【作者】 刘平；

【导师】 彭士琪； 彭建营；

【作者基本信息】 河北农业大学 ， 果树学， 2002， 博士

【摘要】 赞皇大枣是目前发现的枣品种中唯一的自然三倍体品种，品质优良。研究赞皇大枣原生型的地理分布和群体遗传变异可为这一珍贵资源的利用和保护提供理论依据。本研究在地理分布调查的基础上结合染色体数目分析，首次弄清了赞皇大枣的倍性及赞皇大枣原生型的地理分布。同时借助RAPD（Random Amplified Polymorphic DNA）技术研究了赞皇大枣及相关类群的群体遗传多样性、群体遗传结构、群体间遗传学关系，探讨了三倍体赞皇大枣的起源与进化。主要结果如下： 1、首次完成了对有代表性的42个赞皇大枣类型的染色体数目研究，结果表明：供试赞皇大枣均为三倍体，即2n=3x=36。赞皇大枣原生型的主要地理分布区在河北省赞皇县，其中以赞皇县西阳泽乡、院头镇最为集中，南清河乡、城关镇、西龙门乡、张愣乡、黄北坪乡、元氏的苏村乡、高邑的西富村乡、临城的西竖乡等地均有赞皇大枣原生型的地理分布。 2、建立了适于枣及酸枣种质的RAPD优化反应体系：每20μL反应体系中，Tris-HCl（pH8.3）100mmol·L^-1，KCl 50mmol·L^-1，EDTA（pH8.0）20mmol·L^-1，Mg^2-2.0mmol·L^-1；Taq DNA聚合酶1.2U；dNTPs 100μmol·L^-1；引物0.2μmol·L^-1；模板40～60ng。PCR扩增程序为：第一步：94℃ 3min；第二步：94℃ 30s，36℃ 40s，72℃ 1min，40个循环；第三步：72℃ 5min。 3、共利用22个多态性引物（其中20个引物在赞皇大枣各类型之间表现多态性）对赞皇县及周边县区的50个赞皇大枣类型、26个酸枣类型、6个其它枣品种（类型），河北农大标本园1个冬枣和1个未知品种（类型）进行了RAPD分析。共扩增出284个位点，平均每引物扩增13个位点，位点长度在270bp～2500bp之间。多态性位点261个，占总扩增位点数的92％。 4、共有19个供试类型扩增出了特有标记，占供试类型的22.6％，其中有酸枣类型14个。 5、不同引物的鉴别效率不同。鉴别效率最高的引物是S12，鉴别效率达59.5％，其次是S27、S368，鉴别效率分别为53.6％、46.4％。用这三个引物可把供试84个类型全部分开。 6、首次对赞皇大枣及相关类群的群体遗传多样性进行了研究，从多态位点百分率、基因杂合度、基因型种类及基因型频率、群体内遗传距离等几方面进行剖析。 （1）多态位点百分率 赞皇大枣多态性位点百分率为58％，比酸枣的多态性位 赞皇大枣及相关类群群体遗传变异的分了评价点百分率低刀％，比 8个其它枣品种（类型）的多态性位点百分率低 8％。 （2）基因杂合度 赞皇大枣群体的平均杂合度为 0．105，酸枣群体平均杂合度为0．233，其它枣品种（类型）群体的平均杂合度为0．164。 （3）基因型种类及基因型频率 除引物S12、527、S176外，赞皇大枣扩增的基因型种类要少于酸枣扩增的基因型种类，而且基因型频率的分布不均匀。 （4）群体内遗传距离 赞皇大枣群体内部平均遗传距离为 0．057，小于其它枣品种（类型）群体内的平均遗传距离（0．181）和酸枣群体内的平均遗传距离 （0．254）a 7、首次对赞皇大枣及相关类群的群体遗传学结构进行了研究，研究表明：供试3个群体间的基因多样度为0．05，基因分化系数为0．231，说明有23．1％的变异存在于群体间。本研究中，Nm值小于 l（N＝0 83），说明群体间基因流动不足以防止由于遗传漂变引起的群体间的遗传分化。 8、首次对赞皇大枣及相关类群的群体间遗传学关系进行了研究，结果表明： O)群体间遗传一致度和遗传距离 赞皇大枣与酸枣、赞皇大枣与其它枣品种（类型）、酸枣和其它枣品种（类型）的遗传一致度分别为0．881、0．935和0．916；赞皇大枣与酸枣、赞皇大枣与其它枣品种（类型）、酸枣和其它枣品种（类型）间的遗传距离分别为0．127、0．064和0．088，说明赞皇大枣与其它枣品种（类型）的亲缘关系较近，而与酸枣亲缘关系较远。 （2）聚类分析 根据欧氏距离矩阵，利用离差平方和法对供试类型进行聚类分析，结果如下：取人＝6．0，可将供试类型分为2类c第一类包括赞皇大枣50个类型、榆底串杆、元氏串杆、小串杆、前台二号、吕庄串杆；第二类包括紫铃蛋、冬枣、未知品种（类型）和26个酸枣类型。取入＝5．0，可将供试类型分为三类。第一类包括赞皇大枣50个类型、榆底串杆；第H类包括元氏串杆、小串杆、前台1号、吕庄串杆；第H类包括紫铃蛋、冬枣、未知品种（类型）和26个酸枣类型。 综合分析供试群体的遗传学结构和群体间遗传学关系的研究成果，认为枣和酸枣宜作为两个独立的种。 9、从聚类图和欧氏距离矩阵上看，赞皇大枣与赞皇县境内及周边县区的串杆类型亲缘关系较近，赞皇大枣与的起源可能与赞皇县及附近的串杆枣有关。更多还原

【Abstract】 Zanhuangdazao (Ziziphus jujuba Mill.) was known as a unique triploid cultivar with best quality. Studies on geographic distribution and genetic variation of Zanhuangdazao original forms could provide theoretic evidences for its utility and protection. By combing its geographic distribution with analysis of chromosome number, geographic distribution of Zanhuangdazao original forms was studied for the first time. Genetic diversity > genetic structure^ genetic relationships between populations ^ origin and evolution of Zanhuangdazao and its related populations was also analyzed by using RAPD(Random Amplified Polymorphic DNA) technique in the paper. Main results were as follows:1 .Chromosome number of 42 Zanhuangdazao forms was analyzed for the first time, which suggested that all Zanhuangdazao forms tested were triploid, that was, 2n=3x=36. Zanhuangdazao original forms distributed in Zanhuang county of Hebei province mainly, make Xiyangzcx Yuantou as the towns distributed denselly, next to which were Nanqinghes Chengguan> Xilongmem Zhanglengx Huangbeiping of Zanhuang county > Sucun of Yuanshi county ^ Xifucun of Gaoyi county and Xishu of Lincheng county.2.Optimum system of RAPD amplification for Chinese jujube and wild jujube was established. In 20ul reaction solution: Tris-HCl(pH8.3) lOOmmol ?L"’,KC1 50 mmol -L"’,Mg2+ 2.0 mmol -L"1; Tag DNA polymerase 1.2U; dNTPs 100 pi mol I/1; Primer 0.2 p mol ?L"1; Template 40~60ng. The amplified procedure was : Step 1: 94癈 3 min; Step 2: 94癈30s,36癈40s,72癈 lmin,40 cycles; Step 3:72"C5min.3. Total 22 primers, among which 20 were polymorphic between Zanhuangdazao forms ,were used in RAPD amplification of 50 Zanhuangdazao forn^ 26 wild jujube formsN 6 Chinese jujube cultivars or forms in or nearby Zanhuang county, one Dongzao and an unknown cultivar or form in garden of Agriculture university of Hebei. 284 loci were gained among which 261 loci were polymorphic. Average loci number amplified per primer and length of loci were 13 and 270bp~2500bp respectively.4.Total 19 forms had specific RAPD markers which was 22.6 percentage of the forms tested. Among the 19 forms , there were 14 wild jujube forms.S.The identification efficiency of 22 primers differed with each other. The identification efficiency of primer S12 was highest(59.5%), and then primer S27> S368, whose identification efficiency was 53.6% and 46.4% in order. All the 84 forms tested could be identified by using the three primers.6.Genetic diversity of Zanhuangdazao and its related populations was studied for the first time in several aspects, they were percentage polymorphic loci, gene heterozygosity, kinds and frequency of genotypes^ genetic distance within population.(1 percentage polymorphic loci: Percentage polymorphic loci ofZanhuangdazao(58%) was lower than that of wild jujube and 8 other Chinese jujube cultivars or forms by 31% and 8% respectively.(2)Gene heterozygosity: Gene heterozygosity of Zanhuangdazao population, wild jujube populations, other Chinese jujube cultivars or forms population was 0.105, 0.233 and 0.164 in order.(3)Kinds and frequency of genotypes: Genotypes kinds amplified among Zanhuangdazao forms was less than that of wild jujube except by primer S12^S27>S176. What’s more, the distribution frequency of genotypes in former was also uneven.(4)Genetic distance within populations: Average genetic distance in Zanhuangdazao population was 0.087,which was less than that of Chinese jujube cultivars or forms population (0.181) and wild jujube population (0.254).7.Genetic structure of Zanhuangdazao and its related populations were studied for the first time. The main results were: DST value and GST (coefficient of gene differentiation) of 3 populations was 0.05 and 0.231 respectively, which indicated that 23.1% variation partitioned among populations. Nm<l(Nm=0.83) mean that gene flow among populations was too small to prevent genetic differentiation caused by genetic drift.8.Genetic relationships among Zanhuangdazao and its rela更多还原