环介导等温扩增技术检测铜绿假单胞菌的研究  

Rapid detection of Pseudomonas aeruginosa by loop-mediated isothermal amplification

作　　者：曹科峰[1,2] 沈继录[3] 汪学龙[1] 

机构地区：[1]安徽医科大学病原生物学教研室,合肥230032 [2]安徽省太和县中医院检验科,太和236600 [3]安徽医科大学第一附属医院检验科,合肥230022

出　　处：《安徽医科大学学报》2017年第3期450-453,共4页Acta Universitatis Medicinalis Anhui

基　　金：国家自然科学基金(编号:81171618)

摘　　要：用环介导等温扩增技术(LAMP)快速、灵敏地检测铜绿假单胞菌。利用铜绿假单胞菌gbca基因,特异性设计3对LAMP引物,在反应体系中添加羟基萘酚蓝(HNB)荧光染料,通过反应前后颜色的变化肉眼观察结果,并用琼脂糖凝胶电泳加以验证。对临床标本利用LAMP和PCR法平行检测,验证该法灵敏度、特异性。针对铜绿假单胞菌DNA特异性引物的LAMP法可以扩增,其他无扩增产物。本实验建立的LAMP法具备简便、快速、特异性强和灵敏度高的特点,适于实际应用。A rapid and sensitive method of loop-mediated isothermal amplification（LAMP） was established to de- tect Pseudomonas aeruginosa （P. aeruginosa）. Three pairs of LAMP primers （ inner, outer and ring primers ） were designed according to the gbca gene of P. aeruginosa. Since adding hydroxy naphthol blue （HNB） to the reaction system, a positive reaction was indicated by a colorchange before and after the reaction, and was verified by agarose gel eleetrophoresis. Both LAMP and PCR were applied to detect clinical specimens, the sensitivity and specificity of the detection method were evaluated, and were compared with those of conventional PCR. A LAMP method for de- tecting P. aeruginosa was successfully established. The LAMP method showed specificity for P. aeruginosa without other bacteria amplification. The established LAMP method in this study enables rapid, sensitive and specific detec- tion of P. aeruginosa, and can be applied for grass roots and small scale laboratories as well as field surveillance.

关 键 词：铜绿假单胞菌 环介导等温扩增 检测 

分 类 号：R73-351[医药卫生—肿瘤]