本文選題：森林草莓 + 黃果��； 參考：《沈陽(yáng)農(nóng)業(yè)大學(xué)》2017年博士論文

【摘要】：果色是影響草莓果實(shí)外觀品質(zhì)和商品價(jià)值的核心指標(biāo),草莓果色是花青苷積累的結(jié)果,目前對(duì)于草莓花青苷積累調(diào)控機(jī)制的研究還不充分。森林草莓(Fragaria vesca)是多年生草本植物,基因組小,是研究草莓屬植物以及薔薇科植物的重要模式材料。草莓的黃果性狀由c位點(diǎn)基因控制。為了研究與森林草莓黃果表型相關(guān)的關(guān)鍵基因,我們首先利用黃果森林草莓'Yellow Wonder'驗(yàn)證FvF3H(報(bào)道稱(chēng)與c位點(diǎn)基因共分離)的基因功能,隨后,結(jié)合轉(zhuǎn)錄組高通量測(cè)序、遺傳圖譜構(gòu)建和比較基因組分析進(jìn)一步篩選c位點(diǎn)可能的候選基因。主要結(jié)果如下:1.利用HPLC方法測(cè)定森林草莓'Yellow Wonder'(YW)和'Ruegen'(RG)果實(shí)中花青苷含量和組分,結(jié)果顯示紅果類(lèi)型RG中花青苷含量隨果實(shí)成熟而持續(xù)增加,而黃果類(lèi)型YW果實(shí)成熟過(guò)程中花青苷含量變化很小。RG果實(shí)中花青苷組分以Cy 3-glucoside和Pg3-glucoside為主,還有Pg3-rutinoside和Pg3-malonylglucoside;YW中能夠檢測(cè)到少量的Cy 3-glucoside、Pg 3-glucoside 和 Pg 3-malonylglucoside 三類(lèi)花青苷。2.開(kāi)展FvF3H基因功能驗(yàn)證,發(fā)現(xiàn)在YW中過(guò)量表達(dá)紅果的FvF3H基因沒(méi)有引起果實(shí)花青苷含量的增加,也沒(méi)有引起果色變紅。3.采用Illumina測(cè)序技術(shù)對(duì)森林草莓YW和RG轉(zhuǎn)熟期果實(shí)進(jìn)行轉(zhuǎn)錄組高通量測(cè)序。經(jīng)de novo裝配得到75,426條Unigene,鑒定獲得595個(gè)差異表達(dá)基因,其中224個(gè)在YW中上調(diào)表達(dá),371個(gè)下調(diào)表達(dá)。4.分析轉(zhuǎn)錄組測(cè)序結(jié)果和qRT-PCR結(jié)果,發(fā)現(xiàn)6個(gè)花青苷合成催化酶的編碼基因(C4H、CHS、CHI、F3H、DFR和ANS)在YW中同時(shí)下調(diào)表達(dá);多個(gè)調(diào)控花青苷合成的轉(zhuǎn)錄因子也存在顯著表達(dá)差異,其中MYB1、MADS等轉(zhuǎn)錄因子在YW中下調(diào)表達(dá),MYB1R在YW中上調(diào)表達(dá)。對(duì)花青苷合成相關(guān)結(jié)構(gòu)基因和轉(zhuǎn)錄因子的序列分析發(fā)現(xiàn)RG和YW的MYB10基因CDS序列中存在一個(gè)SNP差異。5.利用構(gòu)建的F2群體材料繪制了森林草莓c位點(diǎn)的基因連鎖圖譜,確定包含c位點(diǎn)基因候選區(qū)域的物理距離是718 kb。6.整合圖譜數(shù)據(jù)與擬南芥、森林草莓全基因組水平的比較基因組分析數(shù)據(jù),發(fā)現(xiàn)轉(zhuǎn)錄因子FvMYB10位于c位點(diǎn)基因的候選區(qū)域內(nèi)。7.發(fā)現(xiàn)FvMYB10基因編碼區(qū)存在TGG→TCG的SNP差異,氨基酸序列也存在相應(yīng)差異Trp→Ser,且該SNP差異與森林草莓果色表型共分離。因此,推斷FvMYB10是c位點(diǎn)基因候選基因。8.分析發(fā)現(xiàn)FvMYB10的突變位點(diǎn)TGG→TCG(Trp→Ser)位于MYB R2 DNA結(jié)合區(qū)域,該區(qū)域?qū)φ{(diào)控花青苷合成基因的表達(dá)有重要作用。因此,FvMYB10 R2區(qū)域的錯(cuò)義突變(Trp→Ser)可能是黃果表型產(chǎn)生的關(guān)鍵所在。9.試驗(yàn)發(fā)現(xiàn)YW和RG間FvMYB10的表達(dá)量并沒(méi)有顯著的差異。說(shuō)明FvMYB10的SNP變異影響果色表型的作用機(jī)理可能更多的是通過(guò)影響其對(duì)花青苷合成基因的調(diào)控作用,而不是通過(guò)影響自身基因表達(dá)來(lái)實(shí)現(xiàn)的。
[Abstract]:Fruit color is the core index that affects the appearance quality and commodity value of strawberry fruit. Strawberry fruit color is the result of anthocyanin accumulation. At present, the regulation mechanism of strawberry anthocyanin accumulation is not enough. Fragaria vesca is a perennial herbaceous plant with small genome. It is an important model material for the study of Strawberry and Rosaceae plants. The yellow fruit traits of strawberry were controlled by c locus gene. In order to study the key genes associated with the phenotype of forest strawberry yellow Wonder', we first used yellow Wonder'to verify the gene function of FvF3H (reported co-isolation with c locus), and then combined with high throughput sequencing of transcriptome. Genetic map construction and comparative genomic analysis were used to screen possible c locus candidate genes. The main results are as follows: 1. The contents and components of anthocyanin in the fruit of Yellow Wonder'(YW) and Ruegen'(RG) were determined by HPLC. The results showed that the content of anthocyanin in red fruit type RG increased with the fruit ripening. However, the content of anthocyanin in YW fruit changed little during maturation. The components of anthocyanin in RG fruit were mainly Cy 3-glucoside and Pg3-glucoside, and a small amount of Cy 3-glucoside PG 3-glucoside and PG 3-malonylglucoside were detected in YW, and a small amount of Cy 3-glucoside PG 3-glucoside and PG 3-malonylglucoside were detected in YW. It was found that FvF3H gene overexpressed in YW did not increase anthocyanin content or red fruit color. The transcriptional high-throughput sequencing of forest strawberry YW and RG was carried out by Illumina sequencing technique. 75426 Unigene genes were obtained by de novo assembly, and 595 differentially expressed genes were identified, of which 224 genes were up-regulated and 371down-regulated genes were expressed in YW. By analyzing the results of transcriptome sequencing and qRT-PCR, it was found that six genes encoding anthocyanin synthase (C _ 4H _ (4) H _ (CHS) CHIF _ (3H) DFR and ans were down-regulated simultaneously in YW, and that there were significant differences in the expression of several transcription factors regulating anthocyanin synthesis. The expression of MYB1R was down-regulated in YW, and MYB1R was up-regulated in YW. Sequence analysis of structural genes and transcription factors associated with anthocyanin synthesis revealed that there was a SNP difference between RG and YW MYB10 gene CDS sequences. The genetic linkage map of c locus of forest strawberry was plotted by using F2 population material, and the physical distance of candidate region containing c locus was 718 kb. 6. The integration map data and the comparative genomic analysis data of Arabidopsis thaliana and forest strawberry showed that the transcription factor FvMYB10 was located in the candidate region of c locus gene. It was found that there were SNP differences of TGG and TCG in the coding region of FvMYB10 gene, and there was also a corresponding difference in amino acid sequence, and the SNP difference was coisolated from the color phenotype of forest strawberry fruit. Therefore, it is inferred that FvMYB10 is a candidate gene of c locus. It was found that the mutation site of FvMYB10, TGG TCG (TRP Ser), was located in the DNA binding region of MYB R2, which played an important role in regulating the expression of anthocyanin synthesis gene. Therefore, the missense mutation of FvMYB10 R2 region (TRP Ser) may be the key to the phenotypic production of yellow fruit .9. The results showed that there was no significant difference in FvMYB10 expression between YW and RG. The results suggested that the mechanism of SNP variation of FvMYB10 affecting fruit color phenotype may be more by affecting its regulation on anthocyanin synthesis gene than by affecting its own gene expression.
【學(xué)位授予單位】：沈陽(yáng)農(nóng)業(yè)大學(xué)
【學(xué)位級(jí)別】：博士
【學(xué)位授予年份】：2017
【分類(lèi)號(hào)】：S668.4

【參考文獻(xiàn)】

相關(guān)期刊論文 前10條

1 徐強(qiáng);郝玉金;黃三文;鄧秀新;;果實(shí)品質(zhì)研究進(jìn)展[J];中國(guó)基礎(chǔ)科學(xué);2016年01期

2 趙貝塔;趙巍;劉鄰渭;;花青素類(lèi)化合物分析方法研究進(jìn)展[J];西北農(nóng)林科技大學(xué)學(xué)報(bào)(自然科學(xué)版);2014年06期

3 豆玉娟;曹飛;馬躍;李賀;劉月學(xué);張志宏;;栽培草莓果實(shí)中特異表達(dá)的bHLH78基因的克隆及過(guò)量表達(dá)載體構(gòu)建[J];分子植物育種;2014年03期

4 劉曉芬;李方;殷學(xué)仁;徐昌杰;陳昆松;;花青苷生物合成轉(zhuǎn)錄調(diào)控研究進(jìn)展[J];園藝學(xué)報(bào);2013年11期

5 李小白;向林;羅潔;胡標(biāo)林;田勝平;謝鳴;孫崇波;;轉(zhuǎn)錄組測(cè)序(RNA-seq)策略及其數(shù)據(jù)在分子標(biāo)記開(kāi)發(fā)上的應(yīng)用[J];中國(guó)細(xì)胞生物學(xué)學(xué)報(bào);2013年05期

6 林紹杰;仇丹;吳月嬋;勞夢(mèng)夢(mèng);;類(lèi)胡蘿卜素光譜信息的量子化學(xué)模擬[J];寧波工程學(xué)院學(xué)報(bào);2012年03期

7 宮硤;薛靜;張曉東;;植物花青素合成途徑中的調(diào)控基因研究進(jìn)展[J];生物技術(shù)進(jìn)展;2011年06期

8 龐紅霞;祝長(zhǎng)青;覃建兵;;植物花青素生物合成相關(guān)基因研究進(jìn)展[J];種子;2010年03期

9 趙文軍;張迪;馬麗娟;柴友榮;;原花青素的生物合成途徑、功能基因和代謝工程[J];植物生理學(xué)通訊;2009年05期

10 張寧;胡宗利;陳緒清;侯曉姝;李勇;陳國(guó)平;;植物花青素代謝途徑分析及調(diào)控模型建立[J];中國(guó)生物工程雜志;2008年01期

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