Expression of an intron-containing gene from P. falciparum using three sequential PCR steps

The amplification of eukaryotic genes, with introns is very tedious and expensive. The only available lab method is through cDNA synthesis, which has a very low success rate in generating a pure gene fragment. Therefore, it is of interest to devise a strategy to amplify a P. falciparum RPL12_mito gene (PfRPL12_mito) containing an intron through three sequential PCR steps using genomic DNA (gDNA) and primers with a minimum 15 bp 5'-overhang. The absence of an intronic segment and a continuous exonic region in the amplified gene fragment was confirmed using Sanger sequencing.