CAJ | 학술논문

根据pRSETA载体序列,设计一对引物,采用PCR技术从重组质粒pRSETA-4CCK33上扩增到鸡缩胆囊素33（cholecystokinin-33,CCK33）四串体基因片段（缩写为4CCK33）。再将4CCK33克隆到原核表达载体pBCX上,成功构建重组质粒pBCX-4CCK33并在大肠杆菌BL21中成功表达,SDS-PAGE检测到分子量约为53 000的融合蛋白,可溶分析表明,融合蛋白主要是以可溶形式表达。Western-blot证实,融合蛋白能与CCK-8阳性血清发生免疫反应。用此纯化的融合蛋白制成油乳剂疫苗,主动免疫肉猪。饲养试验表明,免疫0.5 mg/mL和1.5 mg/mL 4CCK33疫苗的试验肉猪平均日增重与空白对照组相比,分别提高了5.28%和5.92%;日采食量则分别提高3.03%和2.53%,差异显著（P〈0.05）。
근거pRSETA재체서렬,설계일대인물,채용PCR기술종중조질립pRSETA-4CCK33상확증도계축담낭소33（cholecystokinin-33,CCK33）사천체기인편단（축사위4CCK33）。재장4CCK33극륭도원핵표체재체pBCX상,성공구건중조질립pBCX-4CCK33병재대장간균BL21중성공표체,SDS-PAGE검측도분자량약위53 000적융합단백,가용분석표명,융합단백주요시이가용형식표체。Western-blot증실,융합단백능여CCK-8양성혈청발생면역반응。용차순화적융합단백제성유유제역묘,주동면역육저。사양시험표명,면역0.5 mg/mL화1.5 mg/mL 4CCK33역묘적시험육저평균일증중여공백대조조상비,분별제고료5.28%화5.92%;일채식량칙분별제고3.03%화2.53%,차이현저（P〈0.05）。
Four repeats of chicken CCK-33（abbr.4CCK33） gene were successfully amplified by PCR from the pRSETA-4CCK33 recombinant plasmid using the primers designed according to sequences of pRSETA plasmid.The four repeats of CCK-33 gene were inserted into plasmid pBCX to obtain recombinant plasmid in which four repeats of CCK33 gene were fused into.The pBCX-4CCK33 was conducted into E.coli BL-21 to induce four repeats of CCK-33 gene fusion expression.The fusion protein band with molecular weight of 53 000 was detected on the SDS-PAGE gel and most fusion proteins were soluble.Western-blot showed that the expression products can specifically reacted with the antisera of CCK-8.The soluble form was purified.The purified fusion proteins were used as antigen to prepare oil-emulsion vaccines,and the pigs were immunized.Average daily gain in pigs immunized with 0.5 mg/mL and 1.5 mg/mL 4CCK33 vaccine were significantly increased by 5.28% and 5.92% respectively,while that of the feed intake increased by 3.03% and 2.53%,compared with the control group.