【作者】 王黎明；

【导师】 魏湜；

【作者基本信息】 东北农业大学 ， 作物栽培学与耕作学， 2003， 硕士

【摘要】 将具有植株高大、茎秆多汁、含糖量高、抗病、生物产量高且生育期长等特点的热带高粱DNA通过花粉管通道导入到黑龙江省当地高粱品种中，以探讨此项技术用于高粱育种的可行性。 于受体开花受粉后60min左右剪掉柱头，将供体DNA滴于切口处。导入后代出现了数量性状及质量性状等诸多性状的变异，变异性状包括农艺性状、育性、抗病性、含糖量及籽粒品质性状等。各性状的变异比率不同，且D₂代的质量性状分离比例不符合孟德尔遗传规律。 对导入后代的品质性状的测定表明，有4个导入后代的淀粉含量高于受体，占总数的6.9％；3个导入后代的支链淀粉含量高于受体，占总数的5.2％；8个导入后代的单宁含量低于受体，占总数的11.4％。且出现了后代含量均高于或低于供体与受体的结果。 将含糖量高的高粱DNA导入后，获得了15个高于受体含糖量的导入后代，占总数的7.9％。其中有8个导入后代的含糖量既高于受体，又高于供体，这是常规育种所不能得到的。 将抗病性高于受体的DNA导入后，在导入后代中有27个导入后代的抗病性有所提高，占导入后代总数的9.1％，其中5个导入后代的抗病性提高了2级，3个导入后代的抗病性提高了3级，极显著地改变了受体的抗病性。 利用导入后代配制杂交种后，部分不育系、保持系及恢复系的育性均发生了改变。同时导入后代的变异性状可以在杂交后代中表现出来，说明导入后所引起的性状变异是可遗传的。 为了对导入后的变异后代进行验证，对导入后代进行了酯酶及过氧化物酶同工酶的测定。结果表明，导入后代的酶谱明显不同于受体，供体的酶带不同程度地出现在导入后代中，且出现了酶活性减弱及酶带缺失等现象，证明供体DNA片段已进入受体，并参与受体DNA的合成。这与导入后代性状的剧烈分离现象相符。 同时对导入后代进行了RAPD分子验证。在后代的扩增产物中出现了供体具有，而受体没有的特异带，说明供体DNA片段已通过花粉管通道进入受体，并对受体基因的表达产生影响。 利用外源DNA导入技术获得了常规育种无法得到的变异后代，实现了地理远缘的品种间的遗传物质转移，丰富了遗传类型，表明此项技术用于高粱的种质创新和品质改良是可行的，可作为常规育种的一个辅助手段。更多还原

【Abstract】 Exogenous DNA of tropical sorghum with characters of tall plant height, more juice in haulm, high sugar content, diseases resistance, high biomass and long growth duration, was introduced directly into local sorghum varieties through pollen tube pathway, in order to study the feasibility of this technique used in sorghum breading.Stigma was cut when the receptor had been self-pollinated about 60 min, and dripped foreign DNA on the nick. Many characters could be variable, not only quantitative characters, but also qualitative ones, including agronomic characters, fertility, diseases resistance, sugar content, and kernel quality characters. The transferred frequencies of every variable characters were different. The regularity of segregation of qualitative characters in D2 was not accord with Mendel’s regularity of segregation.The determination of quality characters indicated that, 4 transferred progenies’starch content was higher than their receptors, accounting for 6.9 %. 3 transferred progenies’ amylopectin content was higher than their receptors, accounting for 5.1%, 8 transferred progenies’ tannin content was lower than their receptors, accounting for 11.4%. Some transferred progenies’ contents were both higher and lower than their receptors and donors.15 transferred progenies’ sugar content was higher than their receptors, accounting for 7.9%. Of the 15 transferred progenies, 8 transferred progenies’ sugar content was higher than their receptors and donors. These progenies can’t be obtained in conventional breeding.27 transferred progenies’ diseases resistance was increased because of the DNA transferring, accounting for 9.1%. In which 5 transferred progenies’ diseases resistance increased 2 grade, and 3 transferred progenies’ diseases resistance increased 3 grade. The disease resistant of receptor was increased significantly.When the transferred progenies were used in cross breading, some of the sterile lines, maintenance lines and restoring lines’ fertility was changed. Meanwhile, variant characters of their transferred progenies could be appeared in filial generation. This indicated that the variant characters after transference were hereditable.To check the variant progenies, POD and ES isoenzymes were analysed in transferred progenies. The results indicated that the transferred progenies’ bands of isoenzyme were different from their receptors. The donor’s bands of isoenzyme appeared in the transferred progenies in various extends. The activity of isoenzymes was decreased and the bands of isoenzymes were lost. This proved that the DNA fragment of donor had entered to the receptor, and participate in DNA synthesis of receptor. This conformed to the transferredprogenies’ crazy segregation phenomenon.The transferred progenies were checked with RAPD. The specific bands were obtained in progenies’ amplification primers, and the bands didn’t exist in receptor. They inserted into receptor through pollen tube, and affected the expressing of genes in receptors.Variant progenies were obtained by exogenous DNA introduction. And these progenies couldn’t be obtained in conventional breeding. It realized the transference of inheritable matter between varieties of distant geography, and made the inheritable types abundant. The results indicated that it was possible to use this technique to create new germplasm and improve attributes by using the pollen tube passage to introduce exogenous DNA. This method can be used as a complementary method of conventional breeding.更多还原