论文部分内容阅读
从伏马菌素B1(FB1)杂交瘤细胞株F3中扩增出V H和V L基因片段,再经重叠延伸PCR(SOE-PCR)拼接扩增得到单链抗体(ScFv)基因,然后克隆到pCANTAB 5E噬菌粒载体上,转化感受态大肠杆菌TG1,并经辅助噬菌体M13K07超感染,构建FB1毒素噬菌体单链抗体库,ScFv基因插入率为100%,库容约为1.5×107,噬菌体的滴度为2.1×1015PFU·mL-1。免疫亲和筛选和富集后,经ELISA法最终筛选得到5株可分泌阳性噬菌体抗体的细菌。结论:伏马菌素B1噬菌体单链抗体库构建成功。
The VH and VL gene fragments were amplified from Fusamycin B1 (FB1) hybridoma cell line F3. The single chain Fv gene was amplified by overlap extension PCR (SOE-PCR) and cloned into pCANTAB 5E phagemid vector was transformed into competent E. coli TG1 and superinfected with the helper phage M13K07 to construct the FB1 toxin phage scFv library. The insertion rate of ScFv gene was 100% and the storage capacity was about 1.5 × 107. The phage titer Was 2.1 × 10 15 PFU · mL -1. After immunoaffinity screening and enrichment, 5 strains of bacteria secreting positive phage antibody were finally screened by ELISA. Conclusion: The fuscin B1 phage scFv library was constructed successfully.