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目的评价不同浓度叠氮溴化丙啶(PMA)在不同照射时间对大肠杆菌DNA灭活的效果,确定在实际使用中的最佳浓度和照射时间。方法选择相当于107cfu大肠杆菌(8099)的DNA作为灭活对象,加入终浓度为50μmol/ml、100μmol/ml、150μmol/ml和200μmol/ml的PMA,使用波长为460 nm的LED灯,分别照射5 min、10 min和15 min,然后进行Real Time PCR扩增。结果当PMA的浓度达到150μmol/ml,照射5 min时,大肠杆菌Real Time PCR扩增结果为阴性。结论在实际使用中,要完全灭活107cfu的大肠杆菌(8099)的DNA,需要≥150μmol/ml的PMA,照射时间≥5 min较合适。
Objective To evaluate the effect of different concentrations of azido-propidium bromide (PMA) on inactivation of Escherichia coli DNA at different irradiation times, and to determine the optimal concentration and irradiation time in actual use. Methods The DNA of 107cfu Escherichia coli (8099) was selected as inactivation object. PMA with the final concentration of 50μmol / ml, 100μmol / ml, 150μmol / ml and 200μmol / ml was added. 5 min, 10 min and 15 min, followed by Real Time PCR amplification. Results When the concentration of PMA reached 150 μmol / ml and the irradiation time was 5 min, the results of Real Time PCR amplification of E. coli were negative. Conclusion In actual use, to completely inactivate 107cfu of E. coli (8099) DNA, the need for ≥ 150μmol / ml of PMA, irradiation time ≥ 5min more appropriate.