Breeding and Characterization of a New Rice Restorer Line Containing Bt Gene

Bt5198, a new rice restorer line containing Bt gene, was developed from the cross and backcross of the elite restorer line Chenghui 177 with Bt Minghui 63, a transgenic Bt restorer line. The inbred lines were evaluated using PCR amplification, test paper evaluation, insect resistance evaluation in both the laboratory and paddy fields, nursery evaluation of rice blast resistance and pedigree selection of agronomic traits. Larval mortalities on Bt5198 and Bt Minghui 63 were 100% when rice culms were inoculated with the eggs of the striped stem borer (SSB) in the laboratory. Bt5198 was highly resistant against SSB and the yellow stem borer (YSB) under field conditions. The F₁ hybrids derived from Bt5198 and four cytoplasmic male sterile (CMS) lines were also highly resistant to SSB and YSB and had a significant heterosis. Two-year evaluation of rice blast resistance confirmed that the resistance levels of Bt5198 to leaf blast and neck blast were similar to those of Chenghui 177 and significantly better than those of Bt Minghui 63. Seed germination ability and pollen yield of Bt5198 were similar with Chenghui 177, suggesting that the introduction of the Bt gene into the new restorer line had no significant effects on seed vitality or the yield of seed production. To identify the presence of the Bt gene, it was effective to combine test paper examination with the evaluation of insect-resistance, both in the laboratory and under field conditions.     

转基因抗虫性水稻恢复系选育及特性分析

将优质抗稻瘟病水稻恢复系成恢177与转基因水稻Bt明恢63杂交并回交1次,采用PCR分析、试纸条检测、室内和田间人工接虫鉴定抗虫性,并结合系谱法选择农艺性状,病圃接种鉴定稻瘟病抗性,育成具转基因抗虫性的水稻新恢复系Bt5198。采用离体茎秆法接种二化螟卵块,新恢复系Bt5198和Bt明恢63的幼虫死亡率均为100%。在田间人工接虫条件下,该恢复系对二化螟和三化螟均表现高抗,与4个不育系配制的杂种F1仍保持良好的抗虫性,且杂种优势明显。两年稻瘟病抗性鉴定结果表明,新恢复系Bt5198的叶瘟和颈瘟抗性水平与成恢177相当,明显优于Bt明恢63。Bt5198的种子发芽率和花粉量与成恢177相当,Bt基因导入水稻恢复系不会对种子生活力和制种产量造成显著影响。在无选择标记基因的转基因后代中,结合利用试纸条检测和室内、田间人工接虫鉴定是筛选Bt基因抗虫性的有效方法。     

优质恢复系成恢047的选育及利用

成恢047是四川省农科院作物所用IR2588－5－1－2作母本，明恢63作父本杂交选育而成的灿型优质恢复系，该恢复系株型好，抗温性强，配合力好，米质优，已配制出一批优质，高产，抗稻瘟性好的组合，与不良系K17A配组育成的K优047于2000年通过四川省品种审定委员会审定。     

电子记录白背飞虱的取食和产卵行为

摘要： 采用电子记录的方法,并结合蜜露量和产卵量测定,研究了白背飞虱在春江06,Rathu Heenati(RHT)和明恢63上的取食和产卵行为。结果表明：白背飞虱在RHT和春江06上的取食刺探频率显著高于明恢63,且取食韧皮部的时间明显少于明恢63;RHT和春江06对白背飞虱具有很强的拒取食作用,明恢63则较感虫;白背飞虱产卵的次数、每次持续时间、总时间及产卵量都不存在品种间的显著差异,具有杀卵作用的只有春江06。春江06具有杀卵和抑制取食两种抗虫性机制,RHT只具有抑制取食的机制,明恢63是感虫的杂交稻恢复系。     

江苏省稻瘟病菌群体分布及优势小种的毒力研究

用中国7个统一鉴别品种对2000~2002年江苏省5大稻区采集的342个稻瘟病菌单孢分离菌株进行了小种鉴定。结果表明3年累计出现7群30个小种,其中ZG1为优势小种,3年的频率分别为65.00%、56.90%和60.38%,ZB群和ZC群生理小种也是重要的小种类型,有较高的出现频率。用130个ZG1小种的菌株对日本13个已知抗性基因品种进行毒力测定,出现42种类型毒力,其中30.77％的ZG1小种的菌株同时对品种新2号（Pita-k[sup]s[/sup],Pi-sh）、K1（Pi-ta）、Pi-4号（Pi-ta[sup]2[/sup],Pi-sh）致病,为优势毒力类型, 表明品种新2号（Pita-ks,Pi-sh）、K1（Pi-ta）、Pi-4号（Pi-ta[sup]2[/sup],Pi-sh)与江苏省稻瘟病菌有较高的适合度。品种K3（Pi-k[sup]h[/sup]）在江苏省有较好的抗性,其抗性频率为100%。用两种毒力类型的不同菌株2003-184(ZC[sub]5[/sub])和 2003-14-1(ZG[sub]1[/sub]) 混合接种13个日本已知抗性基因品种后,发现对上述两个单个菌株表现抗病的K3（Pi-k[sup]h[/sup]）品种表现为感病,提示不同毒力的菌株之间的相互作用导致菌株毒力的改变可能是品种抗性丧失的原因之一。     

稻褐飞虱实验种群致害性变异

以蜜露排泄量为稻褐飞虱致害性指标对稻褐飞虱实验种群的致害性变异进行了研究。首先测定了褐飞虱3个实验种群(TN1种群、Mudgo种群和ASD7种群)[分别用水稻品种TN1、Mudgo和ASD7强迫饲养82代,其中TN1种群(T)为弱致害力种群,Mudgo种群(M)和ASD7种群(A)为强致害力种群]分别在感虫和抗虫品种上的蜜露排泄量。结果表明,寄主表现感虫还是抗虫,与褐飞虱致害力之间存在交互作用。将不同致害力的褐飞虱种群接在抗虫品种上,才可能观察到较大的致害性差异,据此比较了杂交F1代与亲本种群个体、自交F2代与杂交F1代种群个体在抗虫品种上的致害性差异,以分析褐飞虱致害性变异规律。杂交F1代（包括T♀×M♂,A♀×T♂两种处理）雌成虫的平均致害力均介于两个亲本之间,其值与强致害力亲本的更为接近;杂交F1代雌成虫致害力的变异程度均显著地大于弱致害亲本,而与强致害性的亲本相比几乎无明显的差异;表明相对于弱致害性,褐飞虱的强致害性可能为一显性性状。与杂交F1代相比,T♀×M♂处理中自交F2代雌成虫的致害力无显著变化,而A♀×T♂处理中自交F2代雌成虫的致害力显著大于杂交F1代;两种处理中,自交F2代的方差或显著小于杂交F1代,或稍大于杂交F1代,表明自交F2代种群个体未增加新的遗传分离。     

水稻新广亲和恢复系成恢448的选育及利用

 将美国水稻Lemont与中国籼型恢复系871028杂交,采用常规育种方法和花药培养技术,育成广亲和新恢复系成恢448。该恢复与籼稻和粳稻测验种杂交,F1代结实率分别为85.89%和69.04%;与5种不同细胞质来源的不育系测交,F1代结实率平均为83.39%;已测交筛选出一批强优势组合,其中,汕优448和Ⅱ优448于1997年通过四川省区试,2年平均产量分别比对照增产9.07％和14.51%。讨论了利用美国稻种资源开展超高产杂交水稻育种应注意的技术问题。     

水稻成熟胚组织培养研究

对珍汕 97B、明恢 6 3、H14 93、中花 11等 4个水稻品种的成熟胚在组织培养中愈伤组织诱导率和植株再生率的研究表明 ,在N6培养基中附加体积分数为 2× 10 -3 的 2 ,4 -D时诱导率最高。在相同的诱导培养基和分化培养基上 ,4个水稻品种的愈伤组织诱导率和植株再生率表现出极大的差异。粳稻H14 93表现最好 ,其诱导率和再生率分别达到 90 .6 %和 90 .0 % ;中花 11次之 ,分别为 85 .2 %和 6 6 .7% ;2个籼稻品种的愈伤组织诱导率较低 ,植株再生率几乎为 0。说明本试验中的诱导条件适合于粳稻品种 ,而对籼稻品种不利     

稻褐飞虱致害性变异的研究

 研究了杭州褐飞虱田间种群在抗虫品种ASD7上强迫饲养多代后致害性的变化。结果表明,不同致害性的群体,在ASD7上饲养4代后,均可转变成能为害ASD7的ASD7种群。反之,在无抗性胁迫的感性品种TN1上连续饲养7代,ASD7种群对ASD7的致害性逐渐减弱,最终退化为TN1种群。这表明褐飞虱的致害性是可变异的,水稻品种抗、感虫特性是其转化的诱因。     

中国南方水稻条斑病菌小种分化研究

采用针刺接种的方法,在水稻孕穗期的剑叶上接种测定了国内南方稻区细菌性条斑病菌62个菌株的致病力。根据病菌与鉴别品种互作反应和亲和性的差异,将供试条斑病菌株区分为6个小种。大多数条斑病菌株与鉴别品种的互作反应表现为弱互作模式,部分菌株与品种间存在强互作关系.     

中国水稻干尖线虫部分群体对水稻的致病力测定

 为了解不同地理群体和寄主群体的中国水稻干尖线虫对水稻的致病力差异,通过室内水稻盆栽接种试验,对来自中国6个省、2种不同寄主植物上的水稻干尖线虫8个群体的致病力进行了测定和分析。结果表明,供试的8个水稻干尖线虫群体均能侵染供试的3个水稻品种,但不同群体对水稻同一品种的致病力,以及同一群体对不同水稻品种的致病力均存在差异;不同水稻品种被侵染后的症状也存在差异,仅有辽盐16被所有群体侵染后均表现“干尖”症状,武育粳3号和博优998没有明显“干尖”症状。供试的3个水稻品种接种水稻干尖线虫后均能抽穗,但是株高、穗长、单株穗数和千粒重受到不同程度的影响。此外,水稻干尖线虫对水稻的致病力与群体寄主相关,来自草莓的水稻干尖线虫群体对水稻的致病力明显弱于来自水稻上的群体;在供试的水稻干尖线虫群体中,HN 2群体对水稻品种辽盐16的生长影响最大,但在稻株上的虫量最少,显示该群体繁殖数量不大,但具有较强的致病力。这说明水稻干尖线虫在水稻上的繁殖数与致病力之间并不一定呈正相关。这些结果表明中国的水稻干尖线虫可能存在不同的生理小种或致病型。     

不同致害性褐飞虱种群刺吸电位图（EPG）的比较

 利用刺吸电位图(EPG)比较分析了两个致害性不同的褐飞虱种群（TN1种群、Mudgo种群）在感虫品种TN1和抗性品种Mudgo（含Bph1抗虫基因）上的取食行为。结果表明,种群类型、水稻品种对I波的影响最明显,A波次之,S波再次之;试虫性别对各参数均无显著影响。TN1种群在TN1上及Mudgo种群在TN1、Mudgo上这3种能致害处理下的S波、A波均短于TN1试虫在Mudgo上（不能致害）,其中A波的差异达显著水平;而能致害的3种处理的I波持续时间则显著长于不能致害处理。若以3 h内I波持续时间75 min为分界线,高于该值为能致害,低于该值为不能致害,对TN1种群、Mudgo种群致害性的判别准确率分别为82.6%和100%。提出该参数可用于进一步开发褐飞虱个体致害性检测新技术。     

抗草铵膦转基因水稻明恢86B杂草化潜力评价

 为评估抗草铵膦转基因水稻明恢86B大规模推广后演化为杂草的生态风险,在农田生态环境下比较了明恢86B、明恢86和杂交稻组合汕优63的生存竞争能力、繁育能力、落粒性、种子生存能力以及对常规除草剂氟吡磺隆的耐药性。结果表明,无论在适宜季节还是非适宜季节,明恢86B和明恢86的生存竞争能力和繁殖力都低于杂交稻组合汕优63,尤其在非适宜期时,汕优63的生存竞争力、繁殖力明显强于明恢86B和明恢86。主要表现在植株较高、分蘖较多以及高产。明恢86B的生存竞争力和繁殖力都略低于明恢86。3个材料的落粒性都不强,且尚未见自生苗产生。在浅埋和深埋处理下,明恢86B种子的生存能力弱于汕优63和明恢86。3个材料对除草剂氟吡磺隆的耐药性无明显差异。表明抗草铵膦转基因水稻86B在中国南京地区环境条件下演化为杂草的可能性较小。     

Evaluation of Rice Germplasm for Resistance to the Small Brown Planthopper （Laodelphax striatellus） and Analysis of Resistance Mechanism

One hundred and thirty-eight rice accessions were screened for resistance to the small brown planthopper (SBPH) (Laodelphax striatellus Fallén) by the modified seedbox screening test. Twenty-five rice accessions with different levels of resistance to SBPH were detected, accounting for 18.1% of the total accessions, which included 2 highly resistant, 9 resistant and 14 moderately resistant varieties. Compared with indica rice, japonica rice was more susceptible to SBPH. Antixenosis test, antibiosis test and correlation analysis were performed to elucidate the resistance mechanism. The resistant check Rathu Heenati (RHT), highly resistant varieties Mudgo and Kasalath, and resistant variety IR36 expressed strong antixenosis and antibiosis against SBPH, indicating the close relationship between resistance level and these two resistance mechanisms in the four rice varieties. Antibiosis was the dominant resistance pattern in the resistant varieties Daorenqiao and Yangmaogu due to their high antibiosis but low antixenosis. Dular, ASD7 and Milyang 23 had relatively strong antixenosis and antibiosis, indicating the two resistance mechanisms were significant in these three varieties. The resistant DV85 expressed relatively high level of antixenosis but low antibiosis, whereas Zhaiyeqing 8 and Guiyigu conferred only moderate antibiosis and antixenosis to SBPH, suggesting tolerance in these three varieties. Antibiosis and antixenosis governed the resistance to SBPH in the moderately resistant accession 9311. Antixenosis was the main resistance type in V20A. Tolerance was considered to be an important resistance mechanism in Minghui 63 and Yangjing 9538 due to their poor antibiosis and antixenosis resistance. The above accessions with strong antibiosis or antixenosis were the ideal materials for the resistance breeding.     

白叶枯病菌效应子XopN在拥有OsSWEET11同源基因的水稻品种上发挥毒性作用

【目的】黄单胞菌细胞外泌蛋白质(Xanthomonas outer proteins, Xops)是植物病原黄单胞菌高度保守的毒性效应子或毒性辅助组分,通过细菌Ⅲ型分泌系统分泌到细菌细胞外部,然后转入植物细胞而发挥病理作用。水稻黄单胞菌水稻致病变种即水稻白叶枯病菌的标准菌株PXO99^A分泌的XopN是一种毒性效应子,通过影响寄主免疫反应而使水稻发病。但是,XopN对病菌毒性的影响是否因水稻品种的不同而异,还有待研究。【方法】利用同源双交换技术,先敲除了PXO99^A的XopN基因,获得了∆XopN突变体,又经过遗传互补,得到了回补菌株∆XopN/XopN。通过营养肉汤液体培养,测定了XopN对病菌繁殖能力的影响;根据文献选用14个水稻品种,通过接种实验,测定了PXO99^A对这些品种的毒性与XopN敲除或回补的影响;在XopN发挥毒性作用的水稻品种上,测定了隐性抗病基因OsSWEET11/xa13与显性感病基因OsSWEET11/Xa13受病菌侵染而表达的情况,分析了XopN敲除或回补的影响。【结果】在营养肉汤液体培养过程中,病菌突变体菌株∆XopN的繁殖速度明显低于野生型PXO99^A。PXO99^A、∆XopN和∆XopN/XopN接种水稻后,根据其在水稻叶片组织内的繁殖量及随后产生的白叶枯病症状的严重程度,将供试的14个水稻品种分为两种情况。一是感病程度与病菌XopN是否敲除或回补无关,这有10个水稻品种（IRBB1、IRBB3、IRBB8、IRBB10、IRBB14、IR24、IRBB203、IRBB204、IRBB205和IRBB211）,PXO99^A、∆XopN和∆XopN/XopN对它们的毒性无明显差别。二是XopN对病菌毒性发挥作用的水稻品种,包括高度感病的3个品种（IRBB208、Asominori和日本晴）和低感品种IRBB13。与PXO99^A或∆XopN/XopN相比,∆XopN对这4个水稻品种的毒性大为降低。在IRBB13上,病菌侵染对隐性抗病基因OsSWEET11/xa13在叶片内的表达发生抑制作用,这一效应与XopN的毒性功能相关。相反,日本晴显性感病基因OsSWEET11/Xa13却受病菌侵染的诱导,在叶片内的表达水平大幅度提高。IRBB208和Asominori携带OsSWEET11/Xa13同源基因,该同源基因在叶片内的表达水平也因病菌侵染而大幅提高。在这4个水稻品种上,PXO99^A和∆XopN/XopN能够诱导OsSWEET11/Xa13或其同源基因表达,但∆XopN无此作用。另外,XopN对病菌在非寄主植物烟草上诱发过敏反应有量变贡献,相比PXO99^A或∆XopN/XopN,∆XopN引起过敏反应的程度有所降低。【结论】XopN是一个有限广谱性效应子,在拥有OsSWEET11同源基因的水稻品种上发挥毒性作用。XopN也是病菌繁殖所需要的,对病菌在非寄主植物上诱导过敏反应有一定贡献。     

Genetic Improvement of Rice for Bacterial Blight Resistance: Present Status and Future Prospects

The production and productivity of rice has been challenged due to biotic and abiotic factors. Bacterial blight (BB) disease, caused by Xanthomonas oryzae pv. oryzae, is one of the important biotic stress factors, which reduces rice production by 20%–50%. The deployment of host plant resistance is the most preferred strategy for management of BB disease, and breeding disease resistant varieties remains a very economical and effective option. However, it is difficult to develop rice varieties with durable broad-spectrum resistance against BB using conventional approaches alone. Modern biotechnological tools, particularly the deployment of molecular markers, have facilitated the cloning, characterization and introgression of BB resistance genes into elite varieties. At least 46 BB resistance genes have been identified and mapped from diverse sources till date. Among these, 11 genes have been cloned and characterized. Marker-assisted breeding remains the most efficient approach to improve BB resistance by introducing two or more resistance genes into target varieties. Among the identified genes, xa5, xa13 and Xa21 are being widely used in marker-assisted breeding and more than 70 rice varieties or hybrid rice parental lines have been improved for their BB resistance alone or in combination with genes/QTLs conferring tolerance to other stress. We review the developments related to identification and utilization of various resistance genes to develop BB resistant rice varieties through marker-assisted breeding.     

Detection and characterization of bismerthiazol-resistance of Xanthomonas oryzae pv. oryzae

The resistance to bismerthiazol in the Xanthomonas oryzae pv. oryzae population in China has not been clearly determined. Three bismerthiazol-resistant mutants (1-1-1, 2-1-1, 4-1-1) of X. oryzae pv. oryzae were induced on rice plants and the characteristics were compared with their parental strain ZJ173. After 10 subcultures on fresh bismerthiazol-free nutrient agar plates and after being inoculation onto and re-isolation from untreated rice plants two times, the mutants remained pathogenicity on rice plants treated with 300 μg/ml bismerthiazol, indicating that the in vivo-induced bismerthiazol resistance was stable in vivo. The in vitro characteristics of the bismerthiazol-resistant mutants and a wild-type bismerthiazol-sensitive strain (ZJ173) were also compared. The three mutants grew faster than ZJ173 during the logarithmic growth phase but at the same rate during the decline phase. Bismerthiazol had protective and curative activity against strain ZJ173 but not against the three mutants. To evaluate the current degree of bismerthiazol resistance in field populations of X. oryzae pv. oryzae in China, 781 isolates of X. oryzae pv. oryzae were obtained from 1419 leaf samples collected from Yunnan, Sichuan, Jiangsu, Anhui, Hubei, Guangdong, Hainan, and Hunan Provinces in China from 2007 to 2009. The in vivo sensitivity of 505 of the 781 isolates to bismerthiazol was determined by applying them to rice plants treated with 300 μg/ml bismerthiazol and evaluating blast severity. The tested strains were considered resistant, if they showed <70% inhibition relative to strain ZJ173. Of the 505 isolates, 62 were resistant to bismerthiazol. The resistance frequencies among field population ranged from 0 to 21.1%, differed among the provinces, and was 11.2, 20.0 and 10.4% in 2007, 2008 and 2009, respectively. Resistance determined in vivo was not related to resistance determined in vitro. The in vivo-induced bismerthiazol resistance identified in this study will serve as a base line for further studying bismerthiazol resistance in fields, and the current degree of bismerthiazol resistance in field populations of X. oryzae pv. oryzae in China determined would be useful for the rice growers for control of rice bacterial blight.