【作者】 杨小龙； 高志华； 刘敬泽；

【Author】 YANG Xiao-Long, GAO Zhi-Hua, LIU Jing-Ze College of Life Sciences, Hebei Normal University, Shijiazhuang 050016, China

【机构】 河北师范大学生命科学学院；

【摘要】 为阐明蜕皮激素对蜱类唾液腺的调控机制,本文用体外培养、蛋白含量测定和电泳分析方法,研究了蜕皮激素20-羟基蜕皮酮(20-E)对长角血蜱Haemaphysalis longicornis卵黄发生的影响。取刚饱血雌蜱的卵巢, 在TC199培养基中培养,培养基中加入不同剂量的20-E,培养结束后,对卵巢进行蛋白含量测定和电泳分析。结果显示:1.00 ng/μl的20-E能显著提高卵巢蛋白含量,促进卵巢对卵黄原蛋白的摄取;20-E浓度为5.00 ng/μl 时,其作用相反,并且2.50 ng/μl和5.00 ng/μl的20-E均使卵黄蛋白在电泳中的迁移率发生变化;20-E对卵黄蛋白的亚基也有一定影响,使分子量为80 kD和62 kD的亚基在卵巢中的含量增加。上述研究表明,适当剂量的20-E对长角血蜱卵黄发生有作用,能促进卵巢摄取卵黄原蛋白;而高剂量的20-E则表现为毒副作用更多还原

【Abstract】 Synthesis of the major egg yolk protein, vitellogenin (Vg), and Vg uptake by oocytes are essential steps of egg maturation in all arthropods. While the roles of ecdysteroids during vitellogenesis have been well studied in insects, we know much less about the hormonal regulation of vitellogenesis in ticks. In this study, the effects of ecdysteroids on the vitellogenesis of Haemaphysalis longicornis Neumann has been investigated by a culture method in vitro, Bio-Rad protein assay and polyacrylamide gel electrophoresis. The just engorged females were surface sterilized with 70% ethanol. The ovaries were dissected and rinsed several times with phosphate bufftered saline and put directly into the TC199 medium, which contained a gradient of 20-hydroxyecdysone (0, 0.25, 0.50, 1.00, 1.50, 2.00, 2.50 and 5.00 ng/ μl). Two ovaries were incubated by shaking at 27℃ for 48 h in 100 μl of TC199 medium supplemented with egg crude extracts, fetal serum bovine and the compound of penicillin and streptomycin. After incubation, total protein concentration of the ovaries was determined by the Bio-Rad protein assay using bovine serum albumin as the protein standard, the vitellin of ovaries was detected with polyacrylamide gel electrophoresis under non-reducing, non-denaturing condition and reducing condition. The results showed that 1.00 ng/μl dose could enhance the protein concentration and promote the o-vary to absorb vitellogenin, while 5.00 ng/μl dose appeared opposite roles, 2.50 ng/μl and 5.00 ng/μl doses changed the migration rate of the vitellin in polyacrylamide gel electrophoresis and 20-hydroxyecdysone could also affect the polypeptides of the vitellin. The above results suggest that the appropriate dose of 20-hydroxyecdysone plays a role on the vitellogenesis of H .longicornis, the hormone promotes the ovary to absorb vitellogenin, while the high dose has toxico-logical effect更多还原