Polymerase chain reaction: amplification and sequencing of the VP1 gene of foot-and-mouth disease virus type A

The polymerase chain reaction method (PCR) has been applied to the detection of FMD viral RNA in samples taken from the calf during the clinical stage of FMD. Total RNA was extracted with a guanidinum thiocyanate-phenol-chloroform method and reverse transcribed using AMV-reverse transcriptase. cDNA was used as a template for the amplification by PCR of the 672 bp of the VP1 coding sequence. The amplified fragment of cDNA was cloned in the pBS(+) phagemid-containing sites recognized by Smal endonuclease and expressed in E. coli strain MV1193. The first DNA strand was sequenced and concurrently an amino acid sequence was established. Comparison between VP1 amino acid sequence of FMDV types A and earlier described type O was performed.