全转录组SNP和miRNA调控奶水牛泌乳性能的作用机制

Mammary gland is the oragn of milk production in mammals that is of prime importance to milk yield and quality. Previously, the gland transcriptomes of Murrah buffalo at the lactation and gestation were performed using RNA-seq. Based on this, this project will mainly screen the differentially expressed genes (DEGs), microRNA(miRNA) and single nuclease polymorphims (SNPs) using the bioinformatics analysis. And then the genome-whole association analysis (GWAS) will be carried out using the designed SNP array in the milk buffalo population. Finaly, functional verification test for the above results are conducted by some methods of RNAi, fluorescent reproter vector test and over expression vector test, etc. Therefore, the objective of the project is to identify some new genes (transcripts), miRNA and SNPs that have significantly effects on the milk production traits in buffaloes, clarify the interaction among those and the regulation mechaism of milk performance, and reveal the molecular genetic basis of the milk performance in buffaloes. This findings will provide fundamental basis for improving milk performance and genetic improvement in milk buffalo.

乳腺是泌乳动物的生产器官，是多产奶和产好奶的必要条件。本项目拟在先前开展摩拉水牛妊娠期和泌乳期乳腺RNA测序（转录组测序和miRNA测序）研究的基础上，利用生物信息学分析重点筛查差异表达的基因、miRNA和转录组SNP，并通过定制的SNP芯片对奶水牛群体开展全转录组SNP关联分析研究，最后利用RNAi、荧光报告载体和过表达载体等方法对上述结果开展功能验证研究。本项目的研究旨在鉴定显著影响奶水牛泌乳相关的新基因（转录本）、miRNA及转录组SNP，阐明它们之间的互作关系及调控泌乳性能的作用机制，以揭示奶水牛泌乳性能的分子遗传基础，为提高泌乳水牛的产奶性能以及遗传改良提供理论基础。

乳腺是泌乳动物的生产器官，是多产奶和产好奶的必要条件。发掘水牛产奶性状相关基因对于揭示乳腺发育和泌乳的遗传机制，进而改良该畜种的性状至关重要。本项目整合了全基因组关联分析和转录组测序进行了水牛产奶性状相关基因的发掘。同时整合转录组数据和miRNA测序数据进行了泌乳相关miRNA的挖掘及其靶基因调控互作网络的构建，并结合功能验证试验初步阐明了miRNA对水牛泌乳功能的调控机制。取得了以下研究结果：1）利用乳腺组织转录组测序、全基因组关联分析和加权的基因共表达网络分析（WGCNA）等技术手段鉴定出12个基因（BNIPL, TUBA1C, C2CD4B, DCP1B, MAP3K5, PDCD11, SRGAP1, GDPD5, BARX2, SCARA3, CTU2和RPL27A）作为枢纽基因，涉及产奶相关的多种信号通路；2）基于miRNA-seq技术在妊娠期、泌乳期和干乳期三个乳腺组织文库中鉴定出432个成熟miRNA和189个novel-miRNA。经两两比对分析发现了176个差异表达的miRNA。结合生物信息和RT-PCR技术鉴定出25个与乳腺发育和泌乳相关的差异表达miRNA及其调控网络；3）利用双荧光素酶报告载体试验、干扰和过表达等技术手段证实了MAPK1基因为miR-183靶基因，通过调控内源靶基因MAPK1的相对表达水平，进而影响到乳腺上皮细胞的增殖，乳脂及乳蛋白的分泌，初步阐明了miRNA调控水牛泌乳性能的作用机制。