蔡 曼 柳延濤 王 娟 孔憲輝 王旭文 李衛(wèi)華 余 渝 劉 麗

(新疆農(nóng)墾科學(xué)院棉花研究所；農(nóng)業(yè)部西北內(nèi)陸區(qū)棉花生物學(xué)與遺傳育種重點(diǎn)實(shí)驗(yàn)室；新疆兵團(tuán)棉花遺傳改良與高產(chǎn)栽培重點(diǎn)實(shí)驗(yàn)室1，石河子 832000)(石河子大學(xué)；新疆兵團(tuán)綠洲生態(tài)農(nóng)業(yè)重點(diǎn)實(shí)驗(yàn)室2，石河子 832003)(新疆農(nóng)墾科學(xué)院作物研究所3，石河子 832000)

植物體的脂肪酸代謝是維系其生命活動(dòng)的基本代謝之一，也為人類提供了重要的能量來源。由于人體不能自身合成一些必需的脂肪酸，從植物種子中提取植物油食用便成為必需脂肪酸攝入的主要方式之一。植物種子中儲(chǔ)存的脂肪酸常以三酰甘油酯(Triglyceride acyl groups,TAGs)(即在甘油骨架上附連3個(gè)脂肪酸)的形式存在。油脂是植物種子儲(chǔ)存能量的主要形式，為種子萌發(fā)和幼苗前期生長(zhǎng)提供必不可少的能量來源。除食用外，脂肪酸在工業(yè)生產(chǎn)中也有著重要的價(jià)值，如可作為生產(chǎn)油漆、表面活性劑、潤(rùn)滑劑以及尼龍、醫(yī)藥等原料。因此，改良植物種子中脂肪酸含量及組分以滿足食用及工業(yè)需要的研究工作，越來越得到人們的重視。培育含油量更高、不飽和脂肪酸比例更健康的油料作物新品種是作物育種的任務(wù)之一。

植物脂類代謝途徑非常復(fù)雜。首先在種子的發(fā)育過程中，蔗糖作為合成脂肪酸的主要碳源，從光合作用的器官(如葉片)轉(zhuǎn)運(yùn)到種子細(xì)胞中，通過植物糖酵解產(chǎn)生大量的三酰甘油合成前體，如磷酸二羥丙酮、丙酮酸。丙酮酸經(jīng)過氧化脫羧形成乙酰-CoA，它運(yùn)送到質(zhì)體中進(jìn)行脂肪酸的從頭合成，合成的脂肪酸再運(yùn)送到內(nèi)質(zhì)網(wǎng)與3-磷酸甘油組裝形成三酰甘油，合成的三酰甘油最后運(yùn)輸?shù)接腕w中進(jìn)行儲(chǔ)存。這個(gè)復(fù)雜的生理生化過程受各種功能酶和轉(zhuǎn)錄因子的調(diào)控，這些功能酶和轉(zhuǎn)錄因子在擬南芥[1]、油菜[2-4]、大豆[5]、玉米[6]、麻瘋樹[7]、棉花[8]等作物中均有報(bào)道，其中ACCase、DGAT、FAS等關(guān)鍵酶基因及轉(zhuǎn)錄因子LEC、WRI、Dof等都是研究的熱點(diǎn)。本文綜述了植物種子油脂合成代謝中參與調(diào)控該途徑的關(guān)鍵酶和轉(zhuǎn)錄因子的研究進(jìn)展，為開展植物油脂改良提供借鑒。

1 油脂合成途徑

1.1 脂肪酸的生物合成

脂肪酸是油脂的主要組成成分，其合成主要在質(zhì)體中進(jìn)行，首先由乙酰CoA羧化酶(Acetyl CoA carboxylase,ACCase)催化乙酰CoA形成脂肪酸鏈的二碳單位的直接供體—丙二酸單酰CoA(Malonyl CoA,Mal-CoA)。再由脂肪酸合酶(Fatty acid synthase,FAS)系統(tǒng)，經(jīng)過縮合、還原、脫水、再還原的過程進(jìn)行碳鏈的延伸(圖1)。FAS系統(tǒng)是一個(gè)多酶復(fù)合體，包括?；d體蛋白(Acyl carrier protein,ACP)和6種酶，所有的催化反應(yīng)均在ACP上進(jìn)行。FAS催化連續(xù)循環(huán)的聚合反應(yīng)，每次循環(huán)增加2個(gè)碳的?；兼?，直至合成含有ACP的飽和脂肪酸棕櫚酸(16:0-ACP)和硬脂酸(18:0-ACP)，在去飽和酶的作用下形成不飽和脂肪酸，其中包括棕櫚油酸和油酸等單不飽和脂肪酸及亞油酸和亞麻酸等長(zhǎng)鏈多聚不飽和脂肪酸。然后，在?；?ACP硫酯酶(acyl-ACP thioesterase，F(xiàn)AT)的催化下，將脂肪酸從ACP上釋放出來。依據(jù)作用的底物不同，可將FAT分為FATA和FATB，它們有具有碳鏈長(zhǎng)度特異性[12-13]，并且其活性影響著脂肪酸的組成。

圖1 脂肪酸從頭合成

1.2 三酰甘油的合成

從ACP上釋放的游離脂肪酸在長(zhǎng)鏈脂酰輔酶A合成酶(Long-chain acyl-CoA synthetase,LACS)的作用下形成脂酰-CoA，LACS位于質(zhì)體外膜，它催化形成的脂酰CoA是三酰甘油合成的底物。將三酰甘油的合成前體脂酰CoA和3-磷酸甘油運(yùn)輸?shù)絻?nèi)質(zhì)網(wǎng)中進(jìn)行組裝[14-15]。參與脂酰CoA從質(zhì)體運(yùn)輸?shù)絻?nèi)質(zhì)網(wǎng)的轉(zhuǎn)運(yùn)蛋白研究的還不是很清楚，目前對(duì)其運(yùn)輸?shù)臋C(jī)制有幾個(gè)推測(cè)。其一，通過磷脂酰膽堿(Phosphatidyl cholines,PC)將合成的脂肪酸從質(zhì)體運(yùn)輸?shù)絻?nèi)質(zhì)網(wǎng)，其作用機(jī)制可能是在質(zhì)體膜上，由LPCAT催化將新合成的脂肪酸合成到PC上，然后通過PC實(shí)現(xiàn)從質(zhì)體到內(nèi)質(zhì)網(wǎng)的運(yùn)輸[16]。其二，有報(bào)道ABC(ATPbinding cassette)轉(zhuǎn)運(yùn)蛋白也可以實(shí)現(xiàn)該轉(zhuǎn)運(yùn)過程[17]。擬南芥有12個(gè)ABCA家族(ABC家族的一個(gè)亞家族)成員，它們的功能和作用機(jī)制都還不清楚，有待進(jìn)一步研究。

2 油脂合成代謝中的關(guān)鍵酶

2.1 乙酰CoA羧化酶

乙酰-CoA羧化酶(Acetyl CoA carboxylase,ACCase)是脂肪酸生物合成的關(guān)鍵酶之一。植物種子中ACCase催化乙酰CoA羧化形成丙二酸單酰CoA，是脂肪酸合成以及油脂形成的關(guān)鍵調(diào)控步驟，而且ACCase也是影響植物整個(gè)生命過程的重要基因[18-19]。

ACCase有異質(zhì)型和同質(zhì)型2種類型。在雙子葉植物和非禾本科單子葉植物的質(zhì)體中，ACCase為異質(zhì)型，由4個(gè)亞基構(gòu)成[20]：生物素羧基載體蛋白(BCCP)、生物素羧化酶(BC)、羧基轉(zhuǎn)移酶的2個(gè)亞基α-CT和β-CT。同質(zhì)型ACCase存在于植物細(xì)胞質(zhì)中，其每個(gè)亞基兼具ACCase的所有催化功能，但只有當(dāng)它們聚合成完整的酶后才有活性[21]；還存在于藻類、酵母、動(dòng)物及部分植物的胞質(zhì)中[22]。在大多數(shù)高等植物中，同質(zhì)型ACCase催化產(chǎn)生的丙二酸單酰CoA用于脂肪酸鏈的延伸和類黃酮等許多次生代謝產(chǎn)物的合成，而異質(zhì)型則用于脂肪酸的從頭生物合成[23]。

研究表明，植物種子脂肪酸的合成速率和油脂的積累同ACCase的活性密切相關(guān)性。如在大豆油脂形成的種子發(fā)育早期到中期，高油大豆ACCase的活性是低油大豆的2倍[24]；擬南芥低含油量突變體wrinkled1ACCase的表達(dá)量明顯小于野生型[25]。在油菜中，將油菜種子特異表達(dá)啟動(dòng)子與擬南芥同質(zhì)型ACCase基因融合，然后在大豆轉(zhuǎn)移肽的轉(zhuǎn)運(yùn)下，將ACCase導(dǎo)入油菜葉綠體，獲得的轉(zhuǎn)基因油菜成熟種子的ACCase活性比對(duì)照提高10～20倍，并且含油量增加5%[26]。由于異質(zhì)型比同質(zhì)型的結(jié)構(gòu)更為復(fù)雜，所以對(duì)異質(zhì)型的研究也相對(duì)較少。研究發(fā)現(xiàn)，蓖麻種子發(fā)育過程中，ACCase反應(yīng)活性及BC和BCCP的表達(dá)量與油脂積累存在著一定的對(duì)應(yīng)關(guān)系[27]。將羧基轉(zhuǎn)移酶β亞基(accD)在各種組織的質(zhì)體中過量表達(dá)導(dǎo)致轉(zhuǎn)基因植株葉片脂肪酸含量增加，植株葉片明顯增長(zhǎng)，雖然轉(zhuǎn)基因后代種子的脂肪酸含量與野生型沒有顯著變化，但種子產(chǎn)量提高近2倍，從而提高單株種子的含油量[28]。

2.2 脂肪酸合成酶

脂肪酸合成酶(Fatty acid synthesis，F(xiàn)AS)是一個(gè)多酶復(fù)合體。FAS主要由6種酶構(gòu)成：乙酰CoA-ACP轉(zhuǎn)移酶、丙二酸單酰CoA-ACP轉(zhuǎn)移酶(MCAT)、β-酮脂酰-ACP合酶(KAS)、β-酮脂酰-ACP還原酶(KAR)、β-羥脂酰-ACP脫水酶(HAD)、烯脂酰-ACP還原酶(ENR)。其中KAS、KAR、HAD和ENR分別調(diào)控催化、縮合、還原、脫水和再還原過程，形成了一個(gè)脂肪酸延長(zhǎng)酶系統(tǒng)，其功能與FAS類似，只是將乙酰-CoA用中鏈或長(zhǎng)鏈?；?CoA代替[29]，反應(yīng)過程中各種酰基均以?；?CoA的形式參與反應(yīng)，而不是以?；?ACP形式參與反應(yīng)[30]。

目前，研究最為廣泛的脂肪酸合成酶是植物質(zhì)體FASII的β-酮脂酰-ACP合酶(KAS)，它由3種酶構(gòu)成，分別為KASI、KAS II和KASIII。KASIII催化乙酰CoA結(jié)合丙二酰CoA生成4:0-ACP，KASI催化4:0-ACP到16:0-ACP的碳鏈延長(zhǎng)，而KAS II則催化16:0-ACP到18:0-ACP的生成[31-32]。前人研究表明通過控制KASIII基因表達(dá)可以改善油料作物的脂肪酸組成，如基因敲除突變油菜的KASIII和FATB基因，使得雙突變體的中鏈脂肪酸各成分含量提高[33]。KASI在脂肪酸形成及積累過程具有很重要的作用[34]，擬南芥kasI突變型含油量及育性明顯降低，通過互補(bǔ)表達(dá)實(shí)驗(yàn)恢復(fù)了突變體的育性、含油量和脂肪酸組成；而KASII突變會(huì)影響植株的表型，擬南芥kasII突變型互補(bǔ)表達(dá)麻瘋樹的JcKASII恢復(fù)了野生型的表型[35]。

KAR、HAD和ENR是脂肪酸從頭合成的相關(guān)基因，在碳鏈延伸循環(huán)中起著重要的作用。早前研究發(fā)現(xiàn)，大腸桿菌ENR基因在脂肪酸的碳鏈延伸中起決定性作用[36]。擬南芥ENR蛋白缺失的一個(gè)突變體表現(xiàn)出脂肪酸合成體系的損壞以及脂類含量的下降[37]。KAR和ENR可能是脂肪酸合成所需要的，HAD則可能發(fā)揮著不同的生物化學(xué)功能[38]。Bourgis等[39]分析了油棕(高油植物)和椰棗(低油植物)果皮的油脂合成相關(guān)各基因的表達(dá)量，結(jié)果表明油棕的KAR、HAD和ENR基因表達(dá)水平明顯高于椰棗，其中KAR和HAD的表達(dá)量是椰棗中的8倍。在棉花中，通過同源克隆的方法已經(jīng)得到陸地棉的GhKAR、GhHAD和GhENR基因的cDNA全長(zhǎng)[40]，研究發(fā)現(xiàn)，KAR、HAD和ENR基因在脂肪酸的合成中有重要作用[41]。在棉花中過表達(dá)GhKAR和GhENR基因可以提高棉籽含油量，棉籽中不飽和脂肪酸相對(duì)含量較對(duì)照增加10%左右[42]。

植物體內(nèi)飽和脂肪酸可在去飽和酶的作用下形成不飽和脂肪酸，其中包括棕櫚油酸和油酸等單不飽和脂肪酸及亞油酸和亞麻酸等長(zhǎng)鏈多聚不飽和脂肪酸。植物體去飽和酶存在于質(zhì)體，以脂酰-ACP為底物，從雙鍵向脂肪酸甲基端通過脂酰脫氫酶繼續(xù)去飽和[43]。在內(nèi)質(zhì)網(wǎng)上，單不飽和脂肪酸以磷脂或甘油糖脂的形式繼續(xù)去飽和，如磷脂酰膽堿上的油酸，在內(nèi)質(zhì)網(wǎng)上去飽和成為亞油酸或亞麻酸鏈[44]。

2.3 脂肪酸去飽和酶

飽和脂肪酸由脂酰-ACP去飽和酶催化去飽和，形成單不飽和或多不飽和脂肪酸。脂酰-ACP去飽和酶中研究最多的是硬脂酸脫氫酶(Stearoyl-ACP desaturase,SAD)和脂肪酸脫氫酶(Fatty acid desaturase,FAD)。SAD催化硬脂酸去飽和產(chǎn)生油酸，然后以脂形式存在的油酸被運(yùn)轉(zhuǎn)到類囊體膜中或進(jìn)入細(xì)胞質(zhì)中進(jìn)一步去飽和；若要進(jìn)一步去飽和形成多不飽和脂肪酸則需FAD[45]。SAD和FAD是決定脂肪酸不同組分含量的2個(gè)關(guān)鍵酶，可以通過調(diào)節(jié)它們編碼基因的表達(dá)來改善油脂品質(zhì)。有相關(guān)研究證明，F(xiàn)AD2沉默會(huì)明顯的提高植物油酸的含量，具體為使甘藍(lán)型油菜和芥菜型油菜的油酸質(zhì)量分?jǐn)?shù)分別升高至89%和75%[46]。在棉花中已成功構(gòu)建了種子特異性啟動(dòng)子NAPIN調(diào)控的ihpRNA干擾表達(dá)載體和針對(duì)GhFAD2-1基因的人工miRNA表達(dá)載體[47]。通過抑制SAD基因的表達(dá)可以提高硬脂酸含量，在棉花相關(guān)研究中曾有報(bào)道，利用干涉技術(shù)來抑制SAD1的表達(dá)使得棉籽的硬脂酸質(zhì)量分?jǐn)?shù)從2%增加到40%，油酸質(zhì)量分?jǐn)?shù)從15%提高到77%[48]。SAD可以影響FA組成，還可以增強(qiáng)植物抗逆性，如煙草質(zhì)體中過表達(dá)2種野生型馬鈴薯的Δ9脫氫酶基因后，改變了其脂肪酸組成，增加葉片和種子的不飽和脂肪酸含量，提高了植株耐寒性[49]。在棉花中，通過同源克隆的方法已經(jīng)得到陸地棉的GhSAD2基因的cDNA全長(zhǎng)，但它們對(duì)脂肪酸累積及抗寒性方面的作用及有待進(jìn)一步研究[50]。

2.4 二酰甘油脂酰轉(zhuǎn)移酶

二酰甘油脂酰轉(zhuǎn)移酶(Diacylglycerol acyltransferase,DGAT)是三酰甘油合成的限速酶，其作用是催化二酰甘油加上脂肪酸?；纬扇８视?。1998年首次在小鼠中發(fā)現(xiàn)DGAT的cDNA[51]，隨后在擬南芥中克隆到了DGAT1的cDNA[52]，以后在土壤真菌拉曼被孢霉中也克隆到DGAT2的2個(gè)同源基因DGAT2A和DGAT2B[53]。

植物主要發(fā)現(xiàn)有3種類型的DGAT：DGAT1、DGAT2和DGAT3[54]。DGAT1是植物種子油脂合成的關(guān)鍵酶，過量表達(dá)AtDGAT1，使得轉(zhuǎn)基因擬南芥種子中的DGAT活性比野生型高10%～70%，種子的千粒重和含油量也比野生型高[55]。DGAT2則被認(rèn)為是大量累積特殊脂肪酸(如蓖麻油酸)的主控基因，但DGAT2也在橄欖[56]和油棕[57]中參與常規(guī)TAG的累積。在玉米中表達(dá)真菌粗糙脈孢霉的短鏈S-NcDGAT2，小幅度提高了玉米的含油量，改變了脂肪酸的組成，提高了油酸含量降低了亞油酸含量，并對(duì)產(chǎn)量沒有顯著影響[58]。DGAT3研究的比較少，花生DGAT3在花后8～24 d的種子中特異表達(dá)。然后在種子成熟過程中逐漸降低直至表達(dá)消失，而在葉和根中卻沒有表達(dá)[59]。

對(duì)于DGAT在調(diào)控油脂合成方面的研究，主要集中于DGAT1與DGAT2基因家族，盡管這2種DGAT的蛋白序列存在差異，但它們都具備催化二酰甘油結(jié)合脂酰-CoA形成三酰甘油的功能。一般來說，在大部分植物中DGAT1在三酰甘油合成代謝中的作用更加廣泛，而DGAT2更側(cè)重于特殊脂肪酸的積累，兩者的作用并不相互排斥[60]。Wurie等[61]研究發(fā)現(xiàn)DGAT2作用于DGAT1基因的上游，影響TAG合成與儲(chǔ)藏。研究表明，DGAT的表達(dá)影響植物種子發(fā)育，并影響種子含油量，脂肪酸組成與種子粒重等[62]。

3 三酰甘油生物合成過程中的轉(zhuǎn)錄因子

3.1 LEC

LEC(Leafy cotyledon)是胚胎發(fā)生發(fā)育過程中關(guān)鍵的調(diào)控因子，它控制胚胎發(fā)育過程中的多個(gè)方面，同時(shí)它也對(duì)脂肪酸的合成起著重要的調(diào)控作用。LEC2、FUS3和ABI3屬于植物特異性轉(zhuǎn)錄因子B3超級(jí)家族成員，三者共同構(gòu)成AFL(ABI3/FUS3/LEC2)家族[63]。這些轉(zhuǎn)錄因子可以調(diào)控發(fā)育成熟期種子的性狀，如油脂積累、子葉特性等。其中LEC2和LEC1可互相上調(diào)對(duì)方表達(dá)[64]同時(shí)也可影響其他轉(zhuǎn)錄因子的表達(dá)。已有研究證明LEC1和LEC2是影響WRI表達(dá)的上游調(diào)控基因[65-66]。擬南芥LEC的突變體lec1、lec2和fus3存在顯著的胚胎成熟缺陷[67]。

Li等[68]克隆了花生LEC1兩個(gè)成員的cDNA，研究發(fā)現(xiàn)，LEC1在花生種子發(fā)育的不同時(shí)期表達(dá)有差異，種子的表達(dá)水平最高。過表達(dá)AtLEC1和BnLEC1基因，使得轉(zhuǎn)基因擬南芥植株脂肪酸的種類和脂質(zhì)的含量大幅度提高。有研究表明LEC可能是通過增加流向脂肪酸合成的碳流，從而增加脂肪酸含量[69]，具體可能是通過調(diào)控脂肪酸合成相關(guān)基因來影響油份含量，包括編碼乙酰輔酶A羧化酶、控制脂肪酸合成的關(guān)鍵酶以及參與糖酵解和脂質(zhì)積累的基因[70]。

3.2 WRI

WRI(WRINKLED1)是種子油脂合成及累積調(diào)控的關(guān)鍵轉(zhuǎn)錄因子，它能直接調(diào)控糖酵解和脂肪酸代謝過程，進(jìn)而提高脂肪酸合成相關(guān)基因的整體表達(dá)水平。該基因編碼蛋白含有2個(gè)AP2/EREBP結(jié)構(gòu)域，控制種子蔗糖轉(zhuǎn)化成油脂[71]。

1998年，首次發(fā)現(xiàn)了具有種子皺縮表型的擬南芥突變體，命名為wrinkled1(wri1)，該突變體含油量下降了80%，蔗糖含量增加，且糖酵解相關(guān)的酶活性均普遍下降[72]。通過cDNA芯片等方法分析發(fā)現(xiàn)，在種子油形成中WRI主要在轉(zhuǎn)錄水平上調(diào)控質(zhì)體糖酵解途徑的關(guān)鍵酶[73]、脂肪酸合成相關(guān)的酶來調(diào)控油脂的積累。與種子油類似，在非種子油油棕的中果皮中，WRI通過調(diào)控其下游基因丙酮酸激酶(PK)、丙酮酸脫氫酶(PDH)、生物素羧基載體蛋白1(BCCP1)、ENR、己糖激酶(HXK)的表達(dá)來影響油脂的含量。

近年來，有不少有關(guān)WRI用于作物育種的研究報(bào)道。如在擬南芥中過表達(dá)油菜BnWRI1基因，其種子含油量增加了10%～40%[74]。玉米中過表達(dá)ZmWRI1基因，轉(zhuǎn)基因后代株系中含油量的明顯增加并且沒有造成淀粉含量的下降[75]，并對(duì)其他農(nóng)藝性狀也沒有明顯影響[76]。Li等[77]在油菜中過表達(dá)BnWRI1使花期提前，并且種子含油量提高18%～38%。另外，在單子葉植物二穗短柄草(Brachypodiumdistachyon)的葉片中異位表達(dá)WRI1后，會(huì)使葉片中的TAG含量提高32.5倍，游離的FA含量提高2倍[78]。

3.3 Dof

Dof(DNA binding with one finger)是植物特有的一類轉(zhuǎn)錄因子，參與高等植物的復(fù)雜生理活動(dòng)的調(diào)控，如植物對(duì)激素、光的響應(yīng)，同時(shí)它還參與脂肪酸的合成調(diào)控。在Dof蛋白的N末端有一個(gè)獨(dú)特的由52個(gè)氨基酸組成的高度保守的Dof結(jié)構(gòu)域，而在這個(gè)保守的結(jié)構(gòu)域中CX2CX21CX2C基序形成一個(gè)單鋅指結(jié)構(gòu)，單鋅指結(jié)構(gòu)中1個(gè)Zn2+與4個(gè)Cys殘基共價(jià)結(jié)合[79]。

首先在玉米中鑒定Dof轉(zhuǎn)錄因子[80]。大豆中鑒定到28個(gè)Dof轉(zhuǎn)錄因子，其中GmDof4和GmDof11參與油脂合成。有研究表明，GmDof4和GmDof11能分別與ACCase和長(zhǎng)鏈乙酰CoA合成酶基因的啟動(dòng)子區(qū)域結(jié)合，激活該基因的表達(dá)，從而增加種子油脂的含量[81]。

4 展望

植物種子是植物油的主要來源，改良植物種子脂肪酸組成和提高含油量是油料作物研究的永恒課題。油脂合成是一個(gè)復(fù)雜的生理生化過程，有關(guān)油分改良的研究主要集中在植物脂肪酸合成途徑和三酰甘油合成途徑上，目前，可以通過分子標(biāo)記輔助育種以及轉(zhuǎn)基因手段來提高植物油份含量以及改變油分的組成，達(dá)到作物改造的目的。研究表明，增強(qiáng)三酰甘油途徑中?；D(zhuǎn)移酶的表達(dá)比增強(qiáng)脂肪酸合成能更有效地提高種子含油量[82]。目前油脂合成相關(guān)基因在植物抗逆方面也有大量的研究，如脂肪酸合酶復(fù)合體的關(guān)鍵基因KAR、ENR在碳鏈延伸循環(huán)中起著重要的作用，通過測(cè)定抗寒指標(biāo)，研究其轉(zhuǎn)基因株系維持細(xì)胞滲透調(diào)節(jié)能力有所增強(qiáng)，抗寒性有所提高[41]。研究最多的去飽和酶SAD基因，在銀杏和棉花中低溫脅迫誘導(dǎo)結(jié)果表明，SAD基因在不同程度低溫處理下均有上調(diào)表達(dá)[83]。油脂生物合成的相關(guān)基因在植物抗逆方面也有大量的研究，如乙酰輔酶A羧化酶ACCase在除草劑方面的研究[84]。油脂生物合成涉及大量的基因，篩選出對(duì)種子油份改良作用貢獻(xiàn)大并更能適應(yīng)逆境脅迫的相關(guān)基因是下一步的工作重點(diǎn)。近年來，隨著人們生活水平的不斷提高，對(duì)植物油的品質(zhì)需求也越來越高。在科研工作者的不斷努力下，脂肪酸合成、TAG組裝過程與油脂代謝調(diào)控網(wǎng)絡(luò)越來越清晰，通過現(xiàn)代基因工程技術(shù)來改良植物油脂含量和品質(zhì)、提高植物抗逆性將會(huì)是一條有效途徑。

[1]WANG H Y,GUO J H,LAMBERT K N,et al.Developmental control ofArabidopsisseed oil biosynthesis[J].Planta,2007,226:773-783

[2]KANG F,RAWSTHORNE S.Starch and fatty acid synthesis in plastids from developing embryos of oilseed rape(BrassicanapusL.)[J].Plant Journal,1994,6:795-805

[3]ZHAO J,BECKER H C,ZHANG D,et al.Conditional QTL mapping of oil content in rapeseed with respect to protein content and traits related to plant developmentand grain yield[J].Theoretical and Applied Genetics,2006,113:33-38

[4]WU J G,SHI C H,ZHANG H Z.Partitioning genetic effects due to embryo,cytoplasm and maternal parent for oil content in oilseed rape(BrassicanapusL.)[J].Genetics and Molecular Biology,2006,29:533-538

[5]TEICHERT S A,AKOH C C.Stearidonic acid soybean oil enriched with palmitic acid at the sn-2 position by enzymatic interesterification for Use as human milk fat analogues[J].Journal of Agricultural Food and Chemistry,2011,59:5692-5701

[6]LI H,PENG Z Y,YANG X H,et al.Genome-wide association study dissects the genetic architecture of oil biosynthesis in maize kernels[J].Nature Genetics,2013,45:43-50

[7]QU J,MAO H Z,CHEN W,et al.Development of marker-free transgenicJatrophaplants with increased levels of seed oleic acid[J].Biotechnology for Biofuels,2012(5):10

[8]LIU Z J,GUO B S,ZHANG Y,et al.Research and improvement on the oil content and composition of cottonseed[J].Molecular Plant Breeding,2012,10(6):1038-1048

[9]SIMCOX P D,REID E E,CANVIN D T,et al.Enzymes of the glycolytic and pentose phosphate pathways in proplastids from the developing endosperm ofRicinuscommunisL[J].Plant Physiology,1977,59:1128-1132

[10]STITT M,REES T A.Estimation of the activity of the oxidative pentose phosphate pathway in pea chloroplasts[J].Phytochemistry,1980,19:1583-1585

[11]HUANG Y,WANG J F,ZHANG H S.Advances on plant pentose phosphate pathway and its key enzymes[J].Chinese Bulletin Botany,2004,21(2):139-145

[12]JONES A,DAVIES H M,VOELKER T A.Palmitoyl-Acyl carrier protein(ACP)thioesterase and the evolutionary origin of plant acyl-ACP thioesterases[J].Plant Cell,1995,7(3):359-371

[13]SALAS J J,OHLROGGE J B.Characterization of substrate specicity of plantFatAandFatBacyl-ACP thioesterases[J].Archives Biochemistry Biophysics,2002,403:25-34

[14]BAUD S,LEPINIEC L.Physiological and developmental regulation of seed oil production[J].Progress in Lipid Research,2010,49:235-249

[15]MANUEL A,TRONCOSO P,ARUNA K,et al.Comparative deep transcriptional proling of four developing oilseeds[J].Plant Journal,2011,68:1014-1027

[16]TJELLSTROM H,YANG Z,ALLEN D K,et al.Rapid kineticlabeling of Arabidopsis cell suspension cultures:implicationsfor models of lipid export from plastids[J].Plant Physiology,2012,158:601-611

[17]KIM S,YAMAOKA Y,ONO H,et al.AtABCA9 transporter supplies fatty acids for lipid synthesis to the endoplasmic reticulum[J].Proceedings of the National Academy of Sciences of USA,2013,110:773-778

[18]BAUD S,GUYON V,KRONENBERGER J,et al.Multifunctional acetyl-CoA carboxylase 1 is essential for very long chain fatty acid elongation and embryo development inArabidopsis[J].Plant Journal,2003,33:75-86

[19]WANG F L,WU G T,LANG C X,et al.Acetyl-CoA Carboxylase in Plant[J].Plant Physiology Communications,2006,42(1):10-14

[20]ALBAN C,JOB D,DOUCE R.Biotin metabolism in plants[J].Annual Review of Plant Physiology and Plant Molecular Biology,2000,51:17-47

[21]GUO G G.Basic Biochemistry[M].Beijing:Higher Education Press,2001：211-233

[22]ROESSLER P G,OHLROGGE J B.Cloning and characterization of the gene that encodes in the algaCyclotellacryptica[J].Journal Biological Chemistry,1993,268:19254-19259

[23]NIKOLAU B J,OHLROGGE J B,WURTELE E S.Plant biotin-containing carboxylases[J].Archives Biochemistry Biophysics,2003,414:211-222

[24]GENGENBACH B G,SOMERS D A,WYSE D L,et al.Methods and an acetyl-CoA carboxylase gene for conferring herbicide tolerance and an alteration in oil content of plants:United States, Us6069298[P].2000

[25]SASAKI Y,NAGANO Y.Plant acetyl-CoA carboxylase:structure,biosynthesis,regulation,and gene manipulation for plant breeding[J].Bioscience Biotechnology Biochemistry,2004,68:1175-1184

[26]ROESLER K,SHINTANI D,SAVAGE L,et al.Targeting of theArabidopsishomomeric acetyl-coenzyme A carboxylase to plastids of rapeseeds[J].Plant Physiology,1997,113:75-81

[27]ROESLER K R,SAVAGE L J,SHINTANI D K,et al.Co-purification,co-immunoprecipitation,and coordinate expression of acetyl-coenzyme A carboxylase activity,biotin carboxylase,and biotin carboxyl carrier protein of higher plants[J].Planta,1996,198:517-525

[28]MADOKA Y,TOMIZAWA K I,MIZOI J,et al.Chloroplast transformation with modifiedaccDoperon increased acetyl-CoA carboxylase and causes extension of leaf longevity and increase in seed yield in tobacco[J].Plant Cell Physiology,2002,43:1518-1525

[29]JAMES D W,LIM E,KELLER J,et al.Direcled tagging of theArabidopsisFATTY ACID ELONGA TIONI(EAE1)gene with maize transposoa aclivator[J].Plant Cell,1995(7):309-319

[30]DITTRICH F,ZAJONC D,HUHNE K,et al.Fatty acid elongation in yeast biochemical charateristics of the enzyme system and isolation of elongation-defective mutants[J].European Journal of Biochemistry,1998,252:477-485

[31]PIDKOWITCH M S,NGUYEN H T,HEILMANN I,et al.Modulating seed ?-ketoacyl-acyl carrier protein synthase II level converts the composition of a temperate seed oil to that of a palm-like tropical oil[J].Proceedings of the National Academy of Science USA,2007,104:4742-4747

[32]LI X F.Cloning and analysis of genes of fatty acid biosynthesis-related enzymes(KASI、FatB)and genetic transformation ofahFAD2BinArachishypogaeaL.[D].Beijing:China Agricultural University,2010

[33]WU G Z,XUE H W.Arabidopsisβ-ketoacyl-[Acyl Carrier Protein]synthase is crucial for fatty acid synthesis and plays a role in chloroplast division and embryo development[J].Plant Cell,2010,22:3726-3744

[34]WEI Q,LI J,ZHANG L,et al.Cloning and characterization of a β-ketoacyl-acyl carrier protein synthaseⅡ fromJatrophacurcas[J].Plant Physiology,2012,169:816-824

[35]STOLL C,LüHS W,ZARHLOUL M K,et al.Knockout of KASIII regulation changes fatty acid composition in canola(BrassicanapusL.)[J].European Journal of Lipid Science and Technology,2010,108:277-286

[36]RICHARD J H,CHARLES O R.Enoyl-acyl carrier protein reductase(fabI)plays a determinant role in completing cycles of fatty acid elongation inescherichiacoli[J].Journal of Biological Chemistry,1995,270:26538-26542

[37]MOU Z,HE Y K,DAI Y,et al.Deficiency in fatty acid synthase leads to premature cell death and dramatic alterations in plant morphology[J].Plant Cell,2000,12:405-417

[38]CHI X Y,YANG Q L,PAN L J,et al.Cloning and expression analysis of β-ketoacyl-ACP reductase,β-hydroxyacyl-ACP dehydrase and enoyl-ACP reductase fromArachishypogaeaL.[J].Genomics and Applied Biology,2010(2):1-9

[39]BOURGIS F,KILARU A,CAO X,et al.Comparative transcriptome and metabolite analysis of oil palm and date palm mesocarp that differ dramatically in carbon partitioning[J].Proceedings of the National Academy of Sciences of USA,2011,108(30):12527-12532

[40]ZHAO P.Isolation andanalysis of fatty acid synthasis related genesGhKAR,GhHAD,GhENRin upland cotton[D].Beijing:China Agricultural University,2012

[41]LIU L,WANG Y M,ZHAO Y P,et al.Construction and function ofGhKARandGhENRexpression vector of fatty acid synthase gene in cotton[J].Cotton Science,2016,6:527-537

[42]LIU L.Gene expression profiling during fiber secondary cell wall development and functional verification ofGhKAR,GhHADandGhENRin cotton[D].Beijing:China Agricultural University,2015

[43]GUO G G.Basic Biochemistry[M].Beijing:Higher Education Press,2001:211-233

[44]WANG J Y,ZHU S G,XU C F.Biochemistry[M].Beijing:Higher Education Press,2004:265

[45]LI X F.Cloning and analysis of genes of fatty acid biosynthesis-related enzymes(KASI、FatB)and genetic transformation ofahFAD2BinArachishypogaeaL[D].Beijing:China Agricultural University,2010

[46]OHLROGGE J B,BROWSE J A.Lipid biosynthesis[J].Plant Cell,1995,7:957-970

[47]ZHAO L Q,LI R,LI W,et al.Cloning of delta-12 oleate desaturase geneFAD2-1 and construction of its ihpRNA and amiRNA interference vectors fromGossypiumhirsutum[J].Cotton Science,2011,23(2):189-192

[48]LIU Q,SURINDER P S,ALLAN G G.High-stearic and high-oleic cottonseed oils produced by hairpin RNA-mediated post-transcriptional gene silencing[J].Plant Physiology,2002,129:1732-1743

[49]CRAIG W,LENZI P,SCOTTI N,et al.Transplastomic tobacco plants expressing a fatty acid desaturase gene exhibit altered fatty acid profiles and improved cold tolerance[J].Transgenic Research,2008,17:769-782

[50]CAI M,LI W H,WANG J,et al.Molecular cloning and expression analysis of stearoyl-ACP desaturase gene(GhSAD2)in upland cotton(GossypiumhirsutumL.)[J].Acta Botanica Boreali-Occidentalia Sinica,2016,9:1713-1720

[51]CASES S,SMITH S J,ZHENG Y W,et al.Identification of a gene encoding an acyl-CoA:diacylglycerol acyltransferase,a key enzyme in triacylglycerol synthesis[J].Proceedings of the National Academy of Sciences of USA,1998,95:13018-1323

[52]HOBBS D H,LU C,HILLS M J.Cloning of a cDNA encoding diacylglycerol acyltransferase fromArabidopsisthalianaand its functional expression[J].Febs Letters,1999,452:145-149

[53]LARDIZABAL K D,MAI J T,WAGNER N W,et al.DGAT2 is a new diacylglycerol acyltransferase gene family:purification,cloning,and expression in insect cells of two polypeptides from Mortierella ramanniana with diacylglycerol acyltransferase activity[J].Journal of Biological Chemistry,2001,276:38862-38869

[54]WANG T T,MA X L,LI F L,et al.Advances in plant diacylglycerol acyltransferases and the coding genes[J].Guangdong Agriculture Science,2012,39(6):127-130

[55]JAKO C,KUMAR A,WEI Y D,et al.Seed-specific over-expression of anArabidopsiscDNA encoding a diacylglycerel acyltransferase enhances seed oil content and seed weight[J].Plant Physiology,2001,126:861-874

[56]ALAGNA F,AGOSTINO N,TORCHIA L,et al.Comparative 454 pyrosequencing of transcripts from two olive genotypes during fruit development[J].BMC Genomics,2009,10:399

[57]BOURGIS F,KILARU A,CAO X,et al.Comparative transcriptome and metabolite analysis of oil palm and date palm mesocarp that differ dramatically in carbon partitioning[J].Proceedings of the National Academy of Sciences of USA,2011,108(30):12527-12532

[58]OAKES J,BRACKENRIDGE D,COLLETTI R,et al.Expression of fungal diacylglycerol acyltransferase2 genes to increase kernel oil in maize[J].Plant Physiology,2011,155:1146-1157

[59]SAHA S,ENUGUTTI B,RAJAKUMARI S,et al.Molecular cloning and expression of peanut cytosolic diacylglycerol acyltransferasc[J].Plant Physiology,2006,141:1533-1543

[60]SHOCKEY J M,GIDDA S K,CHAPITAL D C,et al.Tung Tree DGAT1 and DGAT2 have nonredundant functions in triacylglycerol biosynthesis and are localized to different subdomains of the endoplasmic reticulum[J].The Plant Cell,2006,18(9):2294-2313

[61]WURIE H R,BUCKETT L,ZAMMIT V A.Diacylglycerol acyltransferase 2 acts upstream of diacylglycerol acyltransferase 1 and utilizes nascent diglycerides and de novo synthesized fatty acids inHepG2 cells[J].Febs Journal,2012,279(17):3033-3047

[62]TAYLOR D C,ZHANG Y,KUMAR A,et al.Molecular modification of triacylglycerol accumulation by over-expression of DGAT1to produce canola with increased seed oil content under field conditions[J].Botany,2009,87:533-543

[63]PARCY F,VALON C,KOHARA A,et al.TheABSCISICACID-lNSENSITIVE3,FUSCA3,andLEAFYCOTYLEDON1 loci act in concert to control multiple aspects ofArabidopsisseed development[J].Plant Cell,1997(9):1265-1277

[64]TO A,VALON C,SAVINO G,et al.A network of local and redundant gene regulation governsArabidopsisseed maturation[J].Plant Cell,2006,18:1642-1651

[65]WESELAKE R J,TAYLOR D C,RAHMAN M H,et al.Increasing the flow of carbon into seed oil[J].Biotechnology Advances,2009,27:866-878

[66]BAUD S,MENDOZA M S,TO A,et al.WRINKLED1 specifies the regulatory action ofLEAFYCOTYLEDON2 towards fatty acid metabolism during seed maturation inArabidopsis[J].Plant Journal,2007,50:825-838

[67]MEINKE D W,FRANZMANN L H,NICKLE T C,et al.Leafy cotyledon mutants ofArabidopsis[J].Plant Cell,1994,6:1049-1064

[68]LI A Q,XIA H,WANG X J,et al.Cloning and expression analysis of peanut(ArachishypogaeaL.)LEC1[J].Acta Botanica Boreali-Occidentalia Sinica,2009,29(9):1730-1735

[69]TAN H L,YANG X L,ZHANG F X,et al.Enhanced seed oil production in canola by conditional expression of brassica napusLEAFYCOTYLEDON1 andLEC1-LIKEin developing seeds[J].Plant Physiology,2011,156:1577-1588

[70]MU J Y,TAN H L,ZHENG Q,et al.LEAFYCOTYLEDON1 is a key regulator of fatty acid biosynthesis inArabidopsis[J].Plant Physiology,2008,148:1042-1054

[71]LU Y P,LIU F Z,WAN Y S.In silico Analysis of peanut transcription factor geneWRI1[J].Molecular Plant Breeding,2012,3(10):363-370

[72]FOCKS N,BENNING C.Wrinkled1:a novel,low seed oil mutant ofArabidopsiswith efficiency in the seed-specific regulation of carbohydrate metabolism[J].Plant Physiology,1998,118:91-101

[73]RUUSKA S A,GIRKE T,BENNING C,et al.Contrapuntal network of gene expression duringArabidopsisseed filling[J].Plant Cell,2002,14:1191-1206

[74]LIU J,HUA W,ZHAN G M,et al.Increasing seed mass and oil content in transgenicArabidopsisby the overexpression of wri1-like gene fromBrassicanapus[J].Plant Physiology and Biochemistry,2010,48:9-15

[75]POUVREAU B,BAUD S,VERNOUD V,et al.Duplicate maizeWrinkled1 transcription factors activate target genes involved in seed oil biosynthesis[J].Plant Physiology,2011,156:674-686

[76]SHEN B,ALLEN W B,ZHENG P,et al.Expression ofZmLEC1 andZmWRI1 increases seed oil production in maize[J].Plant Physiology,2010,153:980-987

[77]LI Q,SHAO J H,TANG S H,et al.Wrinkled1 accelerates flowering and regulates lipid homeostasis between oil accumulation and membrane lipid anabolism inBrassicanapus[J].Front Plant Science,2015,6(192):1-15

[78]YANG Y,MUNZ J,CASS C,et al.2015,Ectopic expression ofWRlNKLED1 affects fatty acid homeostasis inBrachypodiumdistachyonvegetative tissues[J].Plant physiology,2015,169(3):1836-1847

[79]YANAGISAWA S.Dof domain proteins:plant-specific transcription factors associated with diverse phenomena unique to plants.Plant Cell Physiology,2004,45:386-391

[80]YANAGISAWA S,IZUI K.Molecular cloning of two DNA-binding proteins of maize that are structurally different but interact with the same sequence motif[J].The Journal of Biological Chemistry,1993,268:16028-16036

[81]WANG H W,ZHANG B,HAO Y J,et al.The soybeanDof-type transcription factor genes,GmDof4 andGmDof11,enhance lipid content in the seeds of transgenicArabidopsisplants[J].Plant,2007,52:716-729

[82]WU L S H,HONG G H H,HOU R F,et al.Classification of the single oleosin isoform and characterization of seed oil bodies in gymnosperms[J].Plant Cell physiology,1999,40:326-334

[83]ZHAO N,ZHANG Y,et al.Identification and expression of a stearoyl-ACP desaturase gene responsible for oleic acid accumulation inXanthocerassorbifoliaseeds[J].Plant Physiology and Biochemistry,2015,87:9-16

[84]DU L,LIU W T,YUAN G H,et al.Cross-resistance patterns to ACCase-inhibitors in American sloughgrass(BeckmanniasyzigachneSteud.)homozygous for specific ACCase mutations[J].Pesticide Biochemistry and Physiology,2016,126:42-48.