用nested_PCR方法快速检测鲑鱼肾杆菌.pdfVIP

　26 5 　　　 Vol.26, No.5 　 2002 10 　　　　　　　　　　　JOURNAL OF F SHER ES OF CH NA　　　　　　 　　　　　 Oct., 2002　 :1000-0615(2002)05-0453-06 nested-PCR 刘　荭, 　高隆英, 　史秀杰, 　江育林 (, 　518010) :(nested-PCR)。 BKDR BKDF , 57 kDa 501 bp DNA , BKDR2 BKDF2 314 bp DNA 。15 , DNA 。, PCR 3 -1 1.8×10 CFU·mL , Nested-PCR PCR , 100 。, , 、、。 :;;; :S941.4 　　　:A Detection of Renibacterium salmoninarum by nested-PCR L U Hong, GAO Long-ying, SH Xiu-jie, J ANG Yu-lin (TheKey Laboratory of AquaticAnimalDisease, Shenzhen Exit-entry Inspection and QuarantineBureau, Shenzhen　518010, hina) Abstract:To develop a nested-polymerase chain reaction (nested-PCR )for detection of Renibacterium salmoninarum, the causative agent of bacterial kidney disease, 2 pairs of primers, BKDF and BDKR, BKDR2 and BKDF2, were designed for amplification of 501 bp and 314 bp DNA fragments of the sequence coding the 57 kDa cell surface protein of R .salmoninarum respectively.No specific fragments were amplified when other principal fish bacterial pathogens were used as templates in PCR and nested-PCR tests .The sensitivity of PCR 3 -1 was limited to 1.8×10 CFU·mL when the bacterial DNA was extracted by phenol-chloroform-isopentanol or boiling freezing-thawing method.The sensitivity of nested-PCR was 100 times higher than that of PCR test. The mixture of bacteria and cod eggs was detected using nested-PCR and the results showed that the nested-PCR was an effective and reliable method for the detection of R.salmoninarum. Key words:bacterial kidney disease;Renibacterium salmoninarum;nested-PCR;detecting methods (BKD), 。1930 , 、BKD。, 、、、、、、、、 14 BKD[ 1] 。,, 。(Renibacterium salmoninarum), 。 。、 ,,。 [2] ,,PCR :200 1-12-25 :(K009-2000) :　(197 1-)