Department of Biotechnology, MingDao University, Pettow, ChangHua, Taiwan.
Theriogenology. 2013 May;79(8):1218-1223.e1. doi: 10.1016/j.theriogenology.2013.02.021. Epub 2013 Mar 23.
Our previous transcriptome analysis using a cDNA microarray identified differentially-expressed transcripts in Tsaiya ducks (Anas platyrhynchos); we concluded that the ovalbumin gene might be involved in duck hatchability. In the present study, associations of single nucleotide polymorphism (SNP) genotypes of the duck ovalbumin gene with hatchability were investigated. To confirm the cDNA microarray analysis, real-time polymerase chain reaction (PCR) and Western blot analysis were used to validate ovalbumin gene expression. The messenger RNA and protein expression of the ovalbumin gene were higher (P < 0.05) in the low-hatchability group (1.00 ± 0.19; 30.36 ± 3.51 arbitrary units) than in high-hatchability counterparts (0.56 ± 0.07; 8.53 ± 2.97 arbitrary units), consistent with the previous cDNA microarray analysis. The PCR products (506 base pairs) of ovalbumin gene amplified by the primer pair of TovaF and TovaR from the genomic DNA templates of 10 ducks were sequenced and a g.385 C>T SNP site in the 506-base pair sequence of the ovalbumin gene identified. Genotyping of SNP of 187 ducks was then carried out by PCR restriction fragment length polymorphism and minisequencing methods. Based on SNP genotypes of the duck ovalbumin gene, there were three types: CC, TT, and CT. Birds with the CC and TT genotypes had higher hatchability (79.59 ± 3.40, 76.35 ± 1.77) (P < 0.05) than those with a CT genotype (65.77 ± 2.07). In conclusion, the ovalbumin gene was an important candidate gene that can be used for marker-assisted selection to increase hatchability in Tsaiya ducks.
我们之前使用 cDNA 微阵列进行的转录组分析鉴定了 Tsaiya 鸭（Anas platyrhynchos）中差异表达的转录本；我们得出结论，卵白蛋白基因可能参与了鸭蛋的孵化率。在本研究中，研究了鸭卵白蛋白基因的单核苷酸多态性（SNP）基因型与孵化率的关系。为了验证 cDNA 微阵列分析的结果，使用实时聚合酶链反应（PCR）和 Western blot 分析验证了卵白蛋白基因的表达。卵白蛋白基因的信使 RNA 和蛋白表达在低孵化率组（1.00 ± 0.19；30.36 ± 3.51 任意单位）中较高（P < 0.05），而在高孵化率组中则较低（0.56 ± 0.07；8.53 ± 2.97 任意单位），与之前的 cDNA 微阵列分析结果一致。从 10 只鸭的基因组 DNA 模板中用引物对 TovaF 和 TovaR 扩增的卵白蛋白基因的 PCR 产物（506 个碱基对）被测序，并在卵白蛋白基因的 506 个碱基对序列中鉴定出 g.385 C>T SNP 位点。然后，通过 PCR 限制性片段长度多态性和 minisequencing 方法对 187 只鸭子的 SNP 进行基因分型。基于鸭卵白蛋白基因的 SNP 基因型，有三种类型：CC、TT 和 CT。具有 CC 和 TT 基因型的鸟类孵化率较高（79.59 ± 3.40，76.35 ± 1.77）（P < 0.05），而具有 CT 基因型的鸟类孵化率较低（65.77 ± 2.07）。总之，卵白蛋白基因是一个重要的候选基因，可用于标记辅助选择，以提高 Tsaiya 鸭的孵化率。
