發(fā)布時間：2018-03-21 17:14

  本文選題：種質(zhì)資源　切入點：櫻桃　出處：《四川農(nóng)業(yè)大學》2015年碩士論文　論文類型：學位論文

【摘要】：中國櫻桃(Cerasus pseudocerasus Lindl.)、歐洲甜櫻桃(Cerasus avium Linn.)和毛櫻桃(Cerasus tomentosa Thunb.)隸屬薔薇科(Rosaceae)李亞科(Prunoideae)櫻屬(Cerasus),是櫻桃的3個重要栽培種。目前對其種間系統(tǒng)關系和種內(nèi)遺傳多樣性缺乏基于分子序列的相關深入研究。本研究以中國櫻桃、甜櫻桃和毛櫻桃共95份材料的核糖體內(nèi)轉(zhuǎn)錄間隔區(qū)(ITS, internal transcribed spacer)序列作為研究對象,通過分析其序列變異位點,并構(gòu)建進化樹,探討櫻桃種內(nèi)遺傳多樣性和它們的系統(tǒng)發(fā)育關系,并做二級結(jié)構(gòu)預測驗證分析,以揭示中國櫻桃、甜櫻桃和毛櫻桃種內(nèi)遺傳多樣性和種間遺傳關系,并探討ITS序列在櫻桃遺傳多樣性及其相關類群的系統(tǒng)進化研究中應用價值。主要的研究結(jié)果如下：(1)ITS序列特征。中國櫻桃擴增的ITS序列長度范圍為702-703bp,甜櫻桃的為704-705bp,毛櫻桃為709bp,長度變異較小。所有擴增序列對位排列之后長度為712bp,該片段CG核苷酸堿基占總核苷酸堿基的58.1%。共檢測到71個變異位點,其中54個屬簡約信息位點。這些變異位點在ITS1 (37bp-270bp)區(qū)中有40個,5.8S (271bp-373bp)區(qū)中有3個,ITS2 (374bp-651bp)區(qū)有28個。共定義了37個單倍型,其單倍型多樣性Hd=0.911,核酸多樣性π=0.01444,分離位點多態(tài)性平均數(shù)θw=0.01954。(2)中國櫻桃與歐洲甜櫻桃的ITS序列之間的進化距離為0.011-0.027,歐洲甜櫻桃與毛櫻桃之間的為0.047-0.051,中國櫻桃與毛櫻桃之間的為0.044-0.057。從ITS序列數(shù)據(jù)計算的結(jié)果來看,中國櫻桃與甜櫻桃的ITS序列之間同源性較高,毛櫻桃則與它們同源性較低。(3)NJ系統(tǒng)發(fā)育分析將37個單倍型分為3個分支,Clade I包括中國櫻桃的21個單倍型,并獲得了93%的自展支持率,Clade II包括了甜櫻桃的11個單倍型,自展支持率為96%,Clade III為毛櫻桃的5個單倍型構(gòu)成,自展支持率為100%。(4)在整個單倍型網(wǎng)絡中介圖中,37個單倍型基因樹的拓撲結(jié)構(gòu)呈三臂螺旋分布,毛櫻桃的5個單倍型處于臂長較長的一端。網(wǎng)絡中介圖中共有11處發(fā)生沖突,分別位于排列位點79、88、100、130、135、136、153、185、538、641處。在中國櫻桃分支中單倍型H2處于中心位置,而且大部分群體中都有分布,是核心的古老單倍型,其余單倍型則分散于該分支的外圍,成為網(wǎng)絡圖的外部節(jié)點。(5)二級結(jié)構(gòu)分析。根據(jù)進化樹中位置和單倍型所代表的樣品數(shù)量我們選擇了H1、H2、H22、H37四個單倍型進行二級結(jié)構(gòu)預測分析,5.8S區(qū)域的二級結(jié)構(gòu)基本相同,ITS1區(qū)和ITS2區(qū)的二級結(jié)構(gòu)差異較大,中國櫻桃的單倍型H1、H2最小自由能為-248.00 kcal/mo1和-248.50 kcal/mol,甜櫻桃的單倍型H22最小自由能為-251.60kcal/mol,毛櫻桃的單倍型H37的最小自由能為-256.5 kcal/mol。將得到的最小自由能進行T檢測,三個物種的單倍型之間的最小自由能在P0.05時差異顯著。它們的下半部分的莖環(huán)結(jié)構(gòu)較為相似,上半部分則有較大差異,結(jié)合最小自由能數(shù)據(jù)和結(jié)構(gòu)圖,支持基于ITS序列所做的進化樹。
[Abstract]:Cerasus pseudocerasus Lindl.China (Cerasus avium Linn.) and Cerasus tomentosa Thunb.) belong to the genus Prunoideae (Rosaceae), and are three important cultivated species of Cherry. At present, the interspecific phylogenetic relationship and intraspecific genetic diversity are not based on subdivision. Further study on the correlation of subsequences. The internal transcribed spacer (internal transcribed spacer) sequences of 95 samples of sweet cherry and hairy cherry were studied. The genetic diversity and phylogenetic relationship among them were studied by analyzing the sequence variation sites and constructing an evolutionary tree. In order to reveal the intraspecific genetic diversity and interspecific genetic relationship of Chinese cherry sweet cherry and hairy cherry the secondary structure prediction and verification analysis was done to reveal the genetic diversity of Chinese cherry sweet cherry and hairy cherry. The application value of ITS sequence in the phylogenetic study of cherry genetic diversity and its related groups was discussed. The main results were as follows: the ITS sequence length of Chinese cherry was 702-703bpand that of sweet cherry was 704-705bp. The length of Cherry was 709 BP, the length of which was 712 BP after all the amplified sequences were arranged in opposite position. The CG nucleotide base of this fragment was 58.1% of the total nucleotide base. A total of 71 mutation sites were detected. Among them, 54 belong to parsimonious information loci. These variation loci are 28 out of 40 in the ITS1 37bp-270bp) region, and there are 3 ITS2,374bp-651bp) regions. 37 haplotypes have been defined. Its haplotype diversity is 0.911, nucleic acid diversity 蟺 0.01444, and the average polymorphism of segregation sites 胃 w 0.01954.02) the evolutionary distance between Chinese cherry and European sweet cherry ITS sequence is 0.011-0.027, between European sweet cherry and hairy cherry is 0.047-0.051, between Chinese cherry and hairy cherry is 0.047-0.051, between Chinese cherry and European sweet cherry is 0.047-0.051, between European sweet cherry and European sweet cherry is 0.047-0.051. Between 0.044-0.057. from the results of the ITS sequence data calculations, The ITS sequence of Chinese cherry and sweet cherry was highly homologous, while that of hairy cherry was low. NJ phylogeny analysis showed that 37 haplotypes were divided into 3 branches, including 21 haplotypes of Chinese cherry. The self-development support rate of 93% was 93%, which included 11 haplotypes of sweet cherries, and the self-exhibition approval rating of 96 Clade III was 5 haplotypes of hairy cherries. In the whole haplotype network intermediary map, 37 haplotype gene trees have a three-arm spiral distribution, and five haplotypes of hairy cherry are at one end of longer arm length. There are 11 conflicts in the network intermediary diagram. The haplotype H2 is located in the central position of the Chinese cherry branch, and is distributed in the majority of the population, which is the core of the ancient haplotype, while the other haplotypes are scattered in the periphery of the branch. Based on the location of the evolutionary tree and the number of samples represented by the haplotype, we selected four haplotypes H1H2H2H22H37 to predict the secondary structure and analyze the basic phase of the secondary structure in the 5.8S region. The secondary structure of ITS1 region and ITS2 region were different greatly. The minimum free energy of haplotype H _ 1O _ 2 was -248.00 kcal/mo1 and -248.50 kcal / mol, that of sweet cherry was -251.60 kcal / mol, and that of haplotype H37 was -256.5 kcal / mol. The minimum free energy (MEF) of the haplotypes of the three species was significantly different when P05. The structure of the stem ring in the lower part of the species was similar to that in the upper part, which combined with the minimum free energy data and the structure diagram. Support for evolutionary trees based on ITS sequences.
【學位授予單位】：四川農(nóng)業(yè)大學
【學位級別】：碩士
【學位授予年份】：2015
【分類號】：S662.5

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