Inline index helped in cleaning up data contamination generated during library preparation and the subsequent steps

Background High-throughput sequencing involves library preparation and amplification steps, which may induce contamination across samples or between samples and the environment. Methods We tested the effect of applying an inline-index strategy, in which DNA indices of 6 bp were added to both ends of the inserts at the ligation step of library prep for resolving the data contamination problem. Results Our results showed that the contamination ranged from 0.29 to 1.25% in one experiment and from 0.83 to 27.01% in the other. We also found that contamination could be environmental or from reagents besides cross-contamination between samples. Conclusions Inline-index method is a useful experimental design to clean up the data and address the contamination problem which has been plaguing high-throughput sequencing data in many applications.