Open AccessGene Assembly from Chip‐Synthesized Oligonucleotides
Author(s) -
Eroshenko Nikolai,
Kosuri Sriram,
Marblestone Adam H.,
Conway Nicholas,
Church George M.
Publication year - 2012
Publication title -
current protocols in chemical biology
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.503
H-Index - 14
ISSN - 2160-4762
DOI - 10.1002/9780470559277.ch110190
Subject(s) - oligonucleotide , dna , chip , dna synthesis , computational biology , gene , combinatorial chemistry , microbiology and biotechnology , biology , chemistry , nanotechnology , computer science , biochemistry , materials science , telecommunications
De novo synthesis of long double‐stranded DNA constructs has a myriad of applications in biology and biological engineering. However, its widespread adoption has been hindered by high costs. Cost can be significantly reduced by using oligonucleotides synthesized on high‐density DNA chips. However, most methods for using off‐chip DNA for gene synthesis have failed to scale due to the high error rates, low yields, and high chemical complexity of the chip‐synthesized oligonucleotides. We have recently demonstrated that some commercial DNA chip manufacturers have improved error rates, and that the issues of chemical complexity and low yields can be solved by using barcoded primers to accurately and efficiently amplify subpools of oligonucleotides. This unit includes protocols for computationally designing the DNA chip, amplifying the oligonucleotide subpools, and assembling 500‐ to 800‐bp constructs. Curr. Protoc. Chem. Biol . 4:1‐17 © 2012 by John Wiley & Sons, Inc.