| 引用本文: | 张海波,谭洪新,王兴强,吴嘉敏,秦 松,陈华新,阎斌伦.凡纳滨对虾溶菌酶基因在大肠杆菌中的表达和活性检测[J].海洋科学,2009,33(1):48-53. |
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| 摘要: |
| 将凡纳滨对虾（Litopenaeus vannamei）血淋巴细胞中提取的总RNA，经RT-PCR扩增溶菌酶基因（LvLys基因）的开放阅读框，将其克隆至pMD18-T并测序。用限制性内切酶BamHⅠ和Hind Ⅲ双酶切取目的基因并与表达载体pET-28a(+)连接，构建重组质粒pET-28(a+)/LvLys，转移到BL21(DE3)中诱导表达。经亲和纯化得到凡纳滨对虾重组溶菌酶。抑菌活性检测表明重组溶菌酶对大肠杆菌TOP10有一定抑制作用。 |
| 关键词: 凡纳滨对虾（Litopenaeus vannamei） 溶菌酶基因 原核表达 溶菌活性 |
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| 基金项目:江苏省自然科学基金项目（BK2006548）；江苏省“青蓝工程”项目（QN07008）；国家863计划项目（2006AA100311） |
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| Expression of Litopenaeus vannamei lysozyme gene in Escherichia coli and evaluation of its lytic activity |
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| Abstract: |
| The total RNA extracted from the hemocyte sample of Litopenaeus vannamei（L．Vannamei）was used to amplify the sequence encoding an open reading frame for lysozyme gene（called LvLys gene）by RT-PCR．The sequence was then cloned into pMD18-T vector and was sequenced ．The recombinant plasmid was sequenced and digested by BamHⅠand Hind Ⅲ．The target gene was subsequently connected to the pET-28a (+) vector，which was digested with the corresponding restriction endonuclease．The recombinant plasmid pET-28a(+)/LvLys was transformed into Escherichia coli BL21(DE3) and then induced by isopropylthio-β-D-galactoside (IPTG) and purified through chelating affinity，finally recombinant LvLys protein was analyzed and lytic activity was assayed．The recombinant protein showed a certain antibacterial activity against Escherichia TOP10． |
| Key words: Litopenaeus vannamei lysozyme gene prokaryocyte expression lytic activities |
