卵巢癌人源性核糖体展示抗体库的构建

目的构建库容量大、多样性好的卵巢癌人源性核糖体展示抗体库。方法从10名卵巢癌患者的外周血淋巴细胞中提取总RNA,通过聚合酶链式反应（PCR）技术扩增人全套重链可变区基因（variable region of heavychain,V_H）和轻链可变区基因（variable region of light chain,V_L）,用重叠PCR技术对它们进行拼接,构建卵巢癌人源单链抗体库,并连接T载体转化大肠杆菌,经蓝白筛选,挑选阳性克隆,测序鉴定。结果成功构建卵巢癌人源单链抗体库,库容6.5×10~（13）,经测序验证多样性良好。结论成功构建的卵巢癌人源单链抗体库可用于进一步筛选目标抗原,为开发治疗性人源抗体奠定了实验基础。

Construction of single-chain antibody Fv library derived from ovarian cancer patients by ribosome display

Objective To construct high-capacity scFv（single-chain antibody Fv） library derived from ovarian cancer patients by ribosome display.Methods The total RNA was extracted from peripheral blood lymphocytes isolated from the blood of 10 ovarian cancer patients.Gene fragments encoding whole variable region of heavy chain（V_H） and variable region of light chain（V_L） were amplified by PCR and assembled into a single chain by overlapping PCR.Moreover,the template was verified by blue/white screening and sequencing.Results The human scFv library which contains 6.5×10~（13） members was successfully established.Conclusion This study highlights the ribosomal display technology for the selection of human antibody from patient-derived gene pools.