• 方法与技术 •

### 棉花黄萎病菌实时荧光定量PCR检测方法的建立及应用

1. (1中国药科大学生命科学与技术学院， 南京 210009； 2南京大学医学院附属金陵医院， 南京 210093； 3国家环保总局南京环境科学研究所， 南京 210037)
• 出版日期:2015-07-10 发布日期:2015-07-10

### Development and application of a real-time PCR assay for the detection and quantification of Verticillium dahliae.

SHEN Yuan-jie1, QI Xie-min2, LIU Biao3, ZHOU Guo-hua1,2, LI Tai-ming1**

1. (1School of Life Science and Technology, China Pharmaceutical University, Nanjing 210009, China; 2Jingling Hospital, Medical School of Nanjing University, Nanjing 210093, China; 3Nanjing Institute of Environmental Sciences, Ministry of Environmental Protection, Nanjing 210037, China)
• Online:2015-07-10 Published:2015-07-10

Abstract: A specific real-time PCR assay based on the ITS region of rDNA was established to quantitatively detect Verticillium dahliae of cotton in a naturally infested soil. The standard curve of the realtime PCR system was constructed by a 324 bp PCR amplification product and the specificity, sensitivity and reproducibility of the method were evaluated. The results showed that the realtime PCR assay had good specificity and sensitivity and the detection limit was
100 copies·μL-1. The correlation coefficient of the standard curve was 0.994 and the PCR amplification efficiency reached 91.5%. Using the realtime PCR assay, soil samples from a transgenic cotton field and nontransgenic one were detected. The results indicated that the amount of V. dahliae in the transgenic cotton field was significantly higher than that of conventional cotton, which showed consistency with the observations achieved in the two cotton fields. As a reliable and sensitive assay, the realtime PCR system provided an effective method for the rapid detection of V. dahliae and the control of cotton diseases.