The nanos gene of Bombyx mori and its expression patterns in developmental embryos and larvae tissues

The nanos gene encodes a zinc-finger protein which is required for the migration and differentiation of primordial germ cells as well as for their fate maintenance. In this study, a 1913 bp nanos gene was cloned and characterized in silkworm (Bombyx mori). RT-PCR and Western blot analysis showed that the nanos was expressed in developing embryos and various silkworm larval tissues. The expression patterns of Nanos and Vasa in silkworm larval gonads were analyzed using immunohistochemistry. It was found that, in silkworm larval ovaries, the Nanos and Vasa proteins were expressed in oocytes. While in testes, high expression of Nanos and Vasa was detected in spermatogonia and relatively weaker expression was found in spermatocytes at latter stages.     

非转座子载体介导的转基因家蚕表达hIL-28A

为了探讨非转座子载体介导转基因家蚕表达外源基因的可能性,将hIL-28A克隆进昆虫细胞表达载体pIZT/V5-His,构建了重组载体pIZT/V5-His-hIL-28A.利用精子介导法将该重组载体导入家蚕卵,通过绿色荧光筛选并结合PCR、DNA杂交等分子鉴定,证实成功获得了转基因家蚕.Western blotting结果显示,转基因家蚕表达重组hIL-28A的分子质量为25 ku,ELISA检测结果显示,hIL-28A在G3代转基因蚕、后部丝腺、脂肪组织冻干粉中的含量分别为0.198、0.320和0.238 ng/g.表明通过非转座子载体介导可以将外源基因导入家蚕基因组并实现外源基因的表达.     

嵌合抗体轻链基因在家蚕细胞中的重组与表达

用家蚕修饰型核多角体病毒为载体,在家蚕细胞获得人鼠嵌合的抗人小细胞肺癌抗体轻链基因的表达。ＰＣＲ和Ｓｏｕｔｈｅｒｎ杂交证明了抗体轻链基因已组建家蚕病毒基因组中。Ｗｅｓｔｅｒｎ ｂｌｏｔ和ＥＬＩＳＡ和分析都检测到在重组病毒感染的家蚕细胞中产生了人鼠嵌合的抗体轻链。     

Bac-to-Bac/BmNPV杆状病毒在家蚕幼虫中表达重组鲎C因子

鲎C因子是鲎血细胞中一种对内毒素具有高亲和力的丝氨酸蛋白酶,被内毒素激活后可水解人工合成的三肽底物,因而能替代传统的鲎试剂用于内毒素检测。通过RT-PCR从东方鲎血细胞中扩增出鲎C因子基因(factor C)序列,利用Bac-to-Bac/BmNPV杆状病毒表达系统在家蚕幼虫中表达该蛋白,并对其进行活性测定。结果显示:被感染的家蚕血清稀释后能检测到Factor C活性,稀释500倍的血清可以用于内毒素检测,且其灵敏度能达到0.2 EU/mL,有望开发新型的、低成本的内毒素检测试剂。