【作者】 高立宏；

【导师】 王丕武；

【作者基本信息】 吉林农业大学 ， 作物遗传育种， 2004， 硕士

【摘要】 利用稻瘟病菌培养液提取粗毒素作为选择压，通过组织培养途径对水稻抗瘟性进行细胞突变体筛选，是抗病育种的一种新的方法，与常规育种技术相结合，可以培育新的抗病品种。本实验在2002年～2004年，利用目前稻瘟病优势小种菌株产生的粗毒素混合液对5个水稻品种的幼穗、幼胚、成熟胚三种外植体进行处理，研究稻瘟病粗毒素对水稻愈伤组织诱导及分化的影响，以期为东北水稻抗稻瘟病突变体筛选和水稻抗病品种的选育提供理论依据。取得的主要研究成果如下： 1．稻瘟病菌培养及毒素测定 1)不同的菌株在同一培养基中的生长状况差异显著；不同的的培养基在培养同一菌株时效果也不同；不同的稻瘟病菌生理小种其生长量和颜色各不相同(白色、灰色、褐色、黑色为主，有变种后的呈现黄色或黄绿色)。 2)稻瘟病菌提取液的毒性在浓度在0～50％时，随着毒素浓度的提高毒性增加，当浓度为80％左右时，有峰值出现，而后随着毒素浓度的提高而毒性减弱。在0～100％的整个浓度范围内毒性曲线呈现抛物线形状。 3)毒素毒性大小随培养方法和培养中毒素量不同而有差别。 4)不同的生理小种培养的毒素其毒性具有显著性差异。 2．以稻瘟病菌粗毒素提取液为选择压进行水稻抗稻瘟病突变体筛选 1)稻瘟病菌粗毒素提取液对水稻成熟胚组织培养的影响 粗毒素液对水稻胚芽的生长、胚的诱导、愈伤组织的生成及分化有严重的抑制作用，与对照相比差异性极显著；不同浓度的粗毒素提取液其作用效果不同，随着毒素浓度的增大或是作用时间性的延长，水稻诱导或分化受到的抑制作用越强。水稻成熟胚组织培养中进行抗稻瘟病菌突变体筛选的毒素浓度在30％较好；浸泡水稻成熟胚以处理72h愈伤诱导和分化效果较好；愈伤组织的浸泡以24h可以得到胚性的愈伤和分化出苗。愈伤组织上直接加上毒素并长时间处理对愈伤组织的生长有很大的破坏作用，每个愈伤滴入的毒素应小于0.25ml。在进行抗稻瘟病突变体筛选中，以用含毒素的培养基进行诱导愈伤的处理的筛选取效果较好。 2)稻瘟病菌粗毒素提取液对幼穗的组织培养的影响吉林农业大学硕士学位论文租瘟病菌粗毒素对水稻组织培养的影响及抗性突变体筛选 稻瘟病菌粗毒素对幼穗的愈伤诱导和分化有强烈的破坏作用，并且随着处理时间的增加或毒素浓度的增大受到破坏的程度愈深用稻瘟病菌粗毒素浸泡幼穗对其愈伤诱导和分化都产生很强的抑制作用，能够起到杭性筛选的目的，处理的时间以24h以内可以得到胚性愈伤和分化成苗;筛选时毒素的浓度以30%为宜;用粗毒素直接浸泡幼穗愈伤组织对愈伤的伤害很大，得到的分化植株相对较少，但仍可作为一种杭性愈伤筛选的方法，其处理时间以24h较好。3)稻瘟病菌粗毒素提取液对水稻幼胚的组织培养的影响 稻瘟病菌粗毒素提取液对水稻幼胚的愈伤的诱导和分化的有明显的抑制效应。随着毒素浓度的增加，水稻胚性愈伤获得率和分化能力显著降低。在诱导培养基中加入毒素对幼胚的筛选取效果较好，可以获得较多的胚性愈伤和较多的分化苗。培养基中毒素的浓度处理以30ry0效果好。水稻幼胚的杭性筛选过程中胚性愈伤获得和分化苗数均比幼穗或成熟胚高，是很好的培养材料。更多还原

【Abstract】 Using the crude sphytotoxin extracted from the Phyricularis oryzae.cav filtration, screening of rice blast-resistant mutant by culture, and integrating with the normal breeding technique, the new resistant virus breeding methods was used, and new rice varieties had been breeded. The experiments were carried out from 2002 to 2004, young heads, young and mature embryos were treated by crude phytotoxin mixture extracted from the recently superior fungi of Phyricularis oryzae.cav ,and the effects of Phyricularis oryzae.cav toxin on the rice callus induction and differentiation. The studies were aimed to give the theory dependence to screening of rice-resistant mutant and resistant to Phyricularis oryzae.cav breeding. The results are showed as follows:The growth of different stain on the different media, the growth amount and color of different Phyricularis oryzae.cav fungi was different too(white, gray , brown, black and were he main color ,some turned yellow .yellow and green )When the consistence of Phyricularis oryzae.cav filtration were between 0 to 50%, the toxicity were enhanced with the toxin concentration increasing, the toxin concentration were about 80%, the peak in the curve came, then toxicity decreased with the toxin concentration enhancing. The cure of toxicity were parabola.Toxicity was different with the culture methods and toxin amount in the mediaToxicity was significant different when the toxin was cultured by different biological stain.The growth of sprout ,embryo induction, callus subculture and differentiation were strong. Inhibited by the crude toxin filtration. Contrasted with CK, the results were significant different, and the result was different with toxin concentration changing. With the increasing of the toxin concentration and the prolonging the impact time, the inhibiting was stronger. 30% was the better toxin concentration when screening of rice blast-resistant mutant from the mature embryo culture; 72h was to better time to mutant induction land differentiation when the mature embryo was immersed; but in order to get embryo mutant and differentiation seeding, immersing the mutants 24h was needing. When adding the toxin to the mutant and prolonging the treatment time, the growth of mutant could be destroyed strongly, so the amount of mutant must less than 0.25ml. Contrasted the methods of screening of riceblast-resistant mutant, using the media including toxin to inducting mutant was better.On the young head, the result of experiments was similar to that of the mature embryo. But to young head , mutant was hurt strongly, when it was immersed directly in the toxin, and the differentiation seeding was got little., but this methods still can be used to screening of resistant mutant, 24h was the apposite time.The induction and differentiation of young embryo was evidently inhibited. Mutant forming and differentiating were decreased with the toxicity concentration increasing. Adding the toxin to the media was the good methods to screening of the resistant mutant, by which can got the more differentiation seedings and mutants(30% was the concentration). To screening of rice blast-resistant mutant, the young embryo were the best culture materials than young seeding and mature embryos, which can got more mutant and differentiation seedings.更多还原