Allele-Specific PCR (AS-PCR) for <i>kdr</i> genotyping.

<p><b>A</b>: Schematic representation of the primer location and predicted size of PCR products. Cpp1–Cpp5 indicate PCR primers in which sequences are reported in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0011681#pone-0011681-t005" target="_blank">Table 5</a>. Primer pair Cpp2 and Cpp3 amplifies a 389 bp fragment for the wildtype susceptible allele (for codon TTA). Primer pair Cpp1/Cpp4 yields a 176 bp fragment for resistant L1014F allele (codon TTT). Similarly, primer pair Cpp1/Cpp5 leads to amplification of a 176 bp fragment diagnostic to the L1014S resistant allele (for codon TCA). <b>B</b>: An example of AS-PCR gel. Two PCR reactions were run in parallel for each specimen (lanes 1 and 2 for specimen 1; lanes 3 and 4 for specimen 2…). Samples in lanes 1, 3, 5, 7, 9 and 11 were amplified with the primers Cpp1, Cpp2, Cpp3 and Cpp4 to detect wildtype L1014 (TTA) and resistant L1014F (TTT) alleles, whereas samples in lanes 2, 4, 6, 8, 10 and 12 were amplified with primers Cpp1, Cpp2, Cpp3 and Cpp5 to detect wildtype L1014 (TTA) and resistant L1014S (TCA) alleles. Lanes 13 and 14 were negative control. Genotype results: specimen 1: TTA/TTA; specimen 2: TTT/TTT; specimen 3: TCA/TCA; specimen 4: TTA/TTT; specimen 5: TTA/TCA; and specimen 6: TTT/TCA.