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Cloning and Expression of the Cellulase Gene from the King Oyster Mushroom, Pleurotus eryngii. |
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| รหัสดีโอไอ | |
| Creator | 1. Urarux Romruen 2. Eakaphun Bangyeekhun |
| Title | Cloning and Expression of the Cellulase Gene from the King Oyster Mushroom, Pleurotus eryngii. |
| Publisher | Silpakorn University Research and Development Institute |
| Publication Year | 2559 |
| Journal Title | Silpakorn University Science and Technology Journal |
| Journal Vol. | 10 |
| Journal No. | 2 |
| Page no. | 22-30 |
| Keyword | Pleurotus eryngii, Cellobiohydrolase, Glycosyl hydrolase family 7, Protein expression, Cloning |
| ISSN | 1905-9159 |
| Abstract | A gene encoding for cellobiohydrolase (PEcbh) from P. eryngii was cloned by using RT-PCR 3' and 5' RACE techniques. The result showed that the PEcbh was 1377 bp nucleotide sequence encoded for 459-deduced amino acid. Analysis of predicted protein revealed that PEcbh consisted of a glycosyl hydrolase family 7 domain but lacked of cellulose binding domain, a calculated molecular weight of 49.3 kDa and a pI of 5.3. The PEcbh was cloned into pET28a (+) to obtain a recombinant pET/PEcbh and expressed in E. coli BL21 (DE3). The optimal conditions of PEcbh expression were 0.2 mM IPTG, 1 h induction time at 180 C and 4 h post-induction time. The CMCase activity could be detected, but at a low activity. This is probably due to a lack of the cellulose binding domain in PEcbh. Expression of PEcbh in E. coli BL21 (DE3) and Rosetta (DE3) were compared and the results indicated that CMCase activity in Rosetta (DE3) was higher than in BL21 about 2 times. |
