本文選題：中國荷斯坦奶牛 + 脂肪酸��； 參考：《揚州大學(xué)》2017年碩士論文

【摘要】：本課題以中國荷斯坦奶牛為試驗對象,測定分析奶牛乳中主要成分和脂肪酸比例的變化規(guī)律,采用轉(zhuǎn)錄組測序技術(shù)和生物信息學(xué)方法,研究乳腺組織在泌乳初期(圍產(chǎn)期的泌乳階段)和泌乳盛期這兩個關(guān)鍵時期,其分泌細(xì)胞從血液中獲取脂肪酸(FA import into cells)、脂肪酸從頭合成(Denovo synthesis)、外源固醇轉(zhuǎn)運(Xenobiotic and Cholesterol transport)、脂肪酸激活及胞內(nèi)運送(Acetate and FA activation and intra-cellular transport)、脂肪酸細(xì)胞內(nèi)合成和去飽和(FAsynthesis and desaturation)至脂滴形成和分泌(Lipid droplet formation and secretion)等脂類代謝過程的轉(zhuǎn)錄及轉(zhuǎn)錄后調(diào)控機制,根據(jù)轉(zhuǎn)錄組測序結(jié)果,篩選泌乳早期差異表達(dá)的基因。為厘清奶牛乳腺脂類合成、代謝機制,實現(xiàn)乳成分的定向遺傳調(diào)控,生產(chǎn)優(yōu)質(zhì)原料奶奠定理論基礎(chǔ)。在揚州大學(xué)實驗農(nóng)牧場中分別挑選兩組健康的中國荷斯坦奶牛,第一組30頭(2-3胎次),用于牛奶中乳成分和脂肪酸的測定,第二組3頭(2-3胎次),用于牛奶中乳成分和脂肪酸的測定和乳腺組織的采集。33頭奶牛均采用TMR日糧飼養(yǎng),整個研究期間日糧及飼養(yǎng)管理條件保持相對一致。測定3(初乳)、30、90 d奶牛乳中乳脂率、乳蛋白、乳糖、非脂乳固體和脂肪酸的百分比含量。微創(chuàng)法采集-7(產(chǎn)前)、30、90 d第二組奶牛乳腺組織,利用RNA-seq技術(shù)對試驗牛乳腺組織進(jìn)行深度轉(zhuǎn)錄組測序。主要結(jié)果如下:(1)本實驗利用氣相色譜法在牛奶中共檢測出35種脂肪酸(共41類),對含量高于1%的 18 類顯著差異的脂肪酸 C4:0、C6:0、C8:0、C10:0、C12:0、C14:0、C15:0、C16:0、C16:1,cis-9、C18:0、C18:1,cis-9、C18:2,cis-9,12、SFA、MUFA、PUFA、sc-FA、mc-FA和lc-FA著重研究,結(jié)果顯示:除去C16:1,cis-9的17類脂肪酸的含量在泌乳3 d和30 d存在顯著差異(P0.05);除去C4:0、C6:0和sc-FA的15類脂肪酸的含量在泌乳30 d和90 d差異不顯著(P0.05);除去C4:0和sc-FA的16類脂肪酸的含量在泌乳3 d和90 d差異顯著(P0.05)。(2)對不同時期乳腺組織進(jìn)行轉(zhuǎn)錄組(RNA-seq)測序,新轉(zhuǎn)錄本預(yù)測分析結(jié)果:在不同時期乳腺組織中共預(yù)測得到8706個新轉(zhuǎn)錄本,其中預(yù)測具有編碼能力的有3253個。已知基因結(jié)構(gòu)優(yōu)化結(jié)果:在不同時期乳腺組織中共優(yōu)化了 1664個已知基因結(jié)構(gòu),大部分為基因起始位置到終止位置序列的延長。差異表達(dá)分析結(jié)果如下:-7vs30乳腺組織中有2421個差異基因;30vs90乳腺組織中有33個差異基因。(3)通過對差異表達(dá)基因進(jìn)行KEGG富集分析,共篩選到13條與脂肪酸代謝相關(guān)通路,包括PPAR信號通路(PPAR signaling pathway)、甘油脂類新陳代謝信號通路(Glycerolipid metabolism signaling pathway)、脂肪酸代謝信號通路(Fatty acid metabolism signaling pathway)和不飽和脂肪酸的生物合成信號通路(Biosynthesis of unsaturated fatty acids signaling pathway)等。(4)對通路中與脂肪酸合成代謝相關(guān)的30個基因進(jìn)行分析,結(jié)果顯示:CD36、ABCA1、SLC27A6、ACSL1、ACSS1、ACSS2、FASN、AGPAT6、DGAT2、GPAM、GPAT2、LPIN1和SOCS3基因在泌乳啟動期間表達(dá)量存在顯著差異(P0.05);而SOCS3、FABP4、GPAT2、DGAT1和DGAT2基因在泌乳終止期間存在顯著差異(P0.05)。把這些基因與奶牛乳中脂肪酸成分之間進(jìn)行相關(guān)性分析,結(jié)果顯示:除去FADS2、GPAT2、SREBF1和SREBF2外,其他基因均與所測脂肪酸中的一類或幾類存在顯著或極顯著相關(guān)。本研究發(fā)現(xiàn),CD36、ABCA1、SLC27A6、ACSL1、ACSS1、ACSS2、FASN、AGPAT6、DGAT2、GPAM、GPAT2、LPIN1和SOCS3基因在泌乳的啟動和維持過程中起著重要的作用。PPAR信號通路、甘油脂類新陳代謝信號通路、脂肪酸代謝信號通路和不飽和脂肪酸的生物合成信號通路等多條與脂肪酸代謝相關(guān)通路中富集的基因,可作為深入研究乳腺泌乳機制的候選基因群。
[Abstract]:In this study, Chinese Holstein cows were used to test and analyze the changes in the proportion of main ingredients and fatty acids in milk of dairy cows. The two key periods of mammary tissue in the early lactation (perinatal lactation stage) and lactation period were studied by the sequence of transcriptional sequencing and bioinformatics, and the secretory cells were obtained from the blood. Fatty acids (FA import into cells), fatty acids from ab initio synthesis (Denovo synthesis), exogenous steroid transport (Xenobiotic and Cholesterol transport), fatty acid activation and intracellular delivery (Acetate and), fatty acids in fine cell synthesis and desaturation to lipid droplets The transcriptional and post transcriptional regulation mechanism of lipid metabolism processes such as Lipid droplet formation and secretion and other lipid metabolism processes are selected to screen the genes of differential expression in early lactation, to clarify the lipid synthesis and metabolic mechanism of dairy cows, to realize the steady genetic regulation of milk ingredients and to lay a theoretical foundation for the production of high quality raw milk. In the experimental farm of Yangzhou University, two groups of healthy Chinese Holstein cows were selected. The first group of 30 heads (2-3 parity) was used for the determination of milk ingredients and fatty acids in milk, second groups 3 heads (2-3 parity). The milk ingredients and fatty acids in milk and the mammary gland tissues were collected by TMR diet, and the whole study was carried out. The dietary and feeding management conditions were consistent during the period. The percentage of milk fat, milk protein, lactose, non fat milk solid and fatty acid in milk of 30,90 D dairy cows was measured in 3 (colostrum), milk protein, lactose, non fat milk solid and fatty acid. -7 (prenatal), 30,90 d second group of dairy cow mammary tissues were collected by minimally invasive method, and the deep transcriptional group of bovine breast tissue was sequenced by RNA-seq technology. The results are as follows: (1) in this experiment, 35 fatty acids (41 kinds of fatty acids) were detected by gas chromatography in the milk, and the fatty acids C4:0, C6:0, C8:0, C10:0, C12:0, C14:0, C15:0, C16:0, C16:1, cis-9, and C18:0 were studied. There was a significant difference in the content of 17 kinds of fatty acids in C16:1 and cis-9 in lactation 3 D and 30 d (P0.05); except C4:0, the 15 fatty acids of C6:0 and sc-FA were not significant in lactating 30 d and 90 d (P0.05), and the content of 16 types of fatty acids removed from C4:0 and 16 was significant in lactating 3 and 90. RNA-seq sequencing and new transcriptional prediction analysis: 8706 new transcripts were predicted in different stages of breast tissue, of which 3253 were predicted to have the ability to encode. The optimization results of the known gene structure: 1664 known gene structures were optimized in different mammary tissues at different stages, most of which were from the beginning of the gene to the end. Extension of position sequence. The results of differential expression analysis are as follows: there are 2421 differentially differentially genes in -7vs30 breast tissue; there are 33 differential genes in 30vs90 breast tissue. (3) through KEGG enrichment analysis of differentially expressed genes, 13 pathways related to fatty acid metabolism, including PPAR signaling pathway (PPAR signaling pathway), and triglycerides, were screened. Glycerolipid metabolism signaling pathway, fatty acid metabolism signaling pathway (Fatty acid metabolism signaling pathway) and biosynthetic signaling pathway of unsaturated fatty acids (Biosynthesis of unsaturated). (4) 3 of the pathways associated with fatty acid synthesis and metabolism in the pathway 0 genes were analyzed, and the results showed that CD36, ABCA1, SLC27A6, ACSL1, ACSS1, ACSS2, FASN, AGPAT6, DGAT2, GPAM, GPAT2, there were significant differences in the expression of LPIN1 and genes during lactation initiation. The correlation analysis between fatty acids showed that the other genes were significantly or highly correlated with a class or a few classes in the fatty acids that were removed from FADS2, GPAT2, SREBF1 and SREBF2. This study found that CD36, ABCA1, SLC27A6, ACSL1, ACSS1, ACSS2, FASN, AGPAT6, AGPAT6, The maintenance process plays an important role in the.PPAR signaling pathway, the metabolic signaling pathway of triglycerides, the fatty acid metabolism signaling pathway and the biosynthetic pathway of unsaturated fatty acids, which are enriched in the fatty acid metabolism pathway, which can be used as a candidate gene group for the deep study of mammary gland lactation mechanism.

【學(xué)位授予單位】：揚州大學(xué)
【學(xué)位級別】：碩士
【學(xué)位授予年份】：2017
【分類號】：S823;TS252.2

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