下丘脑关键miRNA对肉鸭剩余采食量的调控作用及分子机制

Residual feed intake (RFI) is an important evaluation index for feed efficiency (FE) of meat ducks. Previous studies indicated that several key miRNAs were related to RFI in some economic animals including cattle, pig and chicken, but the molecular mechanism of miRNA regulating RFI in meat-type ducks is not yet clear. In this project, we will select high RFI (HRFI) and low RFI (LRFI) F2 ducks based on our previous study as experimental materials. The differential expressions of miRNA will be detected by small RNA sequencing (sRNA-seq). Bioinformatics analysis will be used to predict the target genes of miRNAs and to construct a gene regulatory network. Combining with the genetic variation in mRNA 3’UTR of functional genes from our previous GWAS study, dual-luciferase reporter assay system will be used to validate the relationship between key miRNAs and genes. The effect of overexpression and knockdown of the key genes on growth of hypothalamic neurons and myoblasts will be verified in vitro. Feed intake and body weight will be measured after key miRNA analogues injection in vivo. A multi-level research path, including bioinfomatics, molecular, in vitro and in vivo, will be implemented to explore the molecular regulation mechanism of RFI in ducks. This research will aim at providing a theoretical basis for the study of FE in ducks, and promoting the development and application of molecular breeding of the small-sized high quality meat-type ducks.

剩余采食量（RFI）是评价肉鸭饲料利用率的重要指标。已有研究表明，miRNA参与调控牛、猪、鸡等动物的RFI，然而其参与调控肉鸭RFI的分子机理尚不清楚。本项目以F2资源群体中的高、低RFI肉鸭为试验素材，对下丘脑组织进行small RNA测序，分析鉴定差异表达的miRNA，利用生物信息学分析预测miRNA靶基因并构建基因调控网络图；结合前期GWAS获取的RFI功能基因3’UTR遗传变异信息，利用双荧光素酶报告系统确定RFI关键miRNA与功能基因靶向关系，通过过表达和敲低关键靶基因，验证其对下丘脑神经元细胞和成肌细胞生长的作用；同时结合体内注射关键miRNA类似物后的采食量和体重测定结果，从生物信息学、分子、细胞和活体等多层次水平深入系统分析肉鸭RFI调控的分子机理，为肉鸭饲料利用率改良与高效生产提供新思路，为小体型优质肉鸭的分子选育工作提供理论依据。

饲料成本约占肉鸭生产总成本的60%以上，饲料利用率低是我国优质肉鸭产业发展面临的“卡脖子”问题，提高饲料利用率是提高优质肉鸭经济效益的重要手段。本项目首先针对小体型优质肉鸭种质资源进行评价，研究表明小体型肉鸭品质优良、营养价值高，但其生长速度慢、饲料转化率低，严重阻碍了其开发利用和产业发展；随后针对小体型肉鸭不同饲养模式进行研究，结果发现笼养模式可以提高肉鸭的生产性能和部分肉质性状，适用于小体型肉鸭的生产；随后在笼养模式下进行高低剩余采食量（RFI）小体型肉鸭生产性能比较研究，结果发现低剩余采食量（LRFI）组肉鸭的FCR和RFI显著低于高剩余采食量（HRFI）组肉鸭，然而上市体重、屠宰性能、肉质性状等方面无显著差异。因此，对低RFI个体选育可以在不影响小体型肉鸭生产性能和肉品质的前提下显著提高饲料利用率而降低生产成本；随后针对上述高低RFI肉鸭下丘脑组织和盲肠内容物分别进行small RNA测序和16S rRNA测序，针对前期F2资源群体中的个体进行全基因组关联分析。本项目通过转录组、基因组和微生物组进行多组学分析，挖掘影响RFI性状的候选基因、分子标记和肠道菌群，从生物信息学、分子生物学、细胞生物学等多层次水平系统分析肉鸭RFI调控的分子机理，初步明确了miR-182通过抑制DDC基因表达参与肉鸭下丘脑对RFI调节的分子调控途径，为提高肉鸭饲料利用率提供新思路，为小体型肉鸭的分子选育工作提供理论依据。