SLIMamp BRCA1 and BRCA2 library preparation.
A) Example of hg19 positions of 21 overlapping amplicons for BRCA2 exon 11. Oligonucleotides were designed to cover the entire coding regions of BRCA1 and BRCA2 ± 20 bp. The primer regions of each amplicon are represented with thin blocks. All primer binding regions in the exon are covered by their adjacent, overlapping amplicons to ensure 100% coverage of the entire targeted region (~5 Kb in size). The number of amplicons covering each coding exon for BRCA1 and BRCA2 is located in S1 Table. B) Gel electrophoresis of PCR products of two different samples at different library preparation steps. The gene-specific PCR products (lanes 1–4) with the SLIMamp BRCA oligos have an average size of 372 bp (average size of gene-specific region is 331 bp and additional 41 bp for TSCA tags). Bands that represent overlapping amplicons (90–234 bp with tags) are not seen on the gel either before (lanes 1 and 2) or after (lanes 3 and 4) bead purification. Libraries after universal PCR before (lanes 5 and 6) and after purification (lanes 7 and 8) have increased in size due to indexing, which is an additional 94 bp, resulting in an average size of 466 bp. Marker in Lane M is 1Kb Plus ladder.