EFFICIENT PRODUCTION OF NUCLEOTIDES OF SELECT VETERINARY FLAVIVIRUSES USING OVERLAP EXTENTION – POLYMERASE CHAIN REACTION

Tick-borne encephalitis (TBEV), Louping ill (LIV), and West Nile viruses (WNV) are notorious flaviviral agents that continue to plague domestic animals including horses, sheep, goats, cattle, pigs, dogs, birds, and also humans. To efficiently produce DNA fragments of TBEV, LIV, and WNV viruses, an algorithmic DNA primer design was developed using overlap extension-polymerase chain reaction (OE-PCR). Arithmetic formulation, with emphasis on the manipulation of melting temperatures (Tm) to enable advanced production of nucleotide lengths, was constructed. The method was validated using OE-PCR carried out using original and modified electrophoresis, with the latter producing 30 nanogram and prominent 256 bp of DNAs used for sequencing by Sanger method. Algorithmic formulation resulted to production of remarkable amount of artificial DNA products whereby accession numbers to each of the three amplified oligomers with complete nucleotide sequences of select veterinary flavivirus of 256 bp each for WNV, LIL and TBE were registered at DDBJ/NCBI. A relatively high synthesis performance of basically Tm-manipulated algorithmic OE-PCR design using simply designed oligonucleotides has paved for availability of ample amount of DNAs for extensive scientific and experimental explorations of select veterinary significant flaviviruses.  Future study employing the use of synthetic veterinary flaviviral DNAs is suggested.