Selection of differential DNA fragment of Tilletia controversa Kühn and establishment of molecular detection approach
小麦矮腥黑穗菌差异片段筛选与分子检测体系的建立

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 CQUK_2/ CQUK_3 and CQUK_4/ CQUK_5 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.