Rapid single-tube splice variants typing of the BF gene based on dual-primer RT-PCR amplification that influence resistance/susceptibility to Mareks disease in chicken

A new effective splice variants typing based on multiplex allele-specific dual-primer RT-PCR assay was developed in a single tube for the rapid detection of the exon 7 splice variant of the BF gene. With 2 pairs of primers, one pair was used for amplifying cDNA fragments containing exon 7 of the BF gene, the other does not contain exon 7 of the BF gene. The templates were amplified in one tube and the type of splice variants was determined by the length of products to be extended and by analysis of nucleotide sequences of these BFs. Results obtained for all samples showed 100% accuracy compared to those obtained with a semi-nested PCR (snPCR) assay of 100% accuracy, but which need two round PCR assay. The dual-primer RT-PCR assay was more rapid and easy to operate than the snPCR assay.