本文選題：桂花 + 擴(kuò)展蛋白�。� 參考：《農(nóng)業(yè)生物技術(shù)學(xué)報(bào)》2017年08期

【摘要】：擴(kuò)展蛋白(expansin,EXP)是一類引起植物細(xì)胞壁松弛的蛋白質(zhì),在花開(kāi)放過(guò)程花瓣細(xì)胞擴(kuò)展中發(fā)揮重要作用。為了探究桂花(Osmanthus fragrans)中擴(kuò)展蛋白基因在花開(kāi)放中的表達(dá)特征,本研究從已構(gòu)建的桂花品種‘堰虹桂’花芽轉(zhuǎn)錄組數(shù)據(jù)庫(kù)中篩選得到12個(gè)與植物EXP基因同源性較高的Unigene序列,以盛開(kāi)期花瓣為材料克隆得到其cDNA序列。通過(guò)NCBI數(shù)據(jù)庫(kù)中的比對(duì)結(jié)果,將12個(gè)Unigene序列分別命名為OfEXPA1~OfEXPA8,OfEXPB1~OfEXPB2,OfEXLA1,OfEXLB1(Gen Bank登錄號(hào):KY968700~KY968711)。BLASTX查找比對(duì)結(jié)合ORF Finder工具分析發(fā)現(xiàn),除了OfEXPA1、OfEXPA5和OfEXLB1基因的cDNA序列5′端有少量缺失外,另外9個(gè)EXP基因具有cDNA全長(zhǎng)序列。序列分析結(jié)果表明,9個(gè)全長(zhǎng)cDNA推導(dǎo)的蛋白氨基酸殘基數(shù)為240~274個(gè),預(yù)測(cè)的分子量為26.06~29.7 kD,理論等電點(diǎn)為4.77~9.74。桂花OfEXPA1~OfEXPA8和OfEXPB1~OfEXPB2蛋白具有典型的擴(kuò)展蛋白結(jié)構(gòu)特征,N端含有8個(gè)保守的半胱氨酸結(jié)構(gòu)域,C端含有4個(gè)保守的色氨酸結(jié)構(gòu)域,其中大部分EXPA和EXPB蛋白含有組氨酸功能域HFD基序序列。OfEXLA1具有特征基序CDQC,OfEXLB1特征基序尚未見(jiàn)報(bào)道。qRT-PCR分析結(jié)果發(fā)現(xiàn)桂花12個(gè)EXP基因在‘堰虹桂’花芽不同發(fā)育過(guò)程中均有表達(dá),其中OfEXPA1、OfEXPA3、OfEXPA5、OfEXPA6、OfEXPA7、OfEXPA8、OfEXPB1、OfEXPB2和OfEXLB1整體表達(dá)模式較為類似,花芽發(fā)育前期(B1~B6)表達(dá)量顯著高于花瓣擴(kuò)展期(B7~FF),在花瓣擴(kuò)展期表達(dá)量逐漸下降;而OfEXPA2、OfEXPA4和OfEXLA1在花芽發(fā)育前期表達(dá)量較低,而隨著花瓣顯著伸展表達(dá)量急劇增加,在花瓣擴(kuò)展期呈現(xiàn)較高的表達(dá)。結(jié)合系統(tǒng)進(jìn)化樹(shù)分析發(fā)現(xiàn)OfEXPA4和OfEXPA2分別與唐菖蒲(Gladiolus grandiflorus)GgEXPA1和月季(Rosa chinensis)RhEXPA1親緣關(guān)系較近,且OfEXPA2和OfEXPA4這兩個(gè)基因與GgEXPA1和Rh EXPA1基因的表達(dá)模式相似,推測(cè)這2個(gè)基因?yàn)榛ㄩ_(kāi)放過(guò)程中的關(guān)鍵EXP基因家族成員;進(jìn)一步根據(jù)OfEXLA1表達(dá)特征推測(cè)其亦參與調(diào)控桂花花開(kāi)放。研究結(jié)果為全面理解桂花花開(kāi)放機(jī)制、擴(kuò)展蛋白基因功能提供理論依據(jù)。
[Abstract]:Expansinus exp) is a kind of protein that causes cell wall relaxation in plants and plays an important role in the expansion of flower petals.In order to investigate the expression characteristics of expanded protein gene in flowering of Osmanthus fragrans, 12 Unigene sequences with high homology to plant EXP gene were screened from the flower bud transcriptional database of Osmanthus manthus Fragrans.Its cDNA sequence was cloned from petals in full bloom.The other 9 EXP genes had full length cDNA sequences.The results of sequence analysis showed that the deduced amino acid residues of 9 full-length cDNA were 240N 274, the predicted molecular weight was 26.06 ~ 29.7 KD, and the theoretical isoelectric point was 4.77 ~ 9.74.The structural characteristics of OfEXPA1~OfEXPA8 and OfEXPB1~OfEXPB2 proteins in Osmanthus Osmanthus showed that the N-terminal contained eight conserved cysteine domains and four conserved tryptophan domains.Most of the EXPA and EXPB proteins contained histidine functional domain HFD motif. OfEXLA1 had the characteristic motif CDQCnOfEXLB1. The results of qRT-PCR analysis showed that 12 EXP genes of Osmanthus osmanthus were expressed in different stages of flower bud development.However, with the increase of the expression of petal significantly, it showed a higher expression in the period of petal expansion.The phylogenetic tree analysis showed that OfEXPA4 and OfEXPA2 were closely related to Gladiolus grandiflorus)GgEXPA1 and Rosa chinensis)RhEXPA1, respectively, and the expression patterns of OfEXPA2 and OfEXPA4 were similar to those of GgEXPA1 and Rh EXPA1.We speculated that these two genes were the key members of EXP gene family in flowering process, and they were also involved in the regulation of Osmanthus fragrans blooming according to the characteristics of OfEXLA1 expression.The results provide theoretical basis for understanding the mechanism of Osmanthus fragrans blooming and expanding the function of protein gene.
【作者單位】： 浙江農(nóng)林大學(xué)風(fēng)景園林與建筑學(xué)院;浙江理工大學(xué)建筑工程學(xué)院;
【基金】：浙江省自然科學(xué)基金(No.LQ16C160003) 浙江省教育廳科研項(xiàng)目(No.Y201533133) 浙江農(nóng)林大學(xué)科研發(fā)展基金人才啟動(dòng)項(xiàng)目(No.2016FR031) 浙江省農(nóng)業(yè)新品種選育重大科技專項(xiàng)(No.2016C02056-12)
【分類號(hào)】：S685.13

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