Table 2 Recommendations

- Highly standardized DNA extraction and sample processing protocol

- 30 second denaturation step for both PCRs

- Use different index on forward and reverse primers in PCR1

- Normalize dual indexed samples

- Library pooling after first PCR

- 10 cycles in PCR2

- Final cleanup (1:1 AMPure for size selection)

- Final library pooling of triple indexed samples

250 bp paired end sequencing in HiSeq 2500 Rapid run mode