| Efficient One-Step Fusion PCR Based on Dual-Asymmetric Primers and Two-Step Annealing | |
| Liu, Yilan1; Chen, Jinjin2,3; Thygesen, Anders4 | |
| 2018-02-01 | |
| Source Publication | MOLECULAR BIOTECHNOLOGY
 |
| ISSN | 1073-6085 |
| Volume | 60Issue:2Pages:92-99 |
| Abstract | Gene splicing by fusion PCR is a versatile and widely used methodology, especially in synthetic biology. We here describe a rapid method for splicing two fragments by one-round fusion PCR with a dual-asymmetric primers and two-step annealing (ODT) method. During the process, the asymmetric intermediate fragments were generated in the early stage. Thereafter, they were hybridized in the subsequent cycles to serve as template for the target full-length product. The process parameters such as primer ratio, elongation temperature and cycle numbers were optimized. In addition, the fusion products produced with this method were successfully applied in seamless genome editing. The fusion of two fragments by this method takes less than 0.5 day. The method is expected to facilitate various kinds of complex genetic engineering projects with enhanced efficiency. |
| Keyword | Fusion Pcr Dual-asymmetric Primers Two-step Annealing |
| Subtype | Article |
| WOS Headings | Science & Technology ; Life Sciences & Biomedicine |
| DOI | 10.1007/s12033-017-0050-7 |
| Indexed By | SCI |
| Language | 英语 |
| WOS Keyword | OVERLAP EXTENSION ; GENE ; DNA ; POLYMERASE |
| WOS Research Area | Biochemistry & Molecular Biology ; Biotechnology & Applied Microbiology |
| WOS Subject | Biochemistry & Molecular Biology ; Biotechnology & Applied Microbiology |
| Funding Organization | National Basic Research Program (973 Program)(2011CBA00800) |
| WOS ID | WOS:000424251300002 |
| Citation statistics | |
| Document Type | 期刊论文 |
| Identifier | http://ir.ipe.ac.cn/handle/122111/24097 |
| Collection | 生化工程国家重点实验室 |
| Affiliation | 1.Univ Nebraska Lincoln, Dept Chem & Biomol Engn, Lincoln, NE 68588 USA 2.Chinese Acad Sci, Inst Proc Engn, State Key Lab Biochem Engn, Beijing 100190, Peoples R China 3.Univ Nebraska Lincoln, Dept Food Sci & Technol, Lincoln, NE 68588 USA 4.Tech Univ Denmark, Dept Chem & Biochem Engn, Ctr Bioproc Engn, DK-2800 Lyngby, Denmark |
| Recommended Citation GB/T 7714 | Liu, Yilan,Chen, Jinjin,Thygesen, Anders. Efficient One-Step Fusion PCR Based on Dual-Asymmetric Primers and Two-Step Annealing[J]. MOLECULAR BIOTECHNOLOGY,2018,60(2):92-99. |
| APA | Liu, Yilan,Chen, Jinjin,&Thygesen, Anders.(2018).Efficient One-Step Fusion PCR Based on Dual-Asymmetric Primers and Two-Step Annealing.MOLECULAR BIOTECHNOLOGY,60(2),92-99. |
| MLA | Liu, Yilan,et al."Efficient One-Step Fusion PCR Based on Dual-Asymmetric Primers and Two-Step Annealing".MOLECULAR BIOTECHNOLOGY 60.2(2018):92-99. |
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