文献类型： 外文期刊
作者： Xiong, Ai-Sheng 1 ; Yao, Quan-Hong 2 ; Peng, Ri-He 2 ; Duan, Hui 2 ; Li, Xian 3 ; Fan, Hui-Qin; Cheng, Zong-Ming; L 1 ;
作者机构： 1.Shanghai Acad Agr Sci, Agro Biotechnol Res Ctr, Shanghai 201106, Peoples R China
2.Shanghai Acad Agr Sci, Agro Biotechnol Res Ctr, Shanghai 201106, Peoples R China; Univ Connecticut, Dept Plant Sci, Storrs, CT 06269 USA; Univ Illinois, Dept Cell & Struct Biol, Urbana, IL 61801 USA; Univ Tennessee, Dept Plant Sci, Knoxville, TN 37996 USA
3.Shanghai Acad Agr Sci, Agro Biotechnol Res Ctr, Shanghai 201106, Peoples R China; Univ Connecticut, Dept Plant Sci, Storrs, CT 06269 USA; Univ Illinois, Dept Cell & Struct
期刊名称：NATURE PROTOCOLS （ 影响因子：13.491； 五年影响因子：17.24 ）
ISSN： 1754-2189
年卷期： 2006 年 1 卷 2 期
页码：
收录情况： SCI
摘要： Here we describe a simple and rapid method for assembly and PCR-based accurate synthesis (PAS) of long DNA sequences. The PAS protocol involves the following five steps: (i) design of the DNA sequence to be synthesized and of 60-bp overlapping oligonucleotides to cover the entire DNA sequence; (ii) purification of the oligonucleotides by PAGE; (iii) first PCR, to synthesize DNA fragments of 400-500 bp in length using 10 inner (template) and two outer (primer) oligonucleotides; (iv) second PCR, to assemble the products of the first PCR into the full-length DNA sequence; and (v) cloning and verification of the synthetic DNA by sequencing and, if needed, error correction using an overlap-extension PCR technique. This method, which takes similar to 1 wk, is suitable for synthesizing diverse types of long DNA molecule. We have successfully synthesized DNA fragments from 0.5 to 12.0 kb, with high G+C content, repetitive sequences or complex secondary structures. The PAS protocol therefore provides a simple, rapid, reliable and relatively inexpensive method for synthesizing long, accurate DNA sequences.
- 相关文献
作者其他论文 更多>>
-
The complete degradation of 1,2-dichloroethane in Escherichia coli by metabolic engineering
作者：Deng, Yong-Dong;Zhang, Wen -Hui;Zuo, Zhi-Hao;Zhang, Hao;Xu, Jing;Gao, Jian- Jie;Wang, Bo;Li, Zhen-Jun;Fu, Xiao-Yan;Wang, Li -Juan;Wang, Yu;Tian, Yong-Sheng;Peng, Ri-He;Yao, Quan-Hong;Deng, Yong-Dong;Zhang, Wen -Hui;Zuo, Zhi-Hao;Zhang, Hao;Xu, Jing;Gao, Jian- Jie;Wang, Bo;Li, Zhen-Jun;Fu, Xiao-Yan;Wang, Li -Juan;Wang, Yu;Tian, Yong-Sheng;Peng, Ri-He;Yao, Quan-Hong
关键词：Bioremediation; Engineered bacteria; Environmental pollution; Metabolic engineering; Synthetic biology
-
Metabolic engineering of Escherichia coli for 2,4-dinitrotoluene degradation
作者：Zhang, Wen-Hui;Deng, Yong-Dong;Zuo, Zhi-Hao;Tian, Yong-Sheng;Xu, Jing;Wang, Bo;Wang, Li-Juan;Han, Hong-Juan;Li, Zhen-Jun;Wang, Yu;Yao, Quan-Hong;Gao, Jian-Jie;Fu, Xiao-Yan;Peng, Ri-He;Zhang, Wen-Hui;Deng, Yong-Dong;Zuo, Zhi-Hao;Tian, Yong-Sheng;Xu, Jing;Wang, Bo;Wang, Li-Juan;Han, Hong-Juan;Li, Zhen-Jun;Wang, Yu;Yao, Quan-Hong;Gao, Jian-Jie;Fu, Xiao-Yan;Peng, Ri-He;Chen, Zhi-Feng
关键词：4-Dinitrotoluene; Re-synthesize gene; Complete degradation; Bioremediation; Escherichia coli
-
Creation of Environmentally Friendly Super "Dinitrotoluene Scavenger" Plants
作者：Gao, Jian-Jie;Li, Zhen-Jun;Wang, Li-Juan;Xu, Jing;Wang, Bo;Fu, Xiao-Yan;Han, Hong-Juan;Zhang, Wen-Hui;Deng, Yong-Dong;Wang, Yu;Zuo, Zhi-Hao;Peng, Ri-He;Tian, Yong-Sheng;Yao, Quan-Hong;Gao, Jian-Jie;Li, Zhen-Jun;Wang, Li-Juan;Xu, Jing;Wang, Bo;Fu, Xiao-Yan;Han, Hong-Juan;Zhang, Wen-Hui;Deng, Yong-Dong;Wang, Yu;Zuo, Zhi-Hao;Peng, Ri-He;Tian, Yong-Sheng;Yao, Quan-Hong;Gao, Jian-Jie;Li, Zhen-Jun;Wang, Li-Juan;Xu, Jing;Wang, Bo;Fu, Xiao-Yan;Han, Hong-Juan;Zhang, Wen-Hui;Deng, Yong-Dong;Wang, Yu;Zuo, Zhi-Hao;Peng, Ri-He;Tian, Yong-Sheng;Yao, Quan-Hong;Zhu, Bo
关键词：2,4-dinitrotoluene (2,4-DNT); carbon neutrality; complete degradation; rapid phytoremediation; sustainable development goals
-
Effect of betanin synthesis on photosynthesis and tyrosine metabolism in transgenic carrot
作者：Wang, Bo;Wang, Ya-Hui;Deng, Yuan-Jie;Xiong, Ai-Sheng;Wang, Bo;Yao, Quan-Hong
关键词：Genetically modified carrot; Betanin; Unintended effects; Photosynthesis; Transcriptome; Metabolomics
-
Identification of Genes of Molecular Marker TGS0892 on Chromosome 6 and Its Mechanism of Soluble Solids Metabolism in Tomato
作者：Zhang, Rong-Rong;Tao, Jian-Ping;Zhang, Jia-Qi;Liu, Hui;Zhu, Wei-Min;Zhao, Tong-Min;Xiong, Ai-Sheng;Song, Liu-Xia;Zhao, Tong-Min;Zhu, Wei-Min
关键词：tomato; soluble solids; molecular marker; TGS0892; identification; gene expression
-
Complete biodegradation of the oldest organic herbicide 2,4-Dichlorophe- noxyacetic acid by engineering Escherichia coli
作者：Wang, Yu;Tian, Yong-Sheng;Gao, Jian-Jie;Xu, Jing;Li, Zhen-Jun;Fu, Xiao-Yan;Han, Hong-Juan;Wang, Li-Juan;Zhang, Wen-Hui;Deng, Yong-Dong;Qian, Cen;Zuo, Zhi-Hao;Wang, Bo;Peng, Ri-He;Yao, Quan-Hong
关键词：24-Dichlorophenoxyacetic acid; Biodegradation; Environmental pollution; Engineered bacteria; Synthetic biology
-
Genetic engineering of complex feed enzymes into barley seed for direct utilization in animal feedstuff
作者：Peng, Ri-He;Zhang, Wen-Hui;Wang, Yu;Deng, Yong-Dong;Wang, Bo;Gao, Jian-Jie;Li, Zhen-Jun;Wang, Li-Juan;Fu, Xiao-Yan;Xu, Jing;Han, Hong-Juan;Tian, Yong-Sheng;Yao, Quan-Hong;Peng, Ri-He;Zhang, Wen-Hui;Wang, Yu;Deng, Yong-Dong;Wang, Bo;Gao, Jian-Jie;Li, Zhen-Jun;Wang, Li-Juan;Fu, Xiao-Yan;Xu, Jing;Han, Hong-Juan;Tian, Yong-Sheng;Yao, Quan-Hong
关键词：recombinant enzymes; feed additive; feedstuff; nutrient utilization
