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Hsp70基因的表达量在研究中常作为一个重要的生理指标来反映机体的状况。为了研究象耳豆根结线虫Hsp70基因相关功能,本研究前期成功构建了p EASY-E1-Me Hsp70和p ET30a-Me Hsp70两个原核表达载体,通过热激转化将其分别转入大肠杆菌(Escherichia coli)BL21进行表达。对转入Me Hsp70基因的大场杆菌BL21、转入空载的大肠杆菌BL21和大肠杆菌BL21原始菌种在55℃、65℃进行热稳定性测定,在37℃、200 r/min条件下进行生长曲线测定。热稳定性试验结果表明,转入p ET30a-Me Hsp70基因的大场杆菌生存时间较长,而转入p EASY-E1-Me Hsp70的菌株、转入空载的菌株和原始菌株热稳定性都相对较差。通过荧光定量PCR测定发现Me Hsp70基因均有上调表达,转入p ET30a-Me Hsp70基因的大场杆菌Hsp70基因水平高于其他菌株。生长曲线测定试验结果表明,重组菌株与原始菌株生长曲线基本一致,转入p ET30a-Me Hsp70载体的菌株调整期滞留时间长于其他菌株。该项研究表明Me Hsp70基因能够提升大肠杆菌BL21的抗应激能力和影响大肠杆菌BL21的生长,为进一步开展Me Hsp70基因在象耳豆根结线虫生长过程的功能研究提供了一定的理论基础。
Hsp70 gene expression in the study often as an important physiological indicators to reflect the condition of the body. In order to study the Hsp70 gene-related functions of root-knot nematode, two prokaryotic expression vectors, pEASY-E1-Me Hsp70 and p ET30a-Me Hsp70, were successfully constructed and transferred into Escherichia coli Escherichia coli BL21. The E. coli BL21 transformed into Me Hsp70 gene and untransfected E. coli BL21 and E. coli BL21 were tested for their thermostability at 55 ° C and 65 ° C and at 37 ° C and 200 r / min Growth curve determination. The results of the thermostability test showed that the large-field bacillus transformed into the p ET30a-Me Hsp70 gene had a longer survival time, and the strain transformed to pEASY-E1-Me Hsp70 was transferred into the empty strain and the original strain was both thermostable Relatively poor. Me Hsp70 gene was up-regulated by fluorescence quantitative PCR assay, and the Hsp70 gene level was higher in Pseudomonas aeruginosa transferred to p ET30a-Me Hsp70 gene than other strains. The results of the growth curve assay showed that the growth curves of the recombinant strain and the original strain were basically the same, and the strains transferred to the p ET30a-Me Hsp70 carrier had longer residence time than the other strains. This study shows that Me Hsp70 gene can enhance the anti-stress ability of Escherichia coli BL21 and affect the growth of Escherichia coli BL21, which provides a theoretical basis for further study on the function of Me Hsp70 gene in the growth of Meloidogyne incognita.