Study on transformation of anti-nasopharyngeal carcinoma plasmid pFY and bacterial strains screening / 重庆医学

ABSTRACT

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.