Patent ID: 8728736

Claim:
A method for amplifying a plurality of different target sequences, comprising: amplifying a plurality of different target sequences within a single amplification reaction mixture, wherein the amplifying includes contacting the plurality of different target sequences with a plurality of target-specific primers including a cleavable group and with a polymerase under amplification conditions and producing a plurality of amplified target sequences containing a primer-derived sequence at one end, and wherein at least one of the target sequences of the plurality is amplified using one pair of target-specific primers having minimal cross-hybridization with any other primer in the amplification reaction mixture, wherein the five terminal nucleotides at the 3′ end of each of the target-specific primers of the one pair do not overlap with the five terminal nucleotides at the 3′ end of any other primer in the amplification reaction mixture, and wherein at least one of the target-specific primers and the primer-derived sequence of at least one of the amplified target sequences includes a cleavable group; cleaving the cleavable group of a primer-derived sequence of at least one of the amplified target sequence, thereby producing a cleaved amplified target sequence; and producing at least one adapter-ligated amplified target sequence by ligating at least one double-stranded adapter to an end of at least one cleaved amplified target sequence, wherein the at least one double-stranded adapter is not complementary to the at least one amplified target sequence.