论文部分内容阅读
目的:构建肺癌噬菌体展示肽库,筛选出肺癌早期检测的分子标志群。方法:取30例第二军医大学长海医院手术后立即冻存的肺癌组织标本构建肺癌噬菌体展示肽库;用结合protein-A/G的琼脂糖珠分别富集肺癌组及非肿瘤对照组血浆中的抗体进行5轮亲和筛选,富集肺癌特异性的相关肽;随机挑取5轮筛选后的500个噬菌体克隆,用ELISA法进一步筛选肺癌/对照D比值大于2.0的肺癌特异标志物克隆,进行基因序列测定和蛋白功能预测。结果:(1)该噬菌体展示肽库的滴度为3.0×106pfu,库容9.0×106pfu,随机挑选20个噬菌斑经PCR鉴定其重组率为60%。(2)共筛选出19个有意义的噬菌体,测序后,预测其蛋白质功能,9个与癌症相关,8个功能未知,2个与癌症不相关。结论:筛选出的噬菌体克隆所表达的抗原很可能是早期诊断肺癌的标志物。
Objective: To construct a phage display peptide library of lung cancer and screen out molecular markers for early detection of lung cancer. Methods: Lung cancer phage display peptide library was constructed from 30 lung cancer samples frozen immediately after surgery in Changhai Hospital of Second Military Medical University. The plasma of lung cancer group and non-tumor control group were enriched by protein-A / G-bound agarose beads Of the antibodies were screened by 5 rounds of affinity screening to enrich for specific lung cancer related peptides. Five rounds of phage clones after 5 rounds of screening were randomly selected, and lung cancer-specific marker clones with lung cancer / control D ratios greater than 2.0 were further screened by ELISA. Gene sequencing and protein function prediction. Results: (1) The phage display peptide library had a titer of 3.0 × 106pfu and a storage capacity of 9.0 × 106pfu. 20 plaques were randomly selected and the recombination rate was 60% by PCR. (2) A total of 19 phage screened out were screened, sequenced and predicted for protein function. Nine were associated with cancer, eight were unknown, and two were unrelated to cancer. Conclusion: The antigens expressed by the selected phage clones are likely to be the early markers of lung cancer diagnosis.