Safe sequencing system

A method to identify single base substitution, insertion, and deletion mutations in an analyte nucleic acid fragment, comprising:

• attaching a unique identifier sequence (UID) from a pool of UIDs to a first end of each strand of a plurality of analyte DNA fragments using at least two cycles of amplification with first and second primers to form a plurality of uniquely identified analyte DNA fragments, wherein the pool of UIDs are in excess of the analyte DNA fragments during amplification, wherein the first primers comprise:
• a first segment complementary to a desired amplicon;
• a second segment containing the UID;
• and
• a third segment containing a universal priming site for subsequent amplification;
• and wherein the second primers comprise a universal priming site for subsequent amplification;
• wherein each cycle of amplification attaches one universal priming site to a strand;
• amplifying the uniquely identified analyte DNA fragments to form a family of uniquely identified analyte DNA fragments from each uniquely identified analyte DNA fragment;
• and
• determining nucleotide sequences of a plurality of members of the family;
• comparing nucleotide sequences of the family of uniquely identified analyte DNA fragments;
• identifying a nucleotide sequence as accurately representing an analyte DNA fragment when at least 1% of members of the family contain the sequence and the sequence is found in at least two families;
• and
• identifying a single base substitution, insertion, or deletion mutation in the analyte DNA fragment when the nucleotide sequence that accurately represents the analyte DNA fragment is different from a reference sequence by a single base substitution, insertion, or deletion in the analyte DNA fragment.