July 19, 2019  |  

Preparation of next-generation DNA sequencing libraries from ultra-low amounts of input DNA: Application to single-molecule, real-time (SMRT) sequencing on the Pacific Biosciences RS II.

Authors: Raley, Castle and Munroe, David and Jones, Kristie and Tsai, Yu-Chih and Guo, Yan and Tran, Bao and Gowda, Sujatha and Troyer, Jennifer L and Soppet, Daniel R and Stewart, Claudia and others

We have developed and validated an amplification-free method for generating DNA sequencing libraries from very low amounts of input DNA (500 picograms – 20 nanograms) for single- molecule sequencing on the Pacific Biosciences (PacBio) RS II sequencer. The common challenge of high input requirements for single-molecule sequencing is overcome by using a carrier DNA in conjunction with optimized sequencing preparation conditions and re-use of the MagBead-bound complex. Here we describe how this method can be used to produce sequencing yields comparable to those generated from standard input amounts, but by using 1000-fold less starting material.

Journal: BioRxiv
DOI: 10.1101/003566
Year: 2014

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