PCR primers and probe for the A. bicolor subspecies-specific allelic discrimination. PCR primers and probe developed in this study for the allelic discrimination assay of the two subspecies of A. bicolor bicolor and A. bicolor pacifica. Partial DNA nucleotide sequences of the 16S rRNA gene of 19 species/subspecies of Anguilla were aligned. Dots indicate the same nucleotide with those of A. bicolor bicolor in the top line. Base positions are according to the complete sequence of the locus in A. bicolor. Arrows and horizontal bar show the position of primers (AD-Abic-16S-F1278v2 and AD-Abic-16S-R1420) and probe (AD-BBI/BPA-16S-p) binding sites, respectively. The subspecies-specific single substitution in A. bicolor is shown with an arrowhead at the 1,318th position. Accession numbers of sequences for each species are as follows: A. bicolor bicolor, AP007236; A. bicolor pacifica, AP007237; A. anguilla, AP007233; A. australis australis, AP007234; A. australis schmidtii, AP007235; A. bengalensis bengalensis, AP007246; A. bengalensis labiata, AP007245; A. borneensis, AP007238; A. celebesensis, AP007239; A. diffenbachii, AP007240; A. interioris, AP007241; A. japonica, AB038556; A. luzonensis, AB469437; A. marmorata, AP007242; A. megastoma, AP007243; A. mossambica, AP007244; A. obscura, AP007247; A. reinhardtii, AP007248; A. rostorata, AP007249.