日用化学工业（中英文）2024，Vol.54Issue(1)：45-50,6.DOI:10.3969/j.issn.2097-2806.2024.01.006
日勾维多细菌源菌双重PCR快速检测方法建立
Establishment of duplex-PCR method for rapid detection of Pluralibacter gergoviae
摘要
Abstract
A rapid detection method for Pluralibacter gergoviae was established,and this method was used to quickly and effectively detect Pluralibacter gergoviae in samples.Four house-keeping genes(gyrB,infB,atpD and ropB)were selected as target genes,and 4 pairs of primers were designed.The bacterial suspension was used as amplification,and the 4 pairs of primers could specifically bind with the Pluralibacter gergoviae to produce bright DNA bands.Among the four pairs of primers,only gyrB-F/gyrB-R and ropB-F/ropB-R obtained a small amount of non-specific DNA bands when Escherichia coli,Staphylococcus aureus,Pseudomonas aeruginosa,Enterobacter cloacae and Burkholderia cepacia were used as DNA templates.Two pairs of specific primers gyrB-F/gyrB-R and ropB-F/ropB-R(insert)were used to establish the rapid detection method of duplex-PCR for Pluralibacter gergoviae in rinsing cosmetics.The specificity of duplex-PCR was improved by optimizing the annealing temperature of amplification.When the annealing temperature was 60℃,using the primer pairs gyrB-F/gyrB-R and ropB-F/ropB-R(insert)to amplify 4 strains of Pluralibacter gergoviae,two specific target DNA bands were obtained,but Escherichia coli,Staphylococcus aureus,Pseudomonas aeruginosa,Enterobacter cloacae and Burkholderia cepacia would obtain non-specific DNA bands.When the annealing temperature was optimized and set at 65℃,the specific target DNA bands could be amplified when 4 strains of Pluralibacter gergoviae were used as DNA templates.When Escherichia coli,Staphylococcus aureus,Pseudomonas aeruginosa,Enterobacter cloacae and Burkholderia cepacia were used as DNA templates,no DNA bands were detected.When the concentration of Pluralibacter gergoviae in the sample reached 4.4 CFU/mL,duplex-PCR was able to detect Pluralibacter gergoviae in the sample with high sensitivity.Duplex-PCR method could be used to quickly detect the Pluralibacter gergoviae in the rinsing cosmetics such as shower gel and shampoo.关键词
化妆品/日勾维多细菌源菌/双重PCR/快速检测Key words
cosmetics/Pluralibacter gergoviae/duplex-PCR/rapid detection分类
化学化工引用本文复制引用
刘丰,邓源昌,应国红,王晓炜..日勾维多细菌源菌双重PCR快速检测方法建立[J].日用化学工业（中英文）,2024,54(1):45-50,6.基金项目
广东省药品监督管理局2022年科技创新项目(2022TDB67) （2022TDB67）
广东省药品监督管理局2023年科技创新项目(2023TDB53) （2023TDB53）
深圳市药品检验研究院(深圳市医疗器械检测中心)2023年度十大科研课题(YNKT20230205) （深圳市医疗器械检测中心）
