中文摘要：

为了筛选柔嫩艾美耳球虫子孢子入侵相关的蛋白,利用酵母双杂交体系构建了柔嫩艾美耳球虫子孢子酵母双杂交cDNA文库。提取纯化的子孢子总RNA,经MMLV-RT反转录合成cDNA第一链后,利用LD-PCR扩增合成双链cDNA（ds cDNA）,dscDNA经纯化柱CHROMA SPINT＋TE-400 Columns纯化剔除小于200 bp的片段。将纯化后dscDNA、pGADT7-Rec及Carrier DNA共转化到已经制备好的酵母感受态细胞Y187中,dscDNA与pGADT7-Rec以同源重组的方式在细胞内进行连接,经过缺陷性培养基（SD/-Leu）的筛选得到酵母双杂交cDNA文库。结果显示,构建的柔嫩艾美耳球虫子孢子酵母双杂交cDNA文库的转化率为4.33×10^5/μg pGADT7-Rec,文库滴度为3.62×10^7 cfu/m L。随机挑取32个单克隆进行PCR检测,插入片段大小为200~2000bp,重组率为93.75%,并以该文库作为模板,用柔嫩艾美耳球虫5对特异性引物进行PCR扩增,获得了这5个基因片段。结果表明成功构建了柔嫩艾美耳球虫子孢子酵母双杂交cDNA文库,为进一步筛选和研究球虫入侵互作蛋白奠定了基础。

英文摘要：

In order to screen the proteins related to the invasion of Eimeria tenella sporozoites, the yeast two-hybrid cDNA library of E. tenella sporozoites was constructed in the present study. The total RNA was isolated from the sporozoites and used as the template to synthesize the first-strand cDNAs. The dscDNAs were acquired by long-distance polymerase chain reaction （LD-PCR） using the first- strand cDNAs as the template and purified with Chroma Spin TE-400 Column to remove less than 200 bp fragments. The dscDNA,pGADT7-Rec and CarrierDNA were co-transformed into Y187 yeast competent cells. The dscDNA were then connected with pGADT7- Rec by homologous recombination reaction to construct the yeast two-hybrid cDNA library orE. tenella sporozoites. The results showed that the conversion ratio and titer of the eDNA library were 4.33 ×1^05/μg pGADT7-Rec and 3.62 ×10^7cfu/mL, respectively, As demonstrated in PCR amplification, the cDNA library contained approximately 93.75 % recombinant clones and the inserted eDNA fragments were between 200 bp and 2000 bp. Furthermore, 5 gene fragments were obtained by PCR amplication using the eDNA library as template and 5 specific pairs of primers of E. tenella. Therefore, it concluded that a yeast two-hybrid eDNA library of the sporozoites was constructed for screening invasion-related interaction proteins of the sporozoites of E. tenella.