中文摘要：

目的 为筛选微小隐孢子虫保护性抗原基因，构建了微小隐孢子虫T7噬菌体展示文库。方法 用Trizol试剂提取微小隐孢子虫总RNA，分离纯化mRNA，经反转录合成双链eDNA。在双链eDNA末端加上定向EcoR Ⅰ／Hind Ⅲ接头，用EcoR Ⅰ和Hind Ⅲ消化接头，使形成两端分别带有EcoR 和Hind Ⅲ粘性末端的双链eDNA。经Mini Column纯化，收集400bp以上的双链eDNA片段，将其连接于带有EcoRⅠ和Hind Ⅲ末端的T7 Seleet 10～3b载体，经体外包装后，以BLT5403为受体菌构建T7噬菌体展示文库。结果 经测定，库容量为1．2×10^7 pfu／mL，重组率为96．7％，扩增后文库滴度为2．4×10^10pfu／mL。对随机挑取的100个噬菌斑进行PCR鉴定，92％的插入片段大于400bp。结论 成功构建了微小隐孢子虫T7噬菌体展示文库。

英文摘要：

To screen protective antigen gene of Cryptosporidium parvum, the T7 phage display library from C. parvum was constructed, and the total RNA was extracted from C. parvum by Trizol reagent. The mRNA was isolated from total RNA by PolyATract mRNA Isolation Kit and the ds eDNA was synthesized by reverse transcription. In addition the directional EcoR Ⅰ/Hind m linkers were ligated into the ends of ds eDNA and the ds eDNA was digested with EcoR Ⅰ and Hind Ⅲ, which resulted in ds eDNA with EcoR Ⅰ and Hind Ⅲ ends. And the ds eDNA fragments longer than 400bp in length were fractionated by Mini Column, and then ligated into the T7 Select 10-3b vertor with EcoR Ⅰ and Hind m ends.After packaging in vitro, the T7 Select 10-3b vertor was transformed into BLT5403 to construct the T7 phage display library. The experimental results showed that the library contained 1.2 × 10^7 clones, and approximately 96.7% of the library were recombinant. The titer of the amplied library was 2.4 × 10^10 pfu/mL. The eDNA fragments longer than 400bp in length were 92 % of 100 plaques by PCR identification. These results suggests that the T7 phage display library from C. parvum is constructed successfully.