[Selection of differential DNA fragment of Tilletia controversa Kühn and establishment of molecular detection approach].
A reliable and simple polymerase chain reaction method for TCK pathogen was established firstly. A 1322bp unique fragment of TCK was amplified and identified by the technique of semi-specific random amplified polymorphism (RM-PCR). Two pairs of species-specific primers CQUK2/CQUK3 and CQUK4/CQUK5 were designed according to the unique fragment of TCK. The first pair primers were capable to stably amplify target DNA band of 747bp from chromosomal DNA of 18 strains of TCK isolates without any DNA bands obtained from 29 strains of TCT. The second pair primers could produce a 200bp target DNA band stably, while no band was amplified from teliospore or mycelium DNA of TCT of strains. Tilletia genus primers were used as internal control of molecular detection system, which can detect whether the PCR inhibitors exist in testing sample or avoid pseudo-negative and pseudo-positive of PCR reaction. The molecular detection approach could rapidly, accurately detect and identify the DNA of teliospore or mycelium of TCK from wheat tissues.
Nian
Si-ji
SJ
College of Bioengineering of Chongqing University, Genetic Engineering Research Centre of Chongqing University, Key Laboratory of Gene Function and Regulation at Chongqing, Chongqing University, Chongqing 400030, China. sijinian@hotmail.com
Yin
You-ping
YP
Yuan
Qing
Q
Xia
Yu-xian
YX
Wang
Zhong-kang
ZK
D001487:Basidiomycota / Q000235:genetics* / Q000302:isolation & purification; D004271:DNA, Fungal / Q000235:genetics*; D010935:Plant Diseases / Q000382:microbiology*; D019105:Random Amplified Polymorphic DNA Technique / Q000379:methods*; D014908:Triticum / Q000382:microbiology*
D004271:DNA, Fungal