[Optimization of cloning and expression of beta-glucanase gene from Bacillus amyloliquefaciens].
To compare of performance of beta-1,3-1,4-glucanase gene (bgl) in different expression systems, the beta-1,3-1,4-glucanase gene (GenBank Accession No. EU623974) was amplified from Bacillus amyloliquefaciens BS5582 by PCR and was cloned into three vectors pEGX-4T-1, pET20b(+) and pET28a(+) to construct pEGX-4T-1-bgl, pET20b(+)-bgl and pET28a(+)-bgl recombinant plasmids. The pEGX-4T-1-bgl was transformed into three different Escherichia coli host strains. The pET20b (+)-bgl and pET28a (+)-bgl were transformed into E. coli BL21 (DE3) respectively. Recombinant beta-glucanase was expressed by IPTG inducement in these recombinants. E. coli BL21 (DE3)-pET28a (+)-bgl had the highest enzyme activity. In Luria-Bertani medium, the total enzyme activity was (322.0 +/- 8.8) U/mL, which was 40.1% of original strain in optimal shaking flask condition. This recombinant's performance was studied in Terrific Broth medium under inducement of IPTG and lactose at the same time., and the highest total enzyme activity could reach (1883.3 +/- 45.8) U/mL (818.8% of the original), which indicate that the recombinant strain has a good value in industry application.
Li
Yongxian
Y
Key Laboratory of Industrial Biotechnology, Ministry of Education, Wuxi 214122, China.
Xie
Yan
Y
Zhu
Linjiang
L
Zhang
Yixin
Y
Gu
Guoxian
G
Li
Qi
Q
D001407:Bacillus / Q000201:enzymology* / Q000235:genetics; D003001:Cloning, Molecular; D044644:Endo-1,3(4)-beta-Glucanase / Q000235:genetics* / Q000378:metabolism*; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism; D005822:Genetic Vectors / Q000235:genetics; D008969:Molecular Sequence Data; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism
D011994:Recombinant Proteins; D044644:Endo-1,3(4)-beta-Glucanase