Molecular identification of avirulent genes of Avr1a, Avr1k and Avr3a in Phytophthora sojae

Abstract:

[Objective] To provide method for rapid molecular detection of Avr1a, Avr1k and Avr3a, avirulent genes of Phytophthora sojae, and also to provide some references for rapid molecular detection of other avirulent genes of P. sojae. [Methods] According to Genbank sequence table of Avr1a, Avr1k and Avr3a, avirulent genes of P. sojae, specific primers were filtered out respectively for the three genes by using primer design method. 86 strains of P. sojae which have been identified virulence on the three differential soybean varieties were detected by PCR on the basis of optimized reaction system and amplification conditions. A specific detection system was established for detecting Avr1a, Avr1k and Avr3a, avirulent genes of P. sojae. The results of molecular identification and inoculation identification were compared and analized. And then amplified true and false positive bands were recovered respectively and cloning sequenced and compared with the original sequences of the three avirulent genes, to verify if Avr1a, Avr1k and Avr3a were suitable to be detected by the method of molecular markers. [Results] All the specific primers screened out could amplify a band with length of about 550 bp. The coincidence rate of molecular and inoculation identification for the three avirulent genes were Avr1a-45.3%, Avr1k-84.9% and Avr3a-97.7%. The true positive bands of three avirulent genes had over 97 percent of consistency in sequence with the original one. The false positive bands of Avr1a were about 80 percent, and Avr1k and Avr3a were less than 30 percent of consistency with their original sequences. [Conclusion] The detection system established by using the primers filtered out by utilizing the gene sequence of Avr1a, Avr1k and Avr3a can be used to detect Avr3a rapidly, but not be suitable for detection of Avr1a, whether fit for detection of Avr1k needed further study.