Xie Hui, Wang Ya-jing, Tie Chao-nan, Bi Shi-liang, Liu Pei-na
Department of Parasitology, West China School of Preclinical and Forensic Medicine, Sichuan University, Chengdu 610041, China.
Zhongguo Ji Sheng Chong Xue Yu Ji Sheng Chong Bing Za Zhi. 2005 Feb 28;23(1):24-6.
To construct a recombinant plasmid containing ferredoxin gene of Trichomonas vaginalis.
Total DNA was extracted from Trichomonas vaginalis with Chelex-100 method and used as templates for PCR. Primers were designed based on the published sequence of the ferredoxin gene and used to amplify the Trichomonas vaginalis gene using PCR method. The ferredoxin gene obtained by PCR technique was directionally cloned into plasmid pMD-18T simple vector. The constructed recombinant plasmid was transferred into E. coli JM109. The transformants were screened and identified by PCR and restriction analysis. The DNA sequence of the gene was determined by Sanger's method.
The size of amplified ferredoxin gene was 306bp. The correct recombinant plasmid was isolated and confirmed by PCR and restriction analysis. The DNA sequence of cloned gene was the same as the published sequence.
The ferredoxin gene was successfully amplified and cloned into plasmid pMD-18T simple vector. The cloned ferredoxin gene could be used to produce recombinant protein and for study of its function.
构建含阴道毛滴虫铁氧化还原蛋白基因的重组质粒。
采用Chelex-100法从阴道毛滴虫中提取总DNA，并用作PCR模板。根据已发表的铁氧化还原蛋白基因序列设计引物，采用PCR方法扩增阴道毛滴虫基因。通过PCR技术获得的铁氧化还原蛋白基因定向克隆到质粒pMD-18T简单载体中。将构建的重组质粒转入大肠杆菌JM109。通过PCR和限制性分析对转化子进行筛选和鉴定。采用桑格法测定该基因的DNA序列。
扩增的铁氧化还原蛋白基因大小为306bp。通过PCR和限制性分析分离并确认了正确的重组质粒。克隆基因的DNA序列与已发表序列相同。
铁氧化还原蛋白基因成功扩增并克隆到质粒pMD-18T简单载体中。克隆的铁氧化还原蛋白基因可用于生产重组蛋白及其功能研究。
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