Universal short adapters for indexing of polynucleotide samples

A method for sequencing target nucleic acids derived from a plurality of samples, comprising:

• (a) providing a plurality of double-stranded nucleic acid molecules derived from the plurality of samples;
• (b) providing a plurality of transposome complexes, wherein each transposome complex comprises a transposase and two transposon end compositions;
• (c) incubating the double-stranded nucleic acid molecules with the transposome complexes to obtain double-stranded nucleic acid fragments, wherein the double-stranded nucleic acid fragments comprise, at one or both ends, sequences transposed from the transposon end compositions;
• (d) contacting a plurality of index primers with the double-stranded nucleic acid fragments to generate a plurality of index-fragment polynucleotides, wherein
• index primers contacted with double-stranded nucleic acid fragments derived from each sample comprise an index sequence or a combination of index sequences that is uniquely associated with that sample, and
• the index sequence or the combination of index sequences is selected from a set of index sequences;
• (e) pooling the plurality of index-fragment polynucleotides;
• (f) sequencing the pooled index-fragment polynucleotides, thereby obtaining index reads of index sequences and a plurality of target reads of target sequences, each target read being associated with at least one index read;
• and
• (g) using the index reads to determine the target reads' sources of samples.

A computer program product comprising a non-transitory machine readable medium storing program code that, when executed by one or more processors of a computer system, causes the computer system to implement a method for sequencing target nucleic acids derived from a plurality of samples, said program code comprising:

• (a) code for receiving a plurality of index reads and a plurality of target reads of target sequences obtained from target nucleic acids derived from the plurality of samples, wherein
• each target read comprises a target sequence obtained from a target nucleic acid derived from a sample of the plurality of samples,
• each index read comprises an index sequence obtained from a target nucleic acid derived from a sample of the plurality of samples, the index sequence being selected from a set of index sequences,
• each target read is associated with at least one index read,
• each sample of the plurality of samples is uniquely associated with one or more index sequences of the set of index sequences, and
• a Hamming distance between any two index sequences of the set of index sequences is not less than a first criterion value, wherein the first criterion value is at least 2;
• (b) code for identifying, among the plurality of target reads, a subset of target reads associated with index reads aligned to at least one index sequence uniquely associated with a particular sample of the plurality of samples;
• and
• (c) code for determining a target sequence for the particular sample based on the identified subset of target reads.