母源性叶酸缺乏对子代肉鸡脂肪相关基因的影响及表观遗传学调控机制

Folate is methyl donor in synthesizing S-adenosyl-L-methionine (SAM), and animal body needs folate to synthesize DNA and methylate DNA. It is not reported that genes involved in lipid and their mechanisms of epigenetic regulation were affected by folate deficiency (FD) in the maternal diets in offspring of broiler breeders during the laying period. In this project, broiler breeders will be used experimental animals and set up FD and normal folate (NF) model; and following experiments will be conducted: Study effects of FD in maternal diets on offspring fat traits and expression levels of genes involved in lipid, and search for key regulating genes in fat metabolism between the offspring of broiler breeders fed on FD diets and NF diets by microarray technology. Moreover, the partial key differential genes will be detected by real time RT-PCR and analyzed their DNA methylation levels by bisulfite sequencing. Finally, based on above experiment results, relationships of these differential expression genes, their DNA methylation levels and fat metabolism will be evaluate and analyzed. These results may provide some data for using falote in poultry.

叶酸是S-腺苷甲硫胺酸合成的甲基提供者，参与体内DNA合成和甲基化反应。肉种鸡产蛋期日粮叶酸缺乏对其后代脂肪代谢基因表达及表观遗传调控机制尚未见报道。本项目拟以肉种鸡为实验动物，构建叶酸缺乏模型，利用基因芯片、qRT-PCR、亚硫酸盐测序等技术，研究母源性叶酸缺乏对后代仔鸡脂肪性状和脂肪相关基因表达的影响，寻找叶酸对脂肪代谢起调控作用的差异基因，并对部分关键基因进行验证和甲基化分析。最后，综合分析基因表达和甲基化水平与脂肪代谢的关系，研究结果可望为家禽的叶酸营养应用提供理论参考。

母代叶酸缺乏可能影响动物后代某些基因的表达水平和甲基化状态，从而影响其正常生长发育。本项目构建了肉种鸡叶酸缺乏模型，测定了1、7、14、21、28、35和42日龄仔鸡叶酸正常组和叶酸缺乏组7个生长阶段的腹脂重等10项性状指标；利用ELISA方法检测了上述7个生长阶段血清中MTHFR、DHFR、Leptin、Hcy、folic acid (FA)和IGF-2激素的水平，以及腹脂中LPL和HSL的酶活性；测定了21日龄和42日龄仔鸡肝脏、左侧腿肌和左侧胸肌中的含脂率；利用透射电子显微镜观察了42日龄仔鸡肝脏亚显微结构。采用鸡数字表达谱芯片筛选出有功能注释的差异表达基因359个，其中下调基因214个，上调基因145个，并对差异表达基因进行了生物信息学分析；选取16个与叶酸代谢、甲基化等相关的差异表达基因，利用qRT-PCR方法验证了表达谱芯片结果；采用亚硫酸盐测序方法分析了ABCG2、MTHFD2和TYMS差异表达基因的甲基化水平。另外，采用qRT-PCR方法分析了FABPs、LPL、IGF-2、MTHFR、FBLN7和ABCG2基因mRNA在肝脏或腹脂组织中的表达水平。上述研究结果可为家禽营养学和鸡肉品质的改良提供科学依据。