Compositions and methods for accurately identifying mutations

A method for detecting mutations from exposure to a substance, the method comprising:

• (a) preparing a sequencing library from a sample comprising a plurality of double-stranded DNA molecules from a patient exposed to the substance, wherein preparing the sequencing library comprises ligating cypher polynucleotides to the double-stranded DNA molecules to form double-stranded cypher-target nucleic acid complexes, wherein the cypher polynucleotides comprise identifier tags selected from a plurality of distinct identifier tag sequences;
• (b) sequencing first and second strands of the cypher-target nucleic acid complexes to produce a plurality of first-strand sequencing reads and a plurality of second-strand sequencing reads;
• (c) for each cypher-target nucleic acid complex among a plurality of the cypher-target nucleic acid complexes, comparing the first-strand sequencing reads with the second-strand sequencing reads to identify nucleotides in the first strand that have a corresponding complementary nucleotide in the second strand;
• (d) detecting a mutation from exposure to the substance by analyzing an error-corrected sequence generated from the first strand sequencing reads and second strand sequencing reads for each of the cypher-target nucleic acid complexes among a plurality of the cypher-target nucleic acid complexes;
• and comparing the error-corrected sequences to a reference sequence to identify one or more of a mutation, a genomic distribution of mutations, a mutation frequency, sequence heterogeneity, or DNA damage.

A method of identifying effects of DNA damaging compounds, the method comprising:

• (a) providing a sample comprising a plurality of double-stranded DNA molecules from a patient that has been treated with a compound;
• (b) preparing a sequencing library from the sample by ligating cypher polynucleotides to the double-stranded DNA molecules to form double-stranded cypher-target nucleic acid complexes, wherein the cypher polynucleotides comprise identifier tags selected from a plurality of distinct identifier tag sequences;
• (c) for each cypher-target nucleic acid complex among a plurality of the cypher-target nucleic acid complexes, generating a set of copies of a first strand of the cypher-target nucleic acid complex and a set of distinct yet related copies of a complementary second strand of the cypher-target nucleic acid complex;
• (d) sequencing one or more copies of the first and complementary second strands to produce a plurality of first-strand sequencing reads and a plurality of distinct yet related second-strand sequencing reads;
• (e) for each cypher-target nucleic acid complex among a plurality of the cypher-target nucleic acid complexes, comparing the first-strand sequencing reads with the second-strand sequencing reads to identify nucleotides in the first strand that have a corresponding complementary nucleotide in the second strand;
• (f) comparing an error-corrected sequence generated from the first strand sequencing reads and second strand sequencing reads to a reference sequence to determine one or more of a mutation, a genomic distribution of mutations, a mutation frequency, sequence heterogeneity, or DNA damage;
• and
• (g) based on the comparing step, identifying DNA damage from the compound.