample of the subject; and   (c) sequencing the selectively enriched or amplified DNA and obtaining sequence reads with a depth of read of at least 25,000 per target locus, and detecting one or more of the tumor-specific variants present in the cell-free DNA from the sequence reads.   
     
     
         2 . The method of  claim 1 , wherein step (c) comprises sequencing the selectively enriched or amplified DNA and obtaining sequence reads with a depth of read of at least 50,000 per target locus. 
     
     
         3 . The method of  claim 1 , wherein the tumor sample of the subject is from a solid tumor. 
     
     
         4 . The method of  claim 1 , wherein the tumor sample of the subject is a tumor tissue sample. 
     
     
         5 . The method of  claim 1 , wherein the cell-free DNA comprises circulating tumor DNA. 
     
     
         6 . The method of  claim 1 , wherein the tumor-specific variants comprise one or more clonal SNV mutations. 
     
     
         7 . The method of  claim 1 , wherein step (a) further comprises determining clonal heterogeneity of the tumor sample. 
     
     
         8 . The method of  claim 1 , wherein step (b) comprises selectively enriches or amplifies 20 to 200 target loci each encompassing a different tumor-specific variant identified in the tumor sample of the subject. 
     
     
         9 . The method of  claim 1 , wherein step (b) comprises selectively enriches or amplifies 200 to 2,000 target loci each encompassing a different tumor-specific variant identified in the tumor sample of the subject. 
     
     
         10 . The method of  claim 1 , wherein the method further comprises designing PCR primers or hybrid capture probes targeting the tumor-specific variants identified in the tumor sample of the subject, and selectively enriching or amplifying the target loci using the designing PCR primers or hybrid capture probes. 
     
     
         11 . The method of  claim 1 , wherein the method further comprises performing barcoding PCR prior to step (c). 
     
     
         12 . The method of  claim 1 , wherein the method further comprises detecting recurrence and/or metastases of the cancer from the tumor-specific variants detected in the cell-free DNA. 
     
     
         13 . The method of  claim 1 , wherein the subject is a human subject. 
     
     
         14 . The method of  claim 1 , wherein the cancer is colorectal cancer, lung cancer, bladder cancer, or breast cancer. 
     
     
         15 . The method of  claim 1 , wherein the method is capable of detecting a tumor-specific variant at a limit of detection of less than or equal to 0.015%. 
     
     
         16 . A method for preparing a sample of a subject having cancer or suspected of having cancer useful for identifying one or more tumor-specific variants in the blood, plasma, serum, or urine sample, the method comprising:
 (a) performing whole exome sequencing or whole genome sequencing on nucleic acids derived from a tumor sample of the subject and identifying a plurality of tumor-specific variants;   (b) selectively enriching or amplifying a plurality of target loci from cell-free DNA derived from a blood, plasma, serum, or urine sample of the subject to obtain selectively enriched or amplified DNA, wherein 10 to 2,000 of the target loci each encompasses a different tumor-specific variant identified in the tumor sample of the subject; and   (c) sequencing the selectively enriched or amplified DNA and obtaining sequence reads, and detecting one or more of the tumor-specific variants present in the cell-free DNA from the sequence reads, wherein the method is capable of detecting a tumor-specific variant at a limit of detection of less than or equal to 0.05%.   
     
     
         17 . The method of  claim 16 , wherein the method is capable of detecting a tumor-specific variant at a limit of detection of less than or equal to 0.015%. 
     
     
         18 . The method of  claim 16 , wherein the tumor sample of the subject is from a solid tumor. 
     
     
         19 . The method of  claim 16 , wherein the tumor sample of the subject is a tumor tissue sample. 
     
     
         20 . The method of  claim 16 , wherein the cell-free DNA comprises circulating tumor DNA. 
     
     
         21 . The method of  claim 16 , wherein the tumor-specific variants comprise one or more clonal SNV mutations. 
     
     
         22 . The method of  claim 16 , wherein step (a) further comprises determining clonal heterogeneity of the tumor sample. 
     
     
         23 . The method of  claim 16 , wherein step (b) comprises selectively enriches or amplifies 20 to 200 target loci each encompassing a different tumor-specific variant identified in the tumor sample of the subject. 
     
     
         24 . The method of  claim 16 , wherein step (b) comprises selectively enriches or amplifies 200 to 2,000 target loci each encompassing a different tumor-specific variant identified in the tumor sample of the subject. 
     
     
         25 . The method of  claim 16 , wherein the method further comprises designing PCR primers or hybrid capture probes targeting the tumor-specific variants identified in the tumor sample of the subject, and selectively enriching or amplifying the target loci using the designing PCR primers or hybrid capture probes. 
     
     
         26 . The method of  claim 16 , wherein the method further comprises performing barcoding PCR prior to step (c). 
     
     
         27 . The method of  claim 16 , wherein the method further comprises detecting recurrence and/or metastases of the cancer from the tumor-specific variants detected in the cell-free DNA. 
     
     
         28 . The method of  claim 16 , wherein the subject is a human subject. 
     
     
         29 . The method of  claim 16 , wherein the cancer is colorectal cancer, lung cancer, bladder cancer, or breast cancer. 
     
     
         30 . The method of  claim 16 , wherein step (c) comprises sequencing the selectively enriched or amplified DNA and obtaining sequence reads with a depth of read of at least 50,000 per target locus.