Patent ID: 11965157
Assignee: SINGLERA GENOMICS, INC.
Field: Biotechnology (Chemistry)
Classification: CPC C | IPC C

Claim 0:
1. A method, comprising ligating a set of adaptors to a library of single-stranded polynucleotides, wherein:
each single-stranded polynucleotide in the library of single-stranded polynucleotides comprises a target sequence;
the ligation is catalyzed by a single-stranded DNA (ssDNA) ligase;
each single-stranded polynucleotide is blocked at the 5′ end to prevent ligation at the 5′ end;
each adaptor comprises a unique molecular identifier (UMI) sequence that earmarks the single-stranded polynucleotide to which the adaptor is ligated, and is blocked at the 3′ end to prevent ligation at the 3′ end; and
the 5′ end of the adaptor is ligated to the 3′ end of the single-stranded polynucleotide by the ssDNA ligase to form a linear ligation product,
thereby obtaining a library of linear, single-stranded ligation products, wherein each linear, single-stranded ligation product comprises a single adaptor;
further comprising converting the library of linear, single-stranded ligation products into a library of linear, double-stranded ligation products;
further comprising amplifying the library of linear, double-stranded ligation products by a polymerase chain reaction (PCR), using a set of primers each comprising a sequence that is reverse-complement to the adaptor and/or hybridizable to the adaptor, and a primer hybridizable to the target sequence, thereby obtaining an amplified library of linear, double-stranded ligation products comprising sequence information of the target sequence; and
further comprising sequencing the amplified library of linear, double-stranded ligation products, wherein the conversion rate of the sequencing (percentage of single-stranded polynucleotides in the library that gives rise to sequencing reads) is at least 60%.