論文 - 詳細
| RRC ID | 50402 |
|---|---|
| 著者 | Yamashita M, Xu J, Morokuma D, Hirata K, Hino M, Mon H, Takahashi M, Hamdan SM, Sakashita K, Iiyama K, Banno Y, Kusakabe T, Lee JM. |
| タイトル | Characterization of Recombinant Thermococcus kodakaraensis (KOD) DNA Polymerases Produced Using Silkworm-Baculovirus Expression Vector System. |
| ジャーナル | Mol Biotechnol |
| Abstract |
The KOD DNA polymerase from Thermococcus kodakarensis (Tkod-Pol) has been preferred for PCR due to its rapid elongation rate, extreme thermostability and outstanding fidelity. Here in this study, we utilized silkworm-baculovirus expression vector system (silkworm-BEVS) to express the recombinant Tkod-Pol (rKOD) with N-terminal (rKOD-N) or C-terminal (rKOD-C) tandem fusion tags. By using BEVS, we produced functional rKODs with satisfactory yields, about 1.1 mg/larva for rKOD-N and 0.25 mg/larva for rKOD-C, respectively. Interestingly, we found that rKOD-C shows higher thermostability at 95 °C than that of rKOD-N, while that rKOD-N is significantly unstable after exposing to long period of heat-shock. We also assessed the polymerase activity as well as the fidelity of purified rKODs under various conditions. Compared with commercially available rKOD, which is expressed in E. coli expression system, rKOD-C exhibited almost the same PCR performance as the commercial rKOD did, while rKOD-N did lower performance. Taken together, our results suggested that silkworm-BEVS can be used to express and purify efficient rKOD in a commercial way. |
| 巻・号 | 59(6) |
| ページ | 221-233 |
| 公開日 | 2017-6-1 |
| DOI | 10.1007/s12033-017-0008-9 |
| PII | 10.1007/s12033-017-0008-9 |
| PMID | 28484957 |
| MeSH | Animals Bombyx / genetics* Genetic Vectors / genetics* Recombinant Proteins / genetics* Thermococcus / genetics* |
| IF | 2.022 |
| 引用数 | 8 |
| オルトメトリクス指標 |
オルトメトリクス指標項目
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| 最多言及媒体 | Patent(IFI CLAIMS) |
| 各媒体での言及数の合計 | 6 |
| 過去6か月間でのオルトメトリクス指標の変動値 | 3.0 |
| リソース情報 | |
| カイコ | n70 |