CAJ | 학술논문

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30 mg/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。
목적：사선휴대항비인암질립 pFY 적은정고산균주。방법이 CaCl2법제비대장간균 JM109감수태,장항비인암질립 pFY 전화 JM109감수태,대경지평판상획득적균락진행사선,선출부합표준적단균락위균충,진행균충은정성실험。용질립제취시제합검측질립함량。장항비인암질립 pFY 전염도세포 CNE-2중,사담서람(MTT)비색법관찰전염시제급질립재체대세포생장증식적영향。결과사선득도균주적배양액중질립 DNA 함량위30 mg/mL,초라선 DNA 비례위92%。경전영화매절감정,해균주적50자대소휴대질립여원대일치。질립 pFY 대 CNE-2세포주생장유명현적억제작용。결론성공사선출휴대항비인암질립 pFY 적은정고산균주,위대비량제비림상응용급질립전정료기출。
Objective To screen the stable high-producing strains carrying anti-nasopharyngeal carcinoma(NPC)plasmid pFY.Methods Competent E.coli JM109 was prepared by the CaCl2 method and transformed with anti-NPC plasmid pFY.The bacterial colonies obtained from the agar plate were screened for selecting the single colony conforming to the standards as the bac-terial strain and conducting the stability test.The plasmid content was detected by the plasmid extraction reagent kit.Anti-NPC plasmid pFY was transfected into nasopharynegal carcinoma cell line CNE-2.The influence of transfection reagent and the plasmid vector on the cell proliferation was detected by MTT.Results The DNA concentration of plasmids in the culture solution of bacte-rial strain obtained by screening was 30 mg/mL.The proportion of supercoiled DNA was 92%.The identification of electrophoresis and restriction enzymes showed that the plasmids harbored in the 50th progeny of this strain were same as those in the primary. Plasmid pFY had the evident inhibiting effect on the growth of CNE-3 cell line.Conclusion The stable high-producing strains of E. coli carrying anti-NPC plasmid pFY is successfully screened out,which lays the foundation for large-scale preparation of plasmid pFY for clinical utility.