人端粒酶逆转录酶在大肠杆菌中表达的初步研究

Expression of Human Telomerase Reverse Transcriptase Segment in E. coli

端粒酶是迄今为止最为广谱的肿瘤标志物,是一种良好的肿瘤检测和治疗的靶分子.曾报道了人端粒酶逆转录酶(hTERT)一个基因片段的表达和相应的抗体用于端粒酶及hTERT检测的研究,为了获得更多针对hTERT不同表位的抗体,根据hTERT的抗原性进一步选取了hTERT的数个片段来制备抗体,这里初步探讨了这些抗原片段在大肠杆菌中的表达及各种因素对表达的影响.实验结果表明,位于hTERT蛋白中部和C末端的片段可在大肠杆菌中表达,但位于N末端的片段在各种条件下均不能表达.另外,就选取的hTERT各片段而言,诱导条件的优化只能在量上提高表达的水平,不能决定片段是否表达.而密码子偏好性,启动子强弱及mRNA二级结构的稳定性可能对片段的表达具有更大的决定作用.

As the most widely expressed biomarker of tumor cells, telomerase is the ideal target for tumor detection and therapy. Research has been reported on the expression and purification of human telomerase reverse transcriptase segment in E. coli, and also antihTERT antibody generation and immunohistochemistry analysis of tumor tissue with this antibody. In order to prepare antibodies which could recognize different epitopes of hTERT, three different segments from hTERT protein were selected as antigens to produce antibodies. These segments were expressed in different strains of E. coli and under the control of several different promoters. The results revealed that multifactors exist which affect the expression level of the segments. For example, optimization of the induction conditions might slightly affect the expression, while the codon usage, promotor activity and stability of secondary structure of mRNA may play key roles in the expression of these segments.