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| Content Provider | PubMed Central |
|---|---|
| Author | Wang, Qiuyan Wu, Huili Wang, Anming Du, Pengfei Pei, Xiaolin Li, Haifeng Yin, Xiaopu Huang, Lifeng Xiong, Xiaolong |
| Abstract | DNA family shuffling is a powerful method for enzyme engineering, which utilizes recombination of naturally occurring functional diversity to accelerate laboratory-directed evolution. However, the use of this technique has been hindered by the scarcity of family genes with the required level of sequence identity in the genome database. We describe here a strategy for collecting metagenomic homologous genes for DNA shuffling from environmental samples by truncated metagenomic gene-specific PCR (TMGS-PCR). Using identified metagenomic gene-specific primers, twenty-three 921-bp truncated lipase gene fragments, which shared 64–99% identity with each other and formed a distinct subfamily of lipases, were retrieved from 60 metagenomic samples. These lipase genes were shuffled, and selected active clones were characterized. The chimeric clones show extensive functional and genetic diversity, as demonstrated by functional characterization and sequence analysis. Our results indicate that homologous sequences of genes captured by TMGS-PCR can be used as suitable genetic material for DNA family shuffling with broad applications in enzyme engineering. |
| Related Links | http://dx.doi.org/10.1074/jbc.M110.139659 |
| Starting Page | 41509 |
| File Format | |
| ISSN | 1083351X |
| e-ISSN | 1083351X |
| Journal | The Journal of Biological Chemistry |
| Issue Number | 53 |
| Volume Number | 285 |
| Language | English |
| Publisher | American Society for Biochemistry and Molecular Biology |
| Publisher Date | 2010-12-31 |
| Access Restriction | Open |
| Rights Holder | American Society for Biochemistry and Molecular Biology |
| Subject Keyword | Research in Higher Education |
| Content Type | Text |
| Resource Type | Article |
| Subject | Cell Biology Biochemistry Molecular Biology |
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