A novel minisequencing single-nucleotide polymorphism marker of the lysozyme gene detects high hatchability of Tsaiya ducks (Anas platyrhynchos)

Lysozyme, one of the major albumen antimicrobials, can break down the polysaccharide walls of a broad spectrum of bacteria. This study presents a novel lysozyme marker of high hatchability in the form of minisequencing single-nucleotide polymorphisms (SNPs). Recently, lysozyme was identified by complementary DNA microarray analysis as one of several differentially expressed genes noted to influence hatchability and recognized as a marker candidate for animal marker-assisted selection. Higher levels (P < 0.05) of lysozyme mRNA (via real-time polymerase chain reaction analysis) and protein (in Western blotting results) were found to be associated with a high-hatchability phenotype. In the preliminary sequence analysis of this study, TsLy1-1 and TsLy1-2 primer pairs, designed according to the lysozyme sequence, were used to amplify small-scale genomic DNA samples from animals in two extreme groups of hatchability. Sequence analysis of the amplified 763-bp DNA products clearly showed that AA and GG genotypes of SNP g.390A > G were from the ducks of the low- and high-hatchability groups, respectively. The SNP g.390A > G also created a new specificity protein 1 transcription factor binding site in the lysozyme gene. Primer pairs of TsLy2-1 and TsLy2-2 then probed the amplified 763-bp DNA products to produce a shorter fragment for easier minisequencing analysis to divide 114 ducks into GG, GA, and AA genotypes. The GG ducks had the highest hatchability, representing that a new lysozyme SNP marker of good hatchability performance can be used for the purpose of marker-assisted selection in Tsaiya ducks.