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目的比较Gelred、GoldView及溴化乙锭(EB)三种染料在病原菌脉冲场凝胶电泳(PFGE)中的染色效果,选择更为安全和灵敏的核酸染料,并形成标准化操作应用于病原菌分子分型监测网络PulseNet China。方法选择霍乱弧菌作为评价菌株,按照病原菌分子分型监测网络(PulseNet)的PFGE标准操作方案进行电泳,采用Gelred、GoldView及EB三种染料分别染色,分析比较实验结果。结果 Gelred染色的胶块清晰度最好,EB次之,均高于GoldView;Gelred染色的图谱,相邻条带边缘清晰,更容易辨别区分。EB和GoldView染色后必须脱色,使用Gelred染色脱色时间较短,亦可以不经过脱色步骤,但脱色后的电泳条带边缘更加锐利。结论 Gelred是一种更安全、更灵敏的核酸染料,可以取代EB应用于PFGE。制定了使用Gelred进行PFGE染胶的标准方案,用于PulseNet China网络的PFGE分析。
OBJECTIVE To compare the staining results of Gelred, GoldView and ethidium bromide (EB) in the pulsed-field gel electrophoresis (PFGE) of pathogenic bacteria and to select safer and more sensitive nucleic acid dyes and to establish a standard operation for the molecular identification of pathogenic bacteria PulseNet China. Methods Vibrio cholerae was selected as the evaluation strain. Electrophoresis was performed according to the PFGE protocol of PulseNet, and Gelred, GoldView and EB dyes were used respectively to analyze and compare the experimental results. Results Gelred staining showed the best clarity and EB followed by GoldView and Gelred staining. The edges of adjacent bands were clear and easy to distinguish. The EB and GoldView stains must be decolorized, Gelred stain should be used for a short period of decolorization, or the decolorization step may be omitted, but the edge of the decolorized electrophoresis strip is sharper. Conclusion Gelred is a safer and more sensitive nucleic acid dye that can replace EB for PFGE. A standard protocol for gel staining of PFGE was developed for the PFGE analysis of the PulseNet China network.