抗鼻咽癌质粒pFY转化和菌种筛选的研究

发布时间：2018-03-28 00:05

本文选题：鼻咽肿瘤　切入点：感受态细胞　出处：《重庆医学》2015年26期

【摘要】：目的筛选携带抗鼻咽癌质粒pFY的稳定高产菌株。方法以CaCl2法制备大肠杆菌JM109感受态,将抗鼻咽癌质粒pFY转化JM109感受态,对琼脂平板上获得的菌落进行筛选,选出符合标准的单菌落为菌种,进行菌种稳定性实验。用质粒提取试剂盒检测质粒含量。将抗鼻咽癌质粒pFY转染到细胞CNE-2中,四氮唑蓝(MTT)比色法观察转染试剂及质粒载体对细胞生长增殖的影响。结果筛选得到菌株的培养液中质粒DNA含量为30mg/mL,超螺旋DNA比例为92%。经电泳和酶切鉴定,该菌株的50子代所携带质粒与原代一致。质粒pFY对CNE-2细胞株生长有明显的抑制作用。结论成功筛选出携带抗鼻咽癌质粒pFY的稳定高产菌株,为大批量制备临床应用级质粒奠定了基础。
[Abstract]:Objective to screen stable and high-yielding strains carrying anti-nasopharyngeal carcinoma (NPC) plasmid pFY. Methods Escherichia coli JM109 was prepared by CaCl2 method, and the plasmid pFY was transformed into JM109 receptive state to screen the colony on Agar plate. A single colony that meets the standard was selected as the strain, and the stability experiment was carried out. The plasmid content was detected by plasmid extraction kit. The anti-nasopharyngeal carcinoma plasmid pFY was transfected into the cell CNE-2. The effect of transfection reagent and plasmid vector on cell growth and proliferation was observed by MTT colorimetry. Results the plasmid DNA content and superhelix DNA ratio were 30 mg / mL and 92% respectively. The plasmids carried in 50 progenies of this strain were consistent with the original ones. Plasmid pFY could inhibit the growth of CNE-2 cell line obviously. Conclusion A stable and high yield strain carrying plasmid pFY against nasopharyngeal carcinoma was successfully screened. It lays a foundation for large batch preparation of clinical application grade plasmids.
【作者单位】：佛山科学技术学院医学院医学检验系;
【基金】：广东省医学科研基金资助项目(A2013687) 广东省大学生创新创业训练计划项目(1184713046)
【分类号】：R739.63

【参考文献】