DAR-PCR: a new tool for efficient retrieval of unknown flanking genomic DNA

AMB Express

Table 1 Primers used in this study

Primary PCR			Secondary PCR
ISA primer		SSP1	Inner SSP pair (SSP2/SSP3)
gad-ISA1	TGAAAACTAACCGGCTTAC	gad-1: TCCATACCCTCATCTCCATTTCCAT	gad-2: AACTATCACCCCACAACGTCATCTC gad-3: ACCGTTCATAGGCGAAATTGTTTGT
gad-ISA2	TAAACCTGCGTAAAAACT
gad-ISA3	AACCGGCTTTTTAAACT
hygU-ISA1	CGGGCGTACACAAATCTC	hygU-1: GGCGTATATGCTCCGCATTGGTCTT	hygU-2: CGGCAATTTCGATGATGCAGCTTGG hygU-3: GACCGATGGCTGTGTAGAAGTACTC
hygU-ISA2	GCAATCGTCCGATCCCT
hygU-ISA3	AAATCGCCCGCAGAA
hygD-ISA1^§	CTAAACTCCCCAATGTC	hygD-1: GCCATGTAGTGTATTGACCGATTCC	hygD-2: CAGTTCGGTTTCAGGCAGGT hygD-3: CATATCCACGCCCTCCTACA
hygD-ISA1a^§	CTAAACTCCCCAATGTCT
hygD-ISA1b^§	CTAAACTCCCCAATGTCC
hygD-ISA1c^§	CTAAACTCCCCAATGTCTC
hygD-ISA1d^§	CTAAACTCCCCAATGTCCA
hygD-ISA2	AGTGCCGATAAACATAA

Note: ISA roots are underlined. Buds are unmarked nucleotides located at the 3’ ends of ISA roots. ISA primers paired with SSP1 in the same row were used in the primary PCR; corresponding secondary PCRs were performed using the inner SSP pair (SSP2/SSP3) in the same row. ^§ indicates ISA primers derived from the same ISA root