A Modular Assembly Platform for Rapid Generation of DNA Constructs

被引:46
作者
Akama-Garren, Elliot H. [1 ,2 ]
Joshi, Nikhil S. [1 ]
Tammela, Tuomas [1 ]
Chang, Gregory P. [1 ]
Wagner, Bethany L. [1 ]
Lee, Da-Yae [1 ]
Rideout, William M., III [1 ]
Papagiannakopoulos, Thales [1 ]
Xue, Wen [3 ,4 ]
Jacks, Tyler [1 ,2 ,5 ]
机构
[1] MIT, David H Koch Inst Integrat Canc Res, 77 Massachusetts Ave, Cambridge, MA 02139 USA
[2] MIT, Dept Biol, 77 Massachusetts Ave, Cambridge, MA 02142 USA
[3] Univ Massachusetts, RNA Therapeut Inst, Sch Med, Worcester, MA 01605 USA
[4] Univ Massachusetts, Program Mol Med, Sch Med, Worcester, MA 01605 USA
[5] MIT, Howard Hughes Med Inst, Cambridge, MA 02139 USA
来源
SCIENTIFIC REPORTS | 2016年 / 6卷
关键词
CRISPR-CAS9; CLONING; CELLS;
D O I
10.1038/srep16836
中图分类号
O [数理科学和化学]; P [天文学、地球科学]; Q [生物科学]; N [自然科学总论];
学科分类号
07 ; 0710 ; 09 ;
摘要
Traditional cloning methods have limitations on the number of DNA fragments that can be simultaneously manipulated, which dramatically slows the pace of molecular assembly. Here we describe GMAP, a Gibson assembly-based modular assembly platform consisting of a collection of promoters and genes, which allows for one-step production of DNA constructs. GMAP facilitates rapid assembly of expression and viral constructs using modular genetic components, as well as increasingly complicated genetic tools using contextually relevant genomic elements. Our data demonstrate the applicability of GMAP toward the validation of synthetic promoters, identification of potent RNAi constructs, establishment of inducible lentiviral systems, tumor initiation in genetically engineered mouse models, and gene-targeting for the generation of knock-in mice. GMAP represents a recombinant DNA technology designed for widespread circulation and easy adaptation for other uses, such as synthetic biology, genetic screens, and CRISPR-Cas9.
引用
收藏
页数:9
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