Zhaochun Ma

Isothermal amplification method for next-generation sequencing

Significance We have developed a cost-effective in situ isothermal amplification method for next-generation sequencing (NGS) that can generate billions of monoclonal colonies in less than 30 minutes. This method can be used on NGS platforms to make the

Unnatural polymerase substrates that can sustain enzymatic synthesis of double stranded nucleic acids from a nucleic acid template and methods of use

1,000 genome possible and simplify the workflow for clinical genomic applications. We report an approach for generating immobilized monoclonal templates for next- generation sequencing applications. Our isothermal amplification method is based on a template walking mechanism using a pair of low-melting temperature (Tm) solid-surface homopolymer primers and a low-Tm solution phase primer. The method can generate more than one billion submicrometer-sized colonies in a single lane of a next-generation sequencing flowchip. An alternative paired-end sequencing method using interstrand DNA photo cross-linking to covalently link the complementary strands of the original templates to the solid surface is also demonstrated.