荷花‘青玉’花瓣轉(zhuǎn)錄組測(cè)序以及花開(kāi)放差異基因分析
發(fā)布時(shí)間：2019-05-20 10:00
【摘要】：荷花(Nelumbo nucifera Gaern)屬于睡蓮科蓮屬,是多年生草本植物花卉,具有極高的觀賞和經(jīng)濟(jì)價(jià)值。荷花的花期時(shí)間較短,研究荷花花期的生物學(xué)基礎(chǔ)相對(duì)滯后,所以提升荷花開(kāi)花時(shí)間的研究尤為重要。轉(zhuǎn)錄組測(cè)序技術(shù)已成為研究分子功能的主流,極大地加快了生物相關(guān)功能的研究進(jìn)展。本研究通過(guò)高通量Illumina HiSeq2500平臺(tái)測(cè)序,研究‘青玉’花瓣開(kāi)放的暄蕾期與盛花期的轉(zhuǎn)錄水平,最終獲得50,154條unigene,總長(zhǎng)度3,938,0851bp,平均長(zhǎng)度是785.20bp,N50的長(zhǎng)度是1,412bp。轉(zhuǎn)錄本中有23,753條unigene對(duì)比到GO數(shù)據(jù)庫(kù)中,其中三大類中包含56組功能;有32,194條unigene對(duì)比分析到NR蛋白數(shù)據(jù)庫(kù)中,同源蛋白質(zhì)序列對(duì)比較高的是葡萄(Vitis vinifera)39.5%,可可樹(shù)(Theobroma cacao)10.6%等。有10,374條unigene注釋到COG數(shù)據(jù)庫(kù)中,最多的是基因功能預(yù)測(cè)(General function prediction only)。有9,550條unigene注釋到KEGG數(shù)據(jù)庫(kù)中,其中RNA運(yùn)輸(RNA transport)有381條unigene,核糖體(Ribosome)有360條unigene。通過(guò)對(duì)樣品進(jìn)行差異表達(dá)分析,差異表達(dá)基因5,141條,其中上調(diào)基因3,832條,占差異表達(dá)基因的74.5%,下調(diào)基因1,309條,占差異表達(dá)基因的25.5%。2,728條差異基因注釋在GO功能富集中,得到56組GO功能分類,代謝過(guò)程(Metabolic process)、細(xì)胞過(guò)程(Celluar process)、細(xì)胞(cell)、細(xì)胞部分(cell parts)、催化活性(catalytic activity)、結(jié)合性(binding)等功能的unigene較多。在三大類的topGO功能富集分析中細(xì)胞壁(cell wall)、亞鐵紅素結(jié)合(heme binding)、鎘的響應(yīng)(response to cadmium ion)節(jié)點(diǎn)最為顯著。1,660條unigene注釋在COG注釋分類中,最顯著的是基因功能預(yù)測(cè)(General function prediction only),這些基因在作用機(jī)制上有共同點(diǎn)而類聚在一起,從而推測(cè)基因功能。有1,500條差異基因注釋到KEGG分類中,共有148條代謝通路,最高的代謝通路中是氧化磷酸化(Oxidative phosphorylation)與總unigene KEGG富集分析結(jié)果一致。按照基因功能找出差異性較大的調(diào)控荷花‘青玉’花開(kāi)放的相關(guān)關(guān)鍵基因,促進(jìn)乙烯合成的相關(guān)基因ACC和ERF;影響生長(zhǎng)素合成的相關(guān)基因XTH和YUCCA;調(diào)控細(xì)胞分裂素合成的CKX基因;影響細(xì)胞壁生長(zhǎng)的相關(guān)基因EXP。通過(guò)實(shí)時(shí)熒光定量Real-time PCR方法,以‘青玉’和‘重陽(yáng)’兩個(gè)品種花開(kāi)放的6個(gè)時(shí)期,現(xiàn)蕾期、轉(zhuǎn)色期、暄蕾期、初花期、盛花期、謝花期的花瓣為材料,對(duì)表達(dá)量較高的4個(gè)基因驗(yàn)證分析,在‘青玉’的暄蕾期與盛花期的表達(dá)量的變化與轉(zhuǎn)錄組測(cè)序基本一致,說(shuō)明測(cè)序數(shù)據(jù)真實(shí)可靠。在‘青玉’與‘重陽(yáng)’的花朵開(kāi)放的6個(gè)時(shí)期中,不同基因在整個(gè)開(kāi)花過(guò)程的表達(dá)量變化有所差異,說(shuō)明基因在花開(kāi)放中的不同作用,也推測(cè)不同的基因?qū)ㄐ鸵灿姓{(diào)控作用。本研究揭示了控制花開(kāi)放的關(guān)鍵基因,為延長(zhǎng)花期和荷花育種提供有效分子數(shù)據(jù)信息。
[Abstract]:Lotus (Nelumbo nucifera Gaern) belongs to the genus Nymbiaceae. It is a perennial herbaceous flower and has high ornamental and economic value. The flowering time of lotus is short, and the biological basis of lotus flowering is relatively lagging behind, so it is particularly important to improve the flowering time of lotus. Transcriptional group sequencing technology has become the mainstream of molecular function research, which greatly accelerates the research progress of biological related functions. In this study, the transcriptional levels of bud stage and flowering stage of 'Qingyu' petals were studied by sequencing on a high throughput Illumina HiSeq2500 platform. Finally, the total length of 50154 unigene, was 3, 938, 0851 BP, the average length was 785.20 BP, and the length of N50 was 1412 BP. There are 23753 unigene in the transcript compared with the GO database, of which the three categories contain 56 groups of functions. 32194 unigene were compared and analyzed in NR protein database. The highest homologous protein sequence pairs were grape (Vitis vinifera) 39.5%, cocoa tree (Theobroma cacao) 10.6% and so on. There are 10374 unigene comments to the COG database, the most of which are gene function prediction (General function prediction only). There are 9550 unigene comments into the KEGG database, of which 381 unigene, ribosomal (Ribosome) have 381 unigene, ribosomal (Ribosome) and 360unigene. are transported by RNA. Through the differential expression analysis of the samples, there were 5141 differentially expressed genes, of which 3832 were up-regulated genes, accounting for 74.5% of the differentially expressed genes and 1309 down-regulated genes. It accounted for 25.5% of the differentially expressed genes. 2728 differentially expressed genes were annotated in GO. 56 groups of GO functional classification were obtained, and the partial (cell parts), of (cell), cells of (Celluar process), cells was obtained during the metabolic process of (Metabolic process), cells. 2. 2728 differentially expressed genes accounted for 25.5% of the differentially expressed genes. There are more unigene with catalytic activity (catalytic activity), binding (binding) and so on. Among the three types of topGO functional enrichment analysis, the response (response to cadmium ion) node of cell wall (cell wall), ferricopene combined with (heme binding), cadmium was the most significant. 1660 unigene comments were in the COG annotation classification. The most significant thing is that gene function predicts that (General function prediction only), genes have something in common and cluster together in the mechanism of action, thus speculating on gene function. There were 1500 differentially expressed genes in KEGG classification, and there were 148 metabolic pathways. The highest metabolic pathway was oxidative phosphorylation (Oxidative phosphorylation) and the results of total unigene KEGG enrichment analysis were consistent. According to the gene function, the key genes that regulate the flower opening of lotus' Qingyu 'were found out, and the genes related to ethylene synthesis, ACC and ERF;, which affected auxin synthesis, XTH and YUCCA;, were found. CKX gene regulating mitogen synthesis; EXP., a gene related to cell wall growth By real-time fluorescence quantitative Real-time PCR, the petals of 'Qingyu' and 'Chongyang' were used as materials in six periods of flower opening, bud stage, color conversion stage, bud stage, early flowering stage, full flowering stage and Xie flowering stage. For the four genes with high expression, the changes of expression at the bud stage and flowering stage of 'Qingyu' were basically consistent with those of the transcriptional group, which indicated that the sequencing data were true and reliable. In the six periods of flower blooming of 'Qingyu' and 'Chongyang', the expression of different genes was different during the whole flowering process, which indicated that different genes played different roles in flower opening, and it was speculated that different genes also played a regulatory role in flower type. This study revealed the key genes to control flower blooming and provided effective molecular data information for prolonged flowering and lotus breeding.
【學(xué)位授予單位】：河南農(nóng)業(yè)大學(xué)
【學(xué)位級(jí)別】：碩士
【學(xué)位授予年份】：2016
【分類號(hào)】：S682.32;Q943.2
本文編號(hào)：2481575
[Abstract]:Lotus (Nelumbo nucifera Gaern) belongs to the genus Nymbiaceae. It is a perennial herbaceous flower and has high ornamental and economic value. The flowering time of lotus is short, and the biological basis of lotus flowering is relatively lagging behind, so it is particularly important to improve the flowering time of lotus. Transcriptional group sequencing technology has become the mainstream of molecular function research, which greatly accelerates the research progress of biological related functions. In this study, the transcriptional levels of bud stage and flowering stage of 'Qingyu' petals were studied by sequencing on a high throughput Illumina HiSeq2500 platform. Finally, the total length of 50154 unigene, was 3, 938, 0851 BP, the average length was 785.20 BP, and the length of N50 was 1412 BP. There are 23753 unigene in the transcript compared with the GO database, of which the three categories contain 56 groups of functions. 32194 unigene were compared and analyzed in NR protein database. The highest homologous protein sequence pairs were grape (Vitis vinifera) 39.5%, cocoa tree (Theobroma cacao) 10.6% and so on. There are 10374 unigene comments to the COG database, the most of which are gene function prediction (General function prediction only). There are 9550 unigene comments into the KEGG database, of which 381 unigene, ribosomal (Ribosome) have 381 unigene, ribosomal (Ribosome) and 360unigene. are transported by RNA. Through the differential expression analysis of the samples, there were 5141 differentially expressed genes, of which 3832 were up-regulated genes, accounting for 74.5% of the differentially expressed genes and 1309 down-regulated genes. It accounted for 25.5% of the differentially expressed genes. 2728 differentially expressed genes were annotated in GO. 56 groups of GO functional classification were obtained, and the partial (cell parts), of (cell), cells of (Celluar process), cells was obtained during the metabolic process of (Metabolic process), cells. 2. 2728 differentially expressed genes accounted for 25.5% of the differentially expressed genes. There are more unigene with catalytic activity (catalytic activity), binding (binding) and so on. Among the three types of topGO functional enrichment analysis, the response (response to cadmium ion) node of cell wall (cell wall), ferricopene combined with (heme binding), cadmium was the most significant. 1660 unigene comments were in the COG annotation classification. The most significant thing is that gene function predicts that (General function prediction only), genes have something in common and cluster together in the mechanism of action, thus speculating on gene function. There were 1500 differentially expressed genes in KEGG classification, and there were 148 metabolic pathways. The highest metabolic pathway was oxidative phosphorylation (Oxidative phosphorylation) and the results of total unigene KEGG enrichment analysis were consistent. According to the gene function, the key genes that regulate the flower opening of lotus' Qingyu 'were found out, and the genes related to ethylene synthesis, ACC and ERF;, which affected auxin synthesis, XTH and YUCCA;, were found. CKX gene regulating mitogen synthesis; EXP., a gene related to cell wall growth By real-time fluorescence quantitative Real-time PCR, the petals of 'Qingyu' and 'Chongyang' were used as materials in six periods of flower opening, bud stage, color conversion stage, bud stage, early flowering stage, full flowering stage and Xie flowering stage. For the four genes with high expression, the changes of expression at the bud stage and flowering stage of 'Qingyu' were basically consistent with those of the transcriptional group, which indicated that the sequencing data were true and reliable. In the six periods of flower blooming of 'Qingyu' and 'Chongyang', the expression of different genes was different during the whole flowering process, which indicated that different genes played different roles in flower opening, and it was speculated that different genes also played a regulatory role in flower type. This study revealed the key genes to control flower blooming and provided effective molecular data information for prolonged flowering and lotus breeding.
【學(xué)位授予單位】：河南農(nóng)業(yè)大學(xué)
【學(xué)位級(jí)別】：碩士
【學(xué)位授予年份】：2016
【分類號(hào)】：S682.32;Q943.2
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