丝光绿蝇抗菌肽针刺诱导及初步电泳分离

目的探讨丝光绿蝇抗菌肽的诱导及电泳分离方法。方法丝光绿蝇3龄幼虫针刺体壁诱导后，第48小时提取其血淋巴，设置3、5、7、10μl不同的加样量和pH值60、6．5、7．0的琼脂平板，观察对溶壁微球菌的抑菌效果。置于100、80℃水浴30s、1、3和5min后，观察不同温度处理不同时间后的去杂蛋白效果和对溶壁微球菌的抑菌效果，并对各样品进行电泳分离。结果随加样量增加抑菌环直径增大，10μl环太大易变形，不易测量，以加7μl血淋巴对溶壁微球菌的抑菌效果最佳。平板pH值6．0时，对溶壁微球菌的抑菌环直径最大，随pH值增加而抑菌环直径减小。血淋巴100℃水浴30s、1min仍有抑菌活性，处理3和5min无抑菌活性；80℃水浴30s至5min均有抑菌活性，电泳显示100℃处理3和5min的样品与其他相比缺少2条蛋白区带。结论针刺诱导的丝光绿蝇抗菌肽在酸性条件下抑菌活性更强，分离时血淋巴预处理采用80℃水浴1—3min去杂蛋白效果较好，诱导的血淋巴100℃处理3和5min即丧失抗菌活性，电泳显示缺少2条区带，此2条差异区带可能为抗菌肽组分。

Inducing of antibacterial peptide from Lucilia sericata and preliminary isolation by polyacrylamide gel electrophoresis

Objective To induce and preliminarily isolate antibacterial peptide from Lucilia sericata by polyacrylarnide gel electrophoresis. Methods After the 3^rd instar larva of L. sericata being induced for 48 h by pricking the hemolymph was collected and tested against Micrococcus lysodeikticus at different doses and different pH conditions. The bacteriostatic effects were observed and the diameters of clear inhibitory ring were recorded respectively. After being bathed at 100 ℃ and 80 ℃ for 30 s, 1 min, 3 min and 5min respectively, each hemolymph was centrifugated at 10 000 r/min for 10 rain to get rid of unwanted proteins, the supematant was tested against M. lysodeikticus and displayed by polyacrylamide gel electrophoresis. Results The antibacterial ring diameter of induced hemolymph increased along with the increasing of dose, and decreased along with the increasing of pH. The hemolymph didn＇t possess antibacterial activity against M. lysodeikticus after being bathed at 100 ℃ for 3 min and 5 min, but possessed antibacterial activity after other treatments. It was also found that the two samples which were bathed at 100 ℃ for 3 min and 5 min were lack of two special bands compared with other treatments. Conclusion It implies that the induced antibacterial substance in the blowfly should possexsed stronger antibacterial activity in acid condition. Bathing at 80 ℃ for 1 to 3 rain and then centrifuging can get rid of unwanted proteins in a large extent. Maybe the two special bands are the components of the antibacterial peptides induced in the blowfly.