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[近平滑假丝酵母β-半乳糖苷酶基因在大肠杆菌细胞中的克隆与表达]

[The cloning and expression of the beta-galactosidase gene of Candida pseudotropicalis yeasts in Escherichia coli cells].

作者信息

Tretiak K A, Zakal'skiĭ A E, Gudz' S P

出版信息

Mikrobiol Z. 1997 Jan-Feb;59(1):11-7.

PMID:9172860
Abstract

The gene encoding the beta-galactosidase of the yeast Candida pseudotropicalis was cloned on YEp13 shuttle vector as the XhoI-fragment of chromosomal DNA of about 9.5 kb. SalGI-fragment of 7.5 kb with the beta-galactosidase gene was subcloned from the pG2 hybrid plasmid obtained into the pBR322 plasmid and then shortened to give 5.2 kb via deletion on XhoI site. This plasmid constructed was designated pBG2-1. In DNA/DNA hybridization studies the appearance of one XhoI-fragment and seven EcoRI-fragments in DNA C. pseudotropicalis hybridized to the DNA cloned fragment and the absence of hybridization to DNA Escherichia coli were shown. The beta-galactosidase activity of cells of transformants of E. coli was lower than the activity of prototrophic strain. The beta-galactosidase biosynthesis in transformants was insignificantly induced by lactose and IPTG and was not repressed by glucose.

摘要

编码假热带念珠菌β-半乳糖苷酶的基因作为约9.5 kb染色体DNA的XhoI片段克隆到YEp13穿梭载体上。将含有β-半乳糖苷酶基因的7.5 kb SalGI片段从获得的pG2杂交质粒亚克隆到pBR322质粒中,然后通过XhoI位点缺失缩短至5.2 kb。构建的该质粒命名为pBG2-1。在DNA/DNA杂交研究中,显示假热带念珠菌DNA中的一个XhoI片段和七个EcoRI片段与克隆的DNA片段杂交,而与大肠杆菌DNA无杂交。大肠杆菌转化体细胞的β-半乳糖苷酶活性低于原养型菌株的活性。转化体中的β-半乳糖苷酶生物合成受乳糖和IPTG诱导不明显,且不受葡萄糖抑制。

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