Unincorporated nucleotides and primers were treated by mixing 4 μl of the final PCR reaction mixture with 1 μl ExoSAP-IT (USB Corp., Cleveland, Ohio) followed by incubation at 37°C for 30 min and 80°C for 15 min. For sequencing, 1 μl ExoSAP-IT-treated DNA was added to 11 μl 8 pM primer C or D. Sequencing was performed at GENEWIZ, Inc. (South Plainfield, NJ) on an ABI 3730 XL sequencer using the ABI 3730POP7SR basecaller (Applied Biosystems).
