NON-UNIQUE BARCODES IN A GENOTYPING ASSAY
The present disclosure involves ctDNA assays that interrogate many regions from a single sample with high precision and accuracy, while evaluating multiple forms of cancer-related genomic alterations including sequence mutations and structural alterations. The disclosure provides simplified yet robust methods that achieve high sensitivity and specificity by analyzing cancer genes using a limited pool of non-unique barcodes in combination with endogenous barcodes. Samples are captured and sequenced using high coverage next-generation sequencing to allow tumor-specific somatic mutations, amplifications, and translocations to be identified.
1 - 23 . (canceled)
24 . A method of detecting a gene variant in a sample of nucleic acid comprising generating a library of fragments of the nucleic acid by capturing regions of the nucleic acid using endogenous and exogenous sequences and sequencing genomic positions in the fragments, thereby identifying gene variants in the nucleic acid.
25 . The method of claim 24 , wherein the variants identified are selected from tumor-specific somatic mutations, amplifications, and translocations.
26 . The method of claim 24 , wherein identifying genomic positions of the fragments comprises hybrid capture or whole genome sequencing.
27 . The method of claim 26 , wherein hybrid capture involves a panel of well-characterized cancer genes.
28 . The method of claim 27 , wherein the cancer genes are selected from ABL1, AKT1, ALK, APC, AR, ATM, BCR, BRAF, CDH1, CDK4, CDK6, CDKN2A, CSF1R, CTNNB1, DNMT3A, EGFR, ERBB2, ERBB4, ESR1, EZH2, FBXW7, FGFR1, FGFR2, FGFR3, FLT3, GNA11, GNAQ, GNAS, HNF1A, HRAS, IDH1, IDH2, JAK2, JAK3, KDR, KIT, KRAS, MAP2K1, MET, MLH1, MPL, MYC, NPM1, NRAS, NTRK1, PDGFRA, PDGFRB, PIK3CA, PIK3R1, PTEN, PTPN11, RARA, RB1, RET, ROS1, SMAD4, SMARCB1, SMO, SRC, STK11, TERT, TP53 and/or VHL.
29 . The method of claim 24 , wherein sequencing comprises single-end or paired-end sequencing.
30 . The method of claim 24 , wherein nucleic acid comprises cell-free DNA, circulating tumor DNA, tumor-derived DNA, or RNA.