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In order to establish a rapid multiplex RT-PCR assay for detection and differentiation of variant，classical and attenuated vaccine strains of porcine epidemic diarrhea virus (PEDV). Two pairs of specific primers were designed based on S gene sequences and 0RF3 gene sequences of variant，classical and attenuated vaccine strains PEDV published in GenBank，to amplify the partial S gene fragment and ORF3 gene fragment of different PEDV strains，the type of PEDV strain could be differentiated according to the number of the fragment and the size of the fragment. Results showed that the multiplex RT-PCR could detect and differentiate variant， classical and attenuated vaccine strains PEDV. Variant PEDV strain generated two fragments，they were 826 bp of the S gene and 234 bp of the ORF3 gene，respectively. Classical PEDV strain only amplified 234 bp of the ORF3 gene，while attenuated vaccine strain produced 185 bp of the ORF3 gene. The multiplex RT-PCR did not cross-react with TGEV，PRoVA，PKV，PRRSV，PRV，CSFV，PCV-2，JEV，TTV and PPV used in the study. Testing of the sensitivity of RT-PCR indicated as low as 2. 3 x 10 -4 ng/μl nuclear acids could be detected accurately and rapidly. Ninety-one stool specimens collected from different farms in Guangxi Province were detected by the established multiplex RT-PCR，64 samples were positive for PEDV， of which 89. 06 % (57/64 ) was variant PED V strains，4. 69 % (3/64) was classical PEDV strain and 6. 25 % (4/64) was attenuated vaccine PEDV strain. Therefore the multiplex RT-PCR could be used as an effective tool for differentiating diagnosis of PEDV in epidemiological investigations. [ABSTRACT FROM AUTHOR]