中文摘要：

为了制备可用于拟蜘蛛拖丝蛋白体外检测的抗血清,利用前期工作中获得的蜘蛛拖丝蛋白基因重组原核表达载体2S-pET-52b（＋）,采用原核表达方法获得大量重组蜘蛛拖丝蛋白2S-His,对此重组蛋白进行His标签特异性的亲和纯化,再依次经SDS-PAGE、切胶和与佐剂混合后作为抗原;皮下多点注射法将抗原注入新西兰大白兔皮下进行免疫;采集已免疫的兔血清,采用ELISA检测方法对其效价进行检测。结果显示,重组质粒2S-pET-52b（＋）转化BL21-pLysS后获得最佳的表达菌株;在筛选得到的最佳IPTG、温度和时间（分别为1.0 mmol/L、30℃和2 h）条件下诱导获得2S-His重组蛋白;经免疫的4只兔血清中,2只兔的血清效价达1∶25 600和1∶12 800。表明利用2S-pET-52b（＋）重组质粒成功获得可用于拟蜘蛛拖丝蛋白体外检测的抗血清。本研究为加快建立在被毛中特异表达大量蜘蛛拖丝蛋白的新方法提供依据。

英文摘要：

In order to prepare anti-serum to spider dragline silk protein for in vitro detection,a recombinant prokaryotic expression plasmid 2S-pET-52b（＋） was constructed and then expressed in BL21-pLysS cells for producing recombinant protein which was used as antigen after purification by the means of affinity choromatography.The results were showed as followed： recombinant plasmid 2S-pET-52b（＋） was transformed into BL21-pLysS cells,and then selected the most suitable strain and expression conditions including concentration,temperature and time of IPTG induction（1.0 mmol/L,30℃ and 2 h）.Recombinant protein was purified by the means of His-tag-specific affinity choromatography after expressed.Polyclonal antibody against 2S was generated by immunizing New Zealand rabbits with purified recombinant protein 2S-His.ELISA results indicated that the two rabbits titre of anti-serum was up to 1∶ 25 600 and 1∶ 12 800 respectively.In conclusion,anti-serum,which could be used to detect spider dragline silk protein in vitro,was successfully obtained from 2S-pET-52b（＋）.This research laid basis for establishing method of expression spider dragline silk protein gene in hair.