Isothermal amplification method for next-generation sequencing.
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.
Genetic Systems, Life Technologies Inc., South San Francisco, CA 94080, USA. zhaochun.ma@lifetech.com
Zhaochun
Z
Ma
Raymond W
RW
Lee
Bin
B
Li
Paul
P
Kenney
Yufang
Y
Wang
Jonathan
J
Erikson
Swati
S
Goyal
Kaiqin
K
Lao
D000818:Animals; D059014:High-Throughput Nucleotide Sequencing / Q000379:methods*; D006801:Humans
DNA breathing; forming colonies; genome; in situ PCR; polony