The diseases caused by Fusarium oxysporum f.sp.cubense and Dickeya zeae have brought serious challenges to the development of banana industry,and it is necessary to develop a detection method by multiplex polymerase chain reaction(multiplex PCR)technology for them.The specific amplification primers for the molecular multiplex PCR system were designed as followed:FOC-F/-R targeted the sequences of the contig 438(35 631-37 693 bp)(GenBank:AMGP01000438.1)in the DNA of F.oxysporum f.sp.cubense race 1(FOC1)and the contig 195(4 028-6 126 bp)(GenBank:AMGQ01000195.1)in the DNA of F.oxysporum f.sp.cubense race 4(FOC4),and there was a difference of 160 bp between FOC1 and FOC4.The primers gyrB-F/-R were designed based on the gene sequence(GenBank:JQ284039)in D.zeae.The results of multiplex PCR showed that this technology could simultaneously detect FOC1,FOC4 and D.zeae in one PCR amplification reaction.Moreover,the minimum limit of DNA concentration for detection of F.oxysporum f.sp.cubense was 0.1 ng/μL,and the sensitivity for detecting D.zeae was 103 cfu/mL.The detection results were stable and reliable.Therefore,the multiplex PCR detection method can be effectively used to detect F.oxysporum f.sp.cubense and D.zeae in infected banana plant tissues,and can also be used to monitor banana seedling-and soil-borne pathogens in the field,which provides a useful tool for safe planting of banana.
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