【摘要】 通过检测草鱼禁食后神经肽Y(NPY)与血糖的变化,探讨NPY对草鱼食欲的调控机理,并在体外进行草鱼NPY蛋白的重组表达及纯化,为后期草鱼NPY蛋白的功能研究奠定基础.采用草鱼禁食后取血测定血糖值,利用荧光定量PCR的方法检测草鱼脑中NPY基因的mRNA水平;以pET-32a(+)为载体构建重组质粒,转入大肠杆菌感受态细胞,通过1.0 mmol/L IPTG诱导表达、His-Tag亲和层析柱纯化获得重组蛋白.结果显示草鱼NPY基因的表达量和血糖水平呈负相关;IPTG诱导得到质量浓度为1.43 mg/mL的重组草鱼NPY融合蛋白.草鱼禁食后血糖值下降,NPY表达量增加,说明NPY可能参与草鱼摄食调控.更多还原

【Abstract】 The aim of our research is to detect the variation of grass carp Neuropeptide Y( NPY) and blood glucose after fasting,and further explore the mechanism of NPY on the appetite of grass carp. Expression and purification of NPY protein was carried out in vitro,which provides basis for the research of NPY function in grass carp. In the research,blood glucose value of grass carp was tested after fasting,and the mRNA level of NPY gene in the brain of grass carp was detected with quantitative PCR. In addition,plasmid was recombined with pET-32a( +) vector,and then transfer to the Bacterium coli competent cells,from which recombination protein was obtained through 1. 0mmol / L IPTG inducible expression and His-Tag affinity column purification. The results indicated that the gene expression level of NPY had opposite effect on the blood glucose in grass carp. 1. 43 mg / mL fusion protein of recombinant NPY was obtained with the method of inducible expression of IPTG. The research concluded that the blood glucose value decreased while the NPY expression increase after fasting in the grass carp,which suggested that NPY perhaps had certain effect on the feeding regulation of grass carp.更多还原