Journal article

Hi-Plex2: a simple and robust approach to targeted sequencing-based genetic screening

Fleur Hammet, Khalid Mahmood, Thomas R Green, Nguyen-Dumont Tu, Melissa C Southey, Daniel D Buchanan, Andrew Lonie, Katherine L Nathanson, Fergus J Couch, Bernard J Pope, Daniel J Park

BioTechniques | FUTURE SCI LTD | Published : 2019

Abstract

We have previously reported Hi-Plex, a multiplex PCR methodology for building targeted DNA sequencing libraries that offers a low-cost protocol compatible with high-throughput processing. Here, we detail an improved protocol, Hi-Plex2, that more effectively enables the robust construction of small-to-medium panel-size libraries while maintaining low cost, simplicity and accuracy benefits of the Hi-Plex platform. Hi-Plex2 was applied to three panels, comprising 291, 740 and 1193 amplicons, targeting genes associated with risk for breast and/or colon cancer. We show substantial reduction of off-target amplification to enable library construction for small-to-medium-sized design panels not poss..

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Grants

Awarded by Australian National Health and Medical Research Council (NHMRC)


Awarded by national Breast Cancer Foundation (NBCF)


Funding Acknowledgements

This work was supported by the Australian National Health and Medical Research Council (NHMRC) grants GNT1108179, GNT1074383 and GNT1125269, and national Breast Cancer Foundation (NBCF) grant NT-14-016. DD Buchanan is a NHMRC RD Wright Career Development Fellow, TN-D is a National Breast Cancer Foundation (Australia) Career Development Fellow and BJP is a Victorian Health and Medical Research Fellow. MC Southey is a NHMRC Senior Research Fellow (GNT1061177). The authors have no other relevant affiliations or financial involvement with any organization or entity with a financial interest in or financial conflict with the subject matter or materials discussed in the manuscript apart from those disclosed.