【摘要】 目的:研发奶牛抗菌肽生物制剂,防治奶牛子宫内膜疾病。方法:采用酸性尿素聚丙烯酰胺凝胶电泳和琼脂糖弥散法检测子宫内膜组织酸溶性提取物抗菌活性;应用琼脂糖电泳及水平电泳洗脱分离出相应的抗菌条带,HPLC进一步分离纯化,琼脂糖弥散法检测纯化物的抗菌活性,Tricine-SDS-PAGE电泳检测其分子量。结果:粗提物中有两种主要的抗菌成分,其对金黄色葡萄球菌26003、大肠杆菌BL-21、牛致病性大肠杆菌4605、绿脓杆菌10104具有抗菌活性。上层条带的主要色谱峰出现在43、45 min,其中45 min收集的样品有较强的抗菌活性,其相对分子质量约为14 k Da。下层条带主要色谱峰出现在5、35、43、45、46、47、50 min,其中35、43、45 min收集的样品有抗菌活性,其相对分子质量分别约为14-16、6和12 k Da。各个组分对大肠杆菌BL-21和金黄色葡萄球菌26003均具有抑菌活性。结论:从奶牛子宫内膜组织分离出了5个抗菌成分,从相对分子质量、抗菌活性等方面初步认定为抗菌多肽。更多还原

【Abstract】 Objective: To research and develop biological agents of antimicrobial peptides for treatment of endometritis in dairy cows. Methods: The antibacterial activity of acid-soluble extracts from endometrial tissue was detected by acid urea PAGE and agarose diffusion assay. Anti-bacterial bands were isolated from gels with the application of agarose gel electrop-horesis and horizontal electrophoretic elution. Then, the separation and purification were further conducted by HPLC, and antibacterial activity was detected by agarose diffusion assay and molecular weight was determined by Tricine-SDS-PAG.Results: There were two main components with anti-bacterial activity to S.aureus 26003, E. coli BL-21, bovine pathogenic E.coli 4605 and Pseudomonas aeruginosa 10104. The main chromatographic peak of the upper bands appeared at 43 min and 45 min. The sample at 45 min present stronger anti-bacterial activity, and its molecular weight was about 14 k Da. The main chromatographic peak of the lower bands appeared at 5 min, 35 min, 43 min, 45 min, 46 min, 47 min and 50 min, and the samples at 35 min, 43 min and 45 min showed anti-bacterial activity and molecular weight were about 14-16, 6 and 12 k Da,respectively. These components had antibacterial activity on E.coli and Staphylococcus aureus 26003 BL-21. Conclusion:There were five components with antibacterial activities separated from dairy endometrial tissue. Although it also required further study, they were preliminarily considered as antimicrobial peptides according to the evaluation of the molecular weight and antibacterial activities.更多还原