Supplementary file 1: RC-PCR, an innovative and effective method for multiplexing forensically relevant SNP marker systems
Supplementary file 1. The graphical presentation of the RC-PCR technology. The (universal) index primer contains a unique dual index i7/i5 (single indexing also possible), sequence adapter, and universal tail. The RC probe includes an extension blocker with a universal sequence and the reverse complement of the SNP target-specific region (F/R). The indexing and multiplex PCR amplification are performed at the same time in one closed tube. The RC-PCR consists of 2 major steps. 1a – 1c) The universal tail sequences of UIPs hybridize, and the target specific index primers are generated by the Taq polymerase that copies the sequence of the RC probe. In each PCR cycle, new target specific index primers are generated. 2a – 2d) The PCR amplification of SNP-specific amplicons. DNA samples are now tagged with a sample-specific index and Illumina sequence adapter. Now samples can be pooled, purified, and sequenced.