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目的 :构建噬菌体表面表达随机 8肽文库 ;方法 :采用人工合成随机寡核苷酸、基因重组技术 ,噬菌体表面表达技术等 ,构建该文库 ;采用多聚酶链式反应技术、核酸杂交技术和测序分析评价鉴定。多聚酶链式反应所采用的上游引物包括载体克隆位点处部分碱基和部分外源基因的碱基 ;在核酸杂交中设计了噬菌体载体克隆位点互补探针 ,以更加准确地排除无外源基因插入的克隆 ;在测序鉴定中 ,选择了 2个混合探针杂交阳性的克隆。结果 :以上证实获得独特克隆为 2 1× 10 8;亲和富集实验证实所建噬菌体文库可以稳定扩展。结论 :我们成功地构建了噬菌体随机 8肽文库 ,其库容量为 2 1× 10 8。
OBJECTIVE: To construct a random 8-peptide library expressing phage surface. Methods: The library was constructed by using synthetic oligonucleotides, gene recombination technology and phage surface expression technique. The polymerase chain reaction, nucleic acid hybridization and sequencing analysis Identification. The upstream primers used in the polymerase chain reaction include bases at part of the vector cloning sites and some foreign genes. Complementary probes of phage vector cloning sites are designed in the nucleic acid hybridization to more accurately exclude exogenous Cloning of gene insert; in the sequencing identification, two hybrid probes hybrid positive clones were selected. Results: The above confirmed that the unique clone was 2 1 × 10 8; affinity enrichment experiments confirmed that the constructed phage library can be stably expanded. Conclusion: We successfully constructed a phage random 8-peptide library with a capacity of 2 × 10 8.