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【目的】构建恶性疟原虫海南 (FCC1/HN)株 p41 3基因真核表达重组质粒pcDNA3 p41 3;测定p41 3基因序列 ,并了解FCC1/HN株与其它分离株 p41 3序列的差异。【方法】根据 p41 3基因已知序列设计合成 2对引物 ,用PCR技术从FCC1/HN株基因组DNA中扩增 p41 3基因 ;将p41 3基因定向克隆入真核表达载体 pcDNA3 ,转化大肠杆菌DH5α感受态细胞 ;用酶切 ,PCR扩增鉴定筛选到的重组质粒阳性克隆。以正确的重组质粒为模板 ,用双脱氧链末端终止法测定 p41 3基因序列 ,应用软件辅助分析p41 3序列及进行同源性比较。【结果】PCR扩增得到特异的FCC1/HN株 p41 3基因 ;正确的 pcD NA3 p41 3重组质粒被筛选和鉴定。测序表明 ,FCC1/HN株p41 3基因大小为 2 137bp ,编码 375个氨基酸。恶性疟原虫FCC1/HN株与FCBR株 p41 3基因核苷酸序列同性为 98.98% ,编码氨基酸序列同源性为 99.73%。【结论】从恶性疟原虫FCC1/HN株基因组DNA中获取p41 3基因 ,成功构建真核表达重组质粒pcDNA3 p41 3,并测定了FCC1/HN株p41 3基因的序列 ;FCC1/HN株与其它分离株 p41 3基因有高度的同源性
【Objective】 To construct eukaryotic expression plasmid pcDNA3 p41 3 of Plasmodium falciparum Hainan (FCC1 / HN) strain p41 3 and to determine the sequence of p41 3 gene and to understand the difference of the sequence of p41 3 between FCC1 / HN strain and other isolates. 【Method】 Two pairs of primers were designed and synthesized based on the known sequence of p41 3 gene. The p41 3 gene was amplified from genomic DNA of FCC1 / HN strain by PCR. The p41 3 gene was cloned into eukaryotic expression vector pcDNA3 and transformed into E. coli DH5α Competent cells; Restriction endonuclease, PCR amplification to identify the selected recombinant plasmid positive clones. The correct recombinant plasmid was used as a template to determine the sequence of p41 3 gene by dideoxy chain termination method. The software was used to analyze p41 3 sequence and compare homology. [Results] The p41 3 gene of FCC1 / HN strain was obtained by PCR amplification. The correct pcD NA3 p41 3 recombinant plasmid was screened and identified. Sequencing showed that the p41 3 gene of FCC1 / HN strain was 2 137 bp in size and encoded 375 amino acids. Plasmodium falciparum FCC1 / HN strain and FCBR strain p41 3 gene nucleotide sequence identity was 98.98%, encoding amino acid sequence homology was 99.73%. 【Conclusion】 The p41 3 gene was obtained from the genomic DNA of Plasmodium falciparum FCC1 / HN strain and the eukaryotic recombinant plasmid pcDNA3 p41 3 was successfully constructed. The sequence of p41 3 gene of FCC1 / HN strain was determined. Strain p41 3 gene has a high degree of homology