【作者】 陈静；

【导师】 任正隆； 余懋群；

【作者基本信息】 四川农业大学 ， 作物遗传育种， 2001， 博士

【摘要】 与大多数作物一样，由于受高度集约化和农业现代化生产的影响，栽培小麦遗传基础日益狭窄。小麦遗传背景单一，不仅限制了其在产量和品质上的进一步改良，而且使品种抗生物和非生物胁迫的能力逐步下降。小麦野生近缘植物中存在大量的遗传变异，是小麦改良可以利用的丰富有益基因资源。在小麦的近缘种属中，簇毛麦（D.villosum，2n=14，VV）对小麦白粉病表现极高的抗性，并具有多蘖、耐旱、高蛋白含量等多种优良特性。黑麦（S.cereale L.，2n=14，RR）是小麦遗传改良的又一重要外源基因供体，1RS上含有与抗多种病害和提高小麦产量有关的基因，1RS/1BL易位系曾经作为重要的抗源亲本成功地用于小麦育种。尽管1RS/1BL易位系的抗性已经丧失，由于其对品种产量潜力和稳定性的贡献，在小麦育种实践中仍然受到育种家的重视。本研究利用多种分子、细胞生物学技术和遗传分析的方法对探索转移外源基因新方法的理论问题、利用ph1b遗传体系直接遗传转移簇毛麦基因到普通小麦的可能性，并对具有小麦、簇毛麦、黑麦优良基因的三属杂种种质材料的选育，鉴定及其细胞遗传学特点等方面进行了研究，结果如下： 1.利用不同Brdu溶液浓度和不同培养时间组合处理簇毛麦和黑麦根尖有丝分裂细胞，以观察植物染色体上脆性位点的分布。结果发现：仅在50μg/mlBrdu溶液20～21h的培养条件下，9.3％的黑麦根尖细胞染色体产生脆性位点表达，主要分布在三条黑麦染色体的四个不同位点上，频率分别为1.4％、2.1％、4.3％和5.7％。不同浓度Brdu均未能诱导簇毛麦染色体脆性位点表达。这一结果不仅证明植物染色体上也具有脆性位点，而且多条黑麦染色体上脆性位点的表达可能与黑麦进化过程中复杂的染色体易位现象有关，从而为我们利用这一脆性位点诱导体系，通过染色体断裂—重新融合建立外源基因转移的新方法带来启示。 2.对中国春，中国春ph1b突变体与簇毛麦杂种F₁花粉母细胞减数分裂中期Ⅰ染色体配对的观察结果表明，（CSXD.villosum）和（CSph1bXD.villosum）F₁杂种平均染色体配对构型分别2n=28=26Ⅰ+1Ⅱ（Xta=0.84）和2n=28=13.55 I＋5．95II＋0．55ill＋0．22IV（xta二9．72），phlb突变体与簇毛麦杂种 F；中部份同 源染色体配对明显增加。将基因组荧光原位杂交技术GISH)用于杂种 F；u七8， ABDV)的染色体配对分析，清晰、直观地揭示出的 基因对小麦与簇毛麦染色 体部分同源配对具有较强的促进作用，其配对类型和频率分别为 0．98 W（小支） －D（簇毛麦），0．11w－W－D，0．02 D－W－D。phlb基因强烈地诱导了小麦剖份 l。Ji 染色体配对。同时，这一结果也说明簇毛麦中不存在Ph或Ph he基冈。 3．在小麦与簇毛麦杂种F；小抱子发生过程中，观察到较多的减数分裂汁 常现象。杂种厂自交严重不育。CS X g VjllOSurk F杂种回交结实率为 6．67见 ＋肚b之下，山于Ph b基因干扰了杂种厂减数分裂的恢复分裂，（CSph o X fi。1”11OSugh‘．的凹交结实率大大降低，只有 0．61％。 4．尽管杂种 F；回交困难，在含有的 fo组合的回交一代仍然获得了 7株 11C （Zn＝48－72）植株。在 Zn＝72植株根尖细胞有丝分裂 GISH观察中发砚一对小交 －簇毛麦染色体臂间易位以及一个可能的小片段易位，从而证实利用Phlb 11）”以 简单、快速、有效地实现簇毛麦基因向小麦的转移。此外，还对2n3犯的l儿 植株形成的可能途径进行了讨论。 5．通过普通小麦与簇毛麦杂交、回交，选育获得了抗白粉病、优质的小k 种质材料 ADI 34（Zn二42＋Zt）。该材料整个生育期对白粉病免疫，分醛！［t盛，成 穗数较高（平均 7——8个），籽粒蛋白质含量高达 18％，并具有与面包烘烤品质有 关的5＋10高分子量谷蛋白优质亚基，但存在秆高，结实率低和种于皱缩的不良 性状。 6．综合运用抗病性鉴定、GIsH技术、C－分带、APAGB、SCAR叫’oR多种技木， 对AD134的染色体组成进行了准确鉴定。结果表明，AD134是含有小麦、淑。方支 与黑麦染色质的非整倍性三属杂种，附加端体为簇毛麦染色体 6V短臂“VU， 在长期选育的过程中6VS的端部发生了部分缺失。AD134所携的一对小麦一黑在 易位染色体IRS／IBL来自含有IRS／IBL易位的回交亲本小麦，该染色体臂IllS h的抗n粉病拭因pms抗性已经丧失或受小麦遗传背景的抑制不能人达。 7．对AD134与感白粉病小麦正、反杂交的F；代进行抗病性鉴定分析，衣叫 其抗白粉病特性为显性，受6VS上抗病基因的控制，山于本研究所用簇毛支供 体与现有6V／6A（6D）系的簇毛麦亲本来源不同，存在一定遗传差异。ADI：14Ij 能是一有潜?更多还原

【Abstract】 As in most other crops, the genetic variation of cultivated wheat has been eroded under modem agricultural system. Narrow genetic background not only makes wheat increasingly vulnerable to biotic and abiotic stress, but also limits the further improvement of yield and quality. A large of genetic variation in the wild relatives of common wheat are useful resources for wheat improvement. D.villosutn (2n=14,VV), a wild related species of wheat, possesses attractive traits, such as high resistance to powdery mildew, abundant tiller, dough tolerance, high protein content. S.cereale (2n=14,RR), another important alien gene donor, possesses many useful characteristics including resistance to several wheat diseases and tolerance to adverse environment, as well as high yield porential controlled IRS. I R/l B translocation lines as important resistant parents have been used in wheat breeding successfully. In spite of their resistance loss, wheat breeders still think highly of IR/IB lines for their contribution to variety yield and stability. Using many molecular cytobiological techniques and genetic methods, The aim of this study was to search for the preliminary theory of a new method to transfer alien gene and the possibility of direct genetic transfer of genes from D. villosum to wheat using ph lb mutant, as well as the development, identification and cytogenetic characteristics of a trigeneric hybrid germplasm with useful genes from Tavestivum, D.villosum and S.cereale. Results are shown as follows: 1. The miotic root-tip cells of rye (S. cereale L.) and D. villosum were treated under different combinations of Brdu solution concentrations and times for culture. Results indicated that chromosomal fragile sites were observed in 9.3% cells of rye and appeared on four locations of three rye chromosomes, with 1.4%, 2.1%, 4.3% and 5.9% respectively, only afler vigor root-tip cells were treated with Brdu (5OugIml) for 20-21 h, but were not on D. villosum chromosomes under any treatment. This result not only showed that there were fragile sites on plant chromosomes, but also suggested that expresstion of fragile sites on many rye chromosomes could be related to complicated chromosomes translocation in the evolution of rye, and alien gene transfer through reak-reunion?could be induced by the expression of chromosome fragile sites. 2. Meiotic chromosome pairing analysis revealed that the average chromosome 92 configuration were 2n~2813.55 I +5.95 II +0.55111+0.22 IV (Xta=9.72) and 2n2826 I +111 (Xta0.48) in CSphIbXD.villosum and CSph X D.villosum F1 hybrids respectively,indicating that homoeologous chromosome pairing increasing dramatically in PMCs of F1 hybrid involving ph lb. Meiotic GISH analysis of Fl hybrids with phlb gene further detected three types of chromosome pairing with different frequencies, that is, 0.98 W-D, 0.11 W-W-D and 0.02 D-W-D, showing that phlb greatly promoted homoeologous chromosome pairing between wheat and D.villosum, but more dramatically between wheat genomes. P1db or Phlb-like gene didn’t existed in D.villosum, 3. Many abnormal meiotic phenomena were observed during microsprorogenesis of F1 hybrid of wheat and D. villosum. F1 hybrids were self-sterile seriously. Decreased backcross seed set of F1 hybrid with phlb, only 0.61%, was due to phlb interfering the meiotic restitution division compared to 6.67% of F1 hybrid without phlb,. Although the backcross更多还原