【作者】 徐立；

【导师】 余茂德；

【作者基本信息】 西南农业大学 ， 特种经济动物饲养， 2003， 硕士

【摘要】 蚕桑生产是全国许多农村脱贫致富的支柱产业，蚕丝品质是国际市场竞争的重要基础。桑叶的产量，质量直接影响茧丝的产量与品质。 在桑树品种中，人工选育的三倍体新桑品种（2n=3x=42）在经济性状和农艺性状上比二倍体桑品种（2n=2x=28）和自然三倍体桑（2n=3x=42）更为优良。因此，选育人工三倍体桑已成为国际国内桑树品种选育的重要方向。要选育产量高、叶质优的新型人工三倍体桑树新品种，必须大量的选择杂交亲本和在杂交后代中进行大量的筛选，因此，有必要弄清育种亲本的遗传背景和性状遗传规律。但桑树属多年生异花授粉木本植物，基因型高度杂合，生命周期长，植株个体占地面积大，其性状遗传规律研究难度较高，采用常规方法，很难弄清其遗传背景。本研究利用DNA多态性检出率较强的分子标记技术AFLP（Amplified Fragment Length Polymorphism），即扩增片段长度多态性，从DNA分子水平对人工三倍体新桑品种嘉陵16号（2n=3x=42）及其亲本，人工三倍体新桑品种嘉陵20号（2n=3x=42）及其亲本以及与嘉陵20号同父、母本的2个三倍体（2n=3x=42）桑品系与10个二倍体（2n=2x=28）桑树材料进行了遗传背景分析。获得了它们的指纹图谱、遗传距离及遗传相似系数并绘制了它们的UPGMA聚类图，为今后人工三倍体新桑品种选育、杂交亲本的选择与组配提供较有用的参考依据，对选育更具强大杂交优势和多倍体优势的新型人工三倍体桑品种，将起到重要辅助作用。现将其研究结果要点总结如下。 1、本实验通过对嘉陵16号（3X；F₁）和嘉陵20号（3X；F₁）两个人工三倍体新桑品种及它们的亲本进行了AFLP分析，计算了其遗传距离与遗传相似系数，研究结果发现：嘉陵16号的母本西庆一号（4X；♀）与父本育二号（2X；♂）之间的遗传相似系数为0.6607；嘉陵20号的母本7920（2X；♀）与父本桐乡青（4X；♂）间的相似系数为0.6612。根据嘉陵16号和嘉陵20号两个人工三倍体新桑品种及其亲本的AFLP分析结果，我们认为人工三倍体新桑品种选育以及桑树杂交育种亲本间的遗传相似系数以0.6600左右较为合适，这可以作为今后多倍体育种与杂交育种中，选择组配杂交亲本的参考依据。 2．本实验通过对嘉陵16号（3X；FI）及其亲本u)以及嘉陵20号（3X：FI）、B（3X；FI）。C（3X：FI）、10个二倍体（ZX；FI）及其亲本0)的AFLP分析结果发现，所有的F；代都与二倍体亲本的亲缘关系相对较近，可能二倍体与同源四倍体杂交的后代（FI）从二倍体亲本处所继承的遗传物质比从同源四倍体亲本处继承的要多一些。因此，在今后人工多倍体育种和杂交育种中的亲本选配时，要特别重视二倍体亲本的选择，应选择生活力强、经济性状、农艺性状、生物学特性都较优的二倍体作为杂交亲本。 3、根据本实验对两个人工三倍体新桑品种及其亲本的遗传背景的AFLP分析结果，结合前人的研究，发现在人工三倍体桑树品种选育中，亲本的育性、杂交后代的种子发芽率、遗传传递力等方面，其二倍体（ZX）交四倍体（4X）的组配方式优于四倍体（4X）交二倍体（ZX）的组配方式。因此，在桑树多倍体育种中杂交亲本组配时，将具有优良性状的二倍体亲本作为杂交母本 （旱）较好。 4、通过对人工三倍体新桑品种嘉陵20号（3X；FI）及其亲本以及与嘉陵20号（3X；FI）同亲本的2个3X材料和 10个ZX材料的llP分析，并结合前人对其细胞染色体的分析，认为在桑H倍体（2—2——28）交四倍体（小X；下）或四倍体交二倍体的杂交厅代之所以出现三倍体（2。3——42）、二倍体等多样性变化而且三倍体之间和二倍体之间都出现了丰富的多态性和复杂的聚类现象，是因为同源四倍体四同源组的几条染色体发生多种形式的不均衡分离，造成了同源四倍体的配子内染色体数和组成的不平衡，从而造成同源四倍体的部分不育及其子代染色体数、遗传物质DNA等的多样性变化。通过对这3个3X材料和10个ZX材料与父母本之间的遗传距离和遗传相似系数以及它们的UPGMA聚类图的比较分析，提出在人工三倍体新桑品种选育中，可利用AFLP技术分析F；代植株的遗传背景，鉴别F；代植株是否是育种所需要的三倍体植株的技术途径，以达到早期鉴定的目的。 5、在本实验中，通过对供试材料的AFLP分析，发现无论是用二倍体（ZX）桑作母本（g）还是作父本（占），桑树二倍体亲本（＊X；P）的遗传传递力都强于四倍体亲本（4X：P），n代的个体与二倍体亲本的亲缘关系较近。根据本研究的结果，结合前人的研究，提出了桑属植物倍数性演化的推测，外 桑树最先起源于二倍体，再由二倍体桑（ZX）衍生出如自然单倍体（力、自然四倍体O）、自然六倍体（6X）、自然八倍体（SX） 自然二十二倍体（22X）等多种类型。当然，这一推测还有待于今后更多更深入的研究证实。 6、为了使所提DNA的质量能满足AFLP实验的要求，保证实验结果可靠。本实验采用斑迹抽提液附加［一mercaptoethanol、Proteinase K和RN ase作为抽提液，来提取桑叶基因组DNA，获得了质量较高的基因组DNA，A260／A280比值为1．80—1．90之间；其产量为133 V g／g～160 ug／g之间，通过琼脂糖凝胶更多还原

【Abstract】 The industry of silkworm and mulberry is of great importance for formers to get rid of poverty and become rich in some areas of China.The quality of silk is the main basis of competence international wide .Nevertheless, the output and quality of silk are affected directly by the yield and quality of mulberry leaves.The breeding of artificial triploid mulberry (2n=3x=42) has become the highlight in the breeding of mulberry because of its excellent economic charaters and agronomic charaters which are prior to that of diploid mulberry (2n=2x=28) and natural triploid mulberry (2n=3x=42).In order to obtain new breeding which high production and good quality of leaves, it is necessary to study the breeding parents and its genetic backgroun.But mulberry is a kind of ligneous plant which is parennial, extendotropic and having high heterozygosis in genotype. So, it is difficult to study its genetic laws and genetic background with common methods. In this study , genetic background of Jialing16, Jialing20, their parents and Jialing20 brothers are analyzed by the AFLP(Amplified Fragment Length Polymorphism),from which the genetic distance , similarity coefficient and UPGMA dendrogram of the materials are obtained.These will provide reliable evidence for the selection and tissue mating of parents. These will be helpful in the breeding of new artificial triploid mulberry with strong heterosis and miltiploid superiority. Results and conclutions of study are as follows:1. By AFLP analysis of Jialing16, Jialing20 and their parents , it is found the similarity coefficient between 7920 (2X: o)and Tongxiangqing(4x: o ), which are parents of Jialing20, is 0.6612 and the similarity coefficient betweenXiqingl (4x: o ) and Yu2 (2X: o ), which are parents of Jialing16, is 0.6607.By studying AFLP analysis of Jialing16 , Jialing20 and their parents.We come to the conclusion that the ideal genetic similarity coefficient between the mother and the father of F1 is about 0.6600, which can be seen as an evidence reference for the selection of hybrid parents in theprocess of multiploid and hybrid breeding .2. By AFLP analysis of Jialingl6 and their parents , Jialing20 A (3x;F1), B(3x;F1), C(3x;F1), 10 diploids(2x;F1) and their parents (P) .It can also be found that the FI generation has nearer genetic relationship with diploid parents.It is probably because that generation gets more genetic substance from diploid parents than that from teroploid parents.As a result , it should be put more importance on the selection of diploid parents, which should has high quality in economic character , argronomic character and with strong vital capcity.3. By analysis, it has been noted that the filial generation are better in fertility seeds germination capacity and genetic transmission capacity when taking diploid as female parents better than taking diploid as male parents.So, it is better to choose diploid as female parents when pairing the hybrid parents.4. By AFLP analysis of this study , according to former study on its chromosome at the cellular level as well, it is considered that the reason for diversity of diploid (3x)and triploid(2x)in F1 generation of 2xX4x or 4xX2x and the reason for high polymorphism and complex dendrogram between dilploidsand triploids are that unbalanced separation has taken place in the number and the content of chromosomes of autotetraploid, which cause partial sterility and diversity in genetic substance and the number of its progeny’s chromosome.By the analysis of genetic distance. Genetic similarity coefficent and their UPGMA dendrogram of the 3 triploids, 10 diploid and their parents. We suggest , in order to obtain early indentification AFLPanalysis may be used to analyse genetic banckground of F1 generation and identify Fl whether is the proper one that we needed.5. According to the results of this study and former studies , we propose a hypothesis about the diplosis evolution of mullberry derived from diploid at first, then derivated other kinds of indi更多还原