Abstract 1352: Enhanced target capturing throughput and simplified workflow via innovative chemistry on hybrid capture blockers
Authors: Wang Jianpeng , Fan Long , Ye Lumeng , Wang Jiashi , Song Irene , Wu Cedric 
Published: September 2020

Hybrid capture is a popular and well-accepted method to select targeted genomic regions for subsequent next generation sequencing (NGS). Despite its popularity in research and clinical diagnostics, hybrid capture NGS is now facing the challenge by the high throughput demands generated by Novaseq, Illumina's flagship platform that occupies the research and industry market rapidly in recent years. Novaseq create tetrabytes of sequencing data per run, allowing unprecedented level of sample multiplexing, especially for targeted NGS. Conventional blockers, whose sample index-matching region uses either degenerate bases or universal pairing bases, still need to match the length of the sample index for each individual library. When using NovaSeq, scientists need to combine huge number of samples using different types of index system in each single run. However, matching different hybridization blockers with corresponding index system is time consuming and error-prone.

Here we introduce a novel design and chemistry to hybrid capture blockers to maximized the pooling capacity of hybrid capture to match the increasing need of sample throughput and multiplexing. Our new hybrid capture blockers are not only an essential component in the target enrichment workflow to block the permissive interactions between library adapters and target probes, which may lower the capturing efficiency and generate excessive data from the off-target sequence, but also significantly promotes the capturing capacity and efficiency, and simplifies the workflow. Our new blocker, GenFisher Universal Blockers, using our proprietary design and chemistry, create a novel solution to the conjugation between sample indexes on libraries and blockers, enabling blockers bind adapters regardless to the index length and sequence.

In our experiment, NGS libraries are created with a commercially available kit and TruSeq style adapters that contain sample indexes with 3 different lengths: 6nt, 8nt and 10nt. Hybrid captures were then carried out with 8,000-probe GenFisher hybridization panel and kit and Universal Blockers for each individual libraries and the combined library with different indexes. Sequencing data showed that the on-target base percentage was consistently at &lt;75% for all captures for both individual or combined multiplex libraries, whereas hybrid capture without blockers yielded &gt;30% on-target rate. We also compared GenFisher Universal Blockers with the blockers from two other industrial leading providers, which are also claimed to be ‘universal'. Our data showed that GenFisher Universal Blockers was the only truly universal solution that worked with all 3 index lengths unbiasedly while the other two are still index length-sensitive and struggled with 10nt indexes library capturing (on-target ~50%, compared to &lt;65% at 6/8nt). With its versatility, GenFisher Universal Blockers enables higher multiplexing capacity, as well as the freedom of index selection to hybrid capture NGS users.

Citation Format: Jianpeng Wang, Long Fan, Lumeng Ye, Jiashi Wang, Irene Song, Cedric Wu. Enhanced target capturing throughput and simplified workflow via innovative chemistry on hybrid capture blockers [abstract]. In: Proceedings of the Annual Meeting of the American Association for Cancer Research 2020; 2020 Apr 27-28 and Jun 22-24. Philadelphia (PA): AACR; Cancer Res 2020;80(16 Suppl):Abstract nr 1352.