本文選題：吉富羅非魚 + 硬脂酰輔酶A去飽和酶��； 參考：《南京農(nóng)業(yè)大學(xué)》2016年碩士論文

【摘要】：硬脂酰輔酶A去飽和酶(stearoyl-CoA desaturates,SCD)是合成單不飽和脂肪酸(monounsaturated fatty acid,MUFA)的限速酶,是調(diào)節(jié)肝臟脂肪生成和脂類氧化的關(guān)鍵控制點。經(jīng)聯(lián)合國糧農(nóng)組織(FAO)調(diào)查數(shù)據(jù)顯示,2014年全球100多個羅非魚養(yǎng)殖地區(qū)的年產(chǎn)量總和達5446800t,其中中國羅非魚產(chǎn)量在2014年增長至1450000t。由于羅非魚對低溫環(huán)境耐受力較差以及養(yǎng)殖過程中的不當投喂,使羅非魚產(chǎn)業(yè)受到重大的經(jīng)濟損失。吉富羅非魚是我國南方各省主要的淡水經(jīng)濟養(yǎng)殖魚類,研究吉富羅非魚的SCD基因?qū)τ谔岣呖沟蜏啬芰疤岣呱唐肤~的營養(yǎng)價值具有重要意義。本實驗是國內(nèi)首次從吉富羅非魚體內(nèi)克隆得到SCD基因的cDNA序列,同時對基因結(jié)構(gòu)和表達時序特征進行分析。并結(jié)合飼料中的脂肪酸組成和水溫對吉富羅非魚生長、生理、脂肪酸組成與SCD表達的影響,從營養(yǎng)、環(huán)境及分子水平較為深入研究了 SCD的表達調(diào)控網(wǎng)絡(luò)。為羅非魚的脂肪酸調(diào)控以及培育高品質(zhì)、高營養(yǎng)價值的羅非魚提供了一定的理論依據(jù)。1.吉富羅非魚SCD基因的克隆與序列分析本研究利用實時熒光定量PCR(real-time PCR,RT-PCR)和cDNA末端快速擴增法(rapid-amplificationof cDNAends,RACE)技術(shù)克隆獲得了吉富羅非魚的SCD基因的cDNA全長序列。結(jié)果顯示,SCD基因的cDNA總長為1333bp,由173bp5'非編碼區(qū)、152bp 3'非編碼區(qū)以及編碼335個氨基酸的1008bp開放閱讀框組成。與GenBank中,尼羅羅非魚、莫桑比克羅非魚相似度較高,基因序列同源性分別達99%和98%,氨基酸序列同源性分別為99%和97%;與其他魚類SCD基因序列相似度達83%以上,氨基酸序列同源性達79%以上。與其他魚類的SCD基因相似度很高,說明該基因在物種間高度保守。2.吉富羅非魚SCD基因組織及時空表達規(guī)律本研究通過RT-PCR檢測了 SCD基因在吉富羅非魚受精卵及胚胎發(fā)育階段(受精后 1h、12h、24h、48h、72h、96h)、仔稚魚階段(出膜后 1d、5d、10d、15d、20d、25d、30d)及成魚組織(血、肝臟、脾臟、腎臟、心臟、前腸、后腸、性腺、腦、皮膚、肌肉、鰓)中的表達。在胚胎發(fā)育的早期階段1h和12h時,SCD基因的表達量較高,24h時出現(xiàn)明顯下降的趨勢,48h時有所升高,隨后逐漸下降,96h最低。在受精卵及胚胎的早期階段(1-15d)時,SCD表達量較低,并呈現(xiàn)逐步上升的趨勢。20d時SCD的表達量顯著升高,隨后呈先下降后升高的趨勢。表明了羅非魚在發(fā)育各個階段所需的脂肪酸營養(yǎng)不同,從而導(dǎo)致脂肪酸代謝酶的表達水平也有所不同,為吉富羅非魚繁殖階段及魚苗培育階段的脂肪酸營養(yǎng)的提供了理論依據(jù)。分別對雄魚和雌魚組織的SCD表達量進行檢測,結(jié)果表明該基因在吉富羅非魚雄性和雌性的各組織表達量的高低有所不同,在肝臟中表達量均達到最高,其次在雄魚的精巢中表達量較高,而雌魚的肌肉中的表達量僅次于肝臟。在雌魚和雄魚的脾臟中表達量均為最低。證明了吉富羅非魚雌魚與雄魚的脂肪酸代謝機制具有一定差異,為進一步研究羅非魚UFA的合成途徑及調(diào)控機理提供了一定理論基礎(chǔ)。3.溫度對吉富羅非魚SCD基因表達的研究本實驗對吉富羅非魚在22℃、28℃和34℃三種不同養(yǎng)殖溫度下對SCD基因及肌肉脂肪酸相關(guān)性進行分析,同時對生長指標、血常規(guī)及血液生理指標進行了測定。28℃終體重顯著高于22和34℃(P0.05),FCR和PER均高于22和34℃。不同組間的肌肉脂肪酸組成具有顯著性差異(P0.05),SFA含量隨溫度的升高而降低而UFA含量呈相反的變化趨勢。22℃時羅非魚含較高的PUFA,尤其是n-3PUFA。SCD基因的表達及活性在低溫時顯著性升高(P0.05),并且增加吉富羅非魚SCD的表達及活性可提升肌肉MUFA的合成。此外,降低水溫可降低機體TC和TG的含量,對于提高吉富羅非魚抗低溫能力具有一定幫助。4.脂肪源對吉富羅非魚SCD基因表達的研究日糧中添加不同脂肪源對魚類的生長、生理等均具有一定影響,主要是由于飼料中脂肪酸組成不同,及不同魚類對脂肪酸利用不同引起的。本研究分別以椰子油(主要含SFA)、橄欖油(主要含MUFA)、魚油(脂肪酸比例均衡)及魚油+豆油(1:1)(主要含PUFA)為脂肪源,進行60d的養(yǎng)殖實驗。通過分析生長指標,發(fā)現(xiàn)飼料中添加脂肪酸比例較均衡的魚油組生長效果最好。添加不同脂肪源對血液和肝臟的生理指標具有一定影響,添加SFA會引起血液與肝臟中TC、TG含量上升,而添加PUFA可以有效地降低體脂的沉積。并且添加PUFA可以增強機體免疫力,提高抗氧化酶活性,降低氧化損傷。此外,飼料中不同脂肪酸組成對肌肉和肝臟脂肪酸的組成具有顯著性影響,魚體脂肪酸組成中的SFA及UFA分別于飼料脂肪酸組成的SFA及UFA呈正相關(guān)。并且飼料中高水平的PUFA會抑制SCD的表達與活性并降低體脂沉積而增加SFA會促進SCD的表達與活性。因此,均衡日糧中的脂肪酸組成,適當提高多不飽和脂肪酸含量有助于增加吉富羅非魚生長性能,降低血清脂肪沉積。
[Abstract]:The stearyl coenzyme A desaturase (stearoyl-CoA desaturates, SCD) is the speed limiting enzyme for the synthesis of monounsaturated fatty acids (monounsaturated fatty acid, MUFA). It is the key control point for regulating liver adipose formation and lipid oxidation. The annual output of more than 100 global tilapia in 2014 by the United Nations Food and agriculture Organization (FAO) survey data The total amount of Chinese tilapia increased to 5446800t in 2014 to 1450000t., due to the poor tolerance of tilapia to low temperature environment and the improper feeding in the breeding process, the tilapia industry suffered major economic losses. The gene is of great significance to improve the ability to resist low temperature and to improve the nutritional value of commercial fish. This experiment is the first time that the cDNA sequence of SCD gene has been cloned from the fishes of fishes in China, and the structure of the gene and the temporal characteristics of the expression are analyzed. The effect of fatty acid composition and SCD expression, the expression regulation network of SCD was studied in depth from nutrition, environment and molecular level. It provides a certain theoretical basis for the regulation of fatty acids and the cultivation of high nutritional value of tilapia and high nutritive value of tilapia. The cloning and sequence analysis of the SCD gene of Gefu tilapia,.1., is used in real time. PCR (real-time PCR, RT-PCR) and cDNA terminal rapid amplification (rapid-amplificationof cDNAends, RACE) were cloned to obtain the cDNA full length of the SCD gene of June tilapia. The results showed that the cDNA total length of the SCD gene was that of the non coding region, the non coding region and the encoding of the 335 amino acids. In GenBank, the similarity of Nile tilapia and Mozambique tilapia is higher, the homology of the gene sequence is 99% and 98% respectively, the homology of amino acid sequence is 99% and 97%, and the similarity of the SCD gene sequence of other fishes is more than 83% and the homology of the amino acid sequence is above 79%. The similarity of the SCD gene of other fishes is very similar. SCD gene tissue and spatio-temporal expression of.2. Junfu tilapia were highly conserved between species. The SCD gene was detected by RT-PCR in the fertilized egg and embryonic development stage of tilapia (1H, 12h, 24h, 48h, 72h, 96h). The expression of spleen, kidney, heart, foregut, hindgut, hindgut, gonadal, brain, skin, muscle, gill. At the early stage of embryonic development 1H and 12h, the expression of SCD gene is higher, the trend of 24h decreases obviously, 48h increases, and then gradually decreases, and 96h is the lowest. The expression of SCD is low in the early stage of fertilized eggs and embryos (1-15d). The expression of SCD increased significantly at.20d, and then decreased and then increased. It showed that the fatty acid nutrition of the tilapia at various stages of development was different, and the expression level of fatty acid metabolizing enzymes was also different, which was the fatty acid nutrition in the breeding stage and the cultivation stage of the fish. A theoretical basis was provided. The expression of SCD expression in male and female tissues was detected. The results showed that the expression of the gene in the male and female tissues of the male and female was different, the expression in the liver was the highest, followed by the high amount of the sperm in the male's spermary, and the amount of expression in the female's muscles was only the only time. The expression of the spleen in the female and male was the lowest. It was proved that the metabolic mechanism of fatty acid in the female and male fish was different, which provided a theoretical basis for the further study of the synthesis and regulation mechanism of UFA, and the study on the expression of Ji Fuluo non fish SCD gene by.3. temperature was a study of the experiment. The correlation between SCD gene and muscle fatty acids was analyzed at 22, 28 and 34 degrees centigrade. At the same time, the growth index, blood routine and blood physiological index were determined to be higher than 22 and 34 centigrade (P0.05), FCR and PER were higher than 22 and 34. The composition of fatty acid between different groups was significant. P0.05, the content of SFA decreased with the increase of temperature, while the UFA content was in the opposite direction. The higher PUFA in the tilapia at.22 C, especially the expression and activity of n-3PUFA.SCD gene increased significantly at low temperature (P0.05), and the expression and activity of SCD of the fish of Gefu tilapia could increase the synthesis of muscle MUFA. In addition, the water decreased water. Temperature can reduce the content of TC and TG. It has certain help to improve the anti low temperature ability of the fishes of Gefu tilapia, which can help the.4. fat source to express the SCD gene of GF riloticus. The diet of different fats has a certain effect on the growth and physiology of fish. It is mainly due to the difference of fatty acid composition in the feed and the fat of different fish. In this study, we used coconut oil (mainly SFA), olive oil (mainly containing MUFA), fish oil (fatty acid proportion equilibrium) and fish oil + bean oil (1:1) as the fat source, and carried out the culture experiment of 60d. By analyzing the growth index, it was found that the growth effect of fatty acid in the feed was the best. Adding different fat sources has a certain effect on the physiological indexes of blood and liver. Adding SFA can cause the increase of TC and TG in the blood and liver, and the addition of PUFA can effectively reduce the deposition of body fat. And adding PUFA can enhance the immunity of the body, improve the activity of antioxidant enzymes and reduce the oxidative damage. In addition, the composition of different fatty acids in the feed is made up. The composition of fatty acids in the muscle and liver has a significant effect. The SFA and UFA in the fatty acid composition of the fish body are positively related to the SFA and UFA of the fatty acids in the feed. And the high level PUFA in the feed inhibits the expression and activity of SCD and reduces the body fat deposition, and increases the expression and activity of SCD by increasing the SFA. Therefore, the balanced diet is balanced in the diet. Fatty acid composition and proper increase of polyunsaturated fatty acids are beneficial to increase growth performance and reduce serum fat deposition.

【學(xué)位授予單位】：南京農(nóng)業(yè)大學(xué)
【學(xué)位級別】：碩士
【學(xué)位授予年份】：2016
【分類號】：S917.4

【參考文獻】

相關(guān)期刊論文 前10條

1 謝帝芝;陳芳;張慶昊;陳軍亮;董燁瑋;王樹啟;游翠紅;聶國興;李遠友;;魚類LC-PUFA合成代謝調(diào)控機制研究進展[J];汕頭大學(xué)學(xué)報(自然科學(xué)版);2015年02期

2 李思萌;吳立新;姜志強;陳煒;;飼料脂肪源對大菱鲆幼魚生長性能和肌肉脂肪酸組成的影響[J];動物營養(yǎng)學(xué)報;2015年05期

3 王健;張軍;段修軍;張宜輝;張蕊;董飚;龔道清;;番鴨硬酯酰輔酶A去飽和酶1基因克隆、序列分析及填飼對其表達的影響[J];上海交通大學(xué)學(xué)報(農(nóng)業(yè)科學(xué)版);2014年03期

4 魏廣蓮;徐鋼春;顧若波;徐跑;;飼料中添加不飽和脂肪酸對刀鱭幼魚脂肪代謝酶活性和肌肉成分的影響[J];動物營養(yǎng)學(xué)報;2014年01期

5 崔素麗;劉波;徐跑;謝駿;萬金娟;周明;;兩種溫度下大黃素對團頭魴生長、血液指標及肝臟HSP70 mRNA表達的影響[J];水生生物學(xué)報;2013年05期

6 張蕊;張宜輝;邵丹;王來娣;龔道清;;硬脂酰輔酶A去飽和酶基因的功能與調(diào)控[J];生命科學(xué);2013年04期

7 陳海燕;朱邦科;樊啟學(xué);胡培培;宋林;彭聰;宗克金;張云龍;;不同脂肪源對溼幼魚生長、血清生化組成和肝臟的影響[J];華中農(nóng)業(yè)大學(xué)學(xué)報;2013年02期

8 張靜靜;宋玉芹;李楨;朱艷菲;張廷榮;;中國西門塔爾牛SCD1基因多態(tài)性與肉質(zhì)性狀的相關(guān)分析[J];華北農(nóng)學(xué)報;2013年01期

9 施兆鴻;岳彥峰;彭士明;李云航;孫鵬;尹飛;王建鋼;;飼料脂肪水平對褐菖湁血清生化指標、免疫及抗氧化酶活力的影響[J];中國水產(chǎn)科學(xué);2013年01期

10 王亞超;鄧俊良;王利民;錢輝;劉國文;王哲;;胰島素、胰高血糖素和神經(jīng)肽Y對體外培養(yǎng)犢牛肝細胞硬脂酰CoA去飽和酶mRNA表達的影響[J];中國獸醫(yī)學(xué)報;2013年01期

相關(guān)博士學(xué)位論文 前1條

1 王建楓;鋅對大鼠肝臟硬脂酰輔酶A去飽和酶-1（SCD-1）的影響及其調(diào)控機理研究[D];浙江大學(xué);2008年

相關(guān)碩士學(xué)位論文 前9條

1 馬明華;曼氏無針烏賊SCD基因克隆與生物信息學(xué)分析[D];浙江海洋學(xué)院;2013年

2 朱才業(yè);克隆徐淮山羊SCD1基因及轉(zhuǎn)基因小鼠和轉(zhuǎn)基因綿羊的制備[D];揚州大學(xué);2013年

3 王金鳳;嶗山奶山羊ACACA和SCD基因多態(tài)性與泌乳性狀的關(guān)聯(lián)分析[D];山東農(nóng)業(yè)大學(xué);2011年

4 王煜恒;不同脂肪源對異育銀鯽生長、體脂沉積和血液生化指標的影響[D];南京農(nóng)業(yè)大學(xué);2011年

5 付曉英;鵝SCD1基因全長cDNA的克隆、組織表達及填飼對其表達的影響[D];四川農(nóng)業(yè)大學(xué);2010年

6 陳忠琦;奶山羊SCD基因的克隆、多態(tài)性檢測及其與經(jīng)濟性狀的關(guān)聯(lián)分析[D];西北農(nóng)林科技大學(xué);2010年

7 朱越;腺病毒介導(dǎo)的山羊乳腺硬脂酰輔酶A去飽和酶（SCD）基因的過表達研究[D];西北農(nóng)林科技大學(xué);2010年

8 唐威;不同脂肪源在斑點叉尾洶飼料中應(yīng)用的研究[D];湖南農(nóng)業(yè)大學(xué);2010年

9 張瑞超;日糧中添加亞油酸和維生素A對乳腺組織和肌肉組織中硬脂酰輔酶A去飽和酶的影響[D];新疆農(nóng)業(yè)大學(xué);2006年

，

本文編號：1835257