广谱稻瘟病抗性基因<italic>Pigm</italic> 和<italic>Pi2</italic> 的抗谱差异及与<italic>Pi1</italic> 的互作效应

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于苗苗, 戴正元, 潘存红, 陈夕军, 余玲, 张晓祥, 李育红, 肖宁, 龚红兵, 盛生兰, 潘学彪, 张洪熙, 李爱宏. 广谱稻瘟病抗性基因Pigm 和Pi2 的抗谱差异及与Pi1 的互作效应 . 作物学报, 2013, 39(11): 1927-1934[YU Miao-Miao, DAI Zheng-Yuan, PAN Cun-Hong, CHEN Xi-Jun, YU Ling, ZHANG Xiao-Xiang, LI Yu-Hong, XIAO Ning, GONG Hong-Bing, SHENG Sheng-Lan, PAN Xue-Biao, ZHANG Hong-Xi, LI Ai-Hong. Resistance Spectrum Difference between Two Broad-Spectrum Blast Resistance Genes,Pigm andPi2 , and Their Interaction Effect onPi1 . 作物学报, 2013, 39(11): 1927-1934] 复制到剪切板

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广谱稻瘟病抗性基因Pigm 和Pi2 的抗谱差异及与Pi1 的互作效应

于苗苗^1,^2,³, 戴正元¹, 潘存红¹, 陈夕军², 余玲¹, 张晓祥¹, 李育红¹, 肖宁¹, 龚红兵³, 盛生兰³, 潘学彪², 张洪熙¹, 李爱宏^1,^*

¹江里下河地区农业科学研究所 / 国家水稻改良中心南京分中心扬州试验站, 江苏扬州225007

²扬州大学植物功能基因组学教育部重点实验室, 江苏扬州225009

³江苏丘陵地区镇江农业科学研究所, 江苏句容212400

^* 通讯作者(Corresponding author): 李爱宏, E-mail:yzlah@126.com, Tel: 0514-87302340

收稿日期:2013-06-24

基金:本研究由江苏省自然科学基金项目(BK2011427)和江苏省农业科技自主创新资金产业体系类项目(CX(12)1003-4)资助。

摘要

Pigm 是Pi2 基因簇的等位或紧密连锁基因。本研究构建了4个不同遗传背景下Pigm 和Pi2 的系列近等基因系, 204个菌株苗期接种结果显示其抗性频率均超过70%, 但Pigm 和Pi2 的抗谱重叠度仅为54.4%~65.7%, 聚合Pi1/Pigm 和Pi1/Pi2 杂种的抗性频率均超过90%。穗瘟人工接种及病圃自然诱发鉴定表现与苗期接种一致的发病趋势。农艺性状调查结果显示获得的近等基因系与其轮回亲本基本相似, 存在较少的累赘连锁。表明Pigm 是一个与Pi2 抗谱差异明显的广谱抗性基因, 对稻瘟病抗性育种具有重要应用价值。

关键词: 稻瘟病抗性基因; 标记辅助选择; 近等基因系; 抗谱; 互作效应

Resistance Spectrum Difference between Two Broad-Spectrum Blast Resistance Genes,Pigm andPi2 , and Their Interaction Effect onPi1

YU Miao-Miao^1,^2,³, DAI Zheng-Yuan¹, PAN Cun-Hong¹, CHEN Xi-Jun², YU Ling¹, ZHANG Xiao-Xiang¹, LI Yu-Hong¹, XIAO Ning¹, GONG Hong-Bing³, SHENG Sheng-Lan³, PAN Xue-Biao², ZHANG Hong-Xi¹, LI Ai-Hong^1,^*

¹Lixiahe Agricultural Research Institute of Jiangsu Province / Yangzhou Station of Nanjing Sub-center of National Rice Improvement Center, Yangzhou 225007, China

²Key Laboratory of Plant Functional Genomics, Ministry of Education, Yangzhou University, Yangzhou 225009, China

³Zhenjiang Institute of Agricultural Science in Jiangsu Hilly Region, Jurong 212400, China

Abstract

Pigm , an R gene to rice blast disease, is either allelic or tightly linked toPi2. In this study, a series of near-isogenic lines (NILs) containingPigm andPi2 under four genetic backgrounds of rice cultivars were established, respectively. The results from inoculating with 204 representative races at seedling stage showed that the resistance frequency of NILs carryingPigm andPi2 all reached more than 70%, but the overlapping degree of resistance spectrum between them was only 54.4% to 65.7%, and the resistance frequency of hybrids pyramidingPi1/Pigm andPi1 /Pi2 was more than 90%. Disease development from artificial inoculation at heading stage and natural induction in disease nursery showed consistent trend with that from inoculation at seedling stage. Agronomic traits of most NILs were similar to those of recurrent parent, indicating that the target region contains few linkage drags. In conclusion,Pigm is a broad-spectrum blast resistance gene possessing significant resistance spectrum difference fromPi2, and holds important value for rice blast breeding.

Keyword: Blast resistance gene; Marker-assisted selection; Near-isogenic lines; Resistance spectrum; Interaction effect

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引言

发掘新的广谱稻瘟病抗性基因是培育抗性品种及控制病害的有效途径。迄今已有63个抗稻瘟病位点的77个主效基因被分子定位, 并有22个基因被克隆。这些基因分布于除第3染色体外的其他11条水稻染色体上, 并具有成簇分布的特性(http://www. ricedate.cn/gene/, 国家水稻数据中心基因数据库)。其中, 第6染色体短臂近着丝粒的附近分布有 Pi2 、 Pi9 、 Pi22 、 Pi25 、 Pi26 、 Pi40 、 Pi42 、 Pi50、Piz 、 Piz-t 等至少10个抗性基因, 这些基因在抗病基因簇内呈等位或紧密连锁关系, 并有研究结果显示其抗谱存在明显差异^{[1,2,3,4,5,6,7]}。但这种差异大多是在不同遗传背景下获得的初步鉴定结果, 缺乏在同一遗传背景下(如近等基因系水平上)的验证。 Pigm 是从持久抗稻瘟病品种谷梅4号中鉴定的一个新的广谱稻瘟病抗性基因, 定位及等位性测验结果显示其可能与第6染色体的 Pi2 和 Pi9 等位或连锁^[8]。 Pi1 是源自利比亚粳型品种“LAC23”的广谱稻瘟病抗性基因, 定位在第11染色体上MGR4766标记1.3 cM处, 现已经被成功克隆^[9,10], 并有研究显示其与 Pi2 有良好的互补效应^[11,12,13]。本研究构建了4个轮回亲本背景下 Pigm 、 Pi2 的系列近等基因系, 进而比较其抗谱差异, 并分析其与 Pi1 的互作效应, 从而为稻瘟病抗性育种提供参考。

1 材料与方法

1.1 供试材料的选择

Pi1 和 Pi2 基因的供体亲本为W1, 是以金23B为轮回亲本导入 Pi1 和 Pi2 基因的中间材料^[9]。 Pigm 的供体亲本为谷梅4号。受体轮回亲本包括2个光温敏核不育系广占63S、97S, 及2个中籼稻恢复系R084和R609, 均由本项目组保存或培育而成。

1.2 多态性分子标记的发展

结合http://www.gramene.org/网站上分子标记的信息和3个抗性基因的定位或克隆信息, 采用Primer Premier 5.0软件发展分子标记设计引物, 引物由TaKaRa公司负责合成, 用1%~3%的琼脂糖电泳筛选多态性的分子标记。最后针对不同供体, 筛选获得与4个轮回亲本均有良好多态性的标记AP22、ZJ58.7及RM224, 用于后续的标记辅助选择。具体信息列于表1。

表1 筛选的分子标记信息 Table 1 Marker information for MAS

1.3 标记检测

参照卢扬江等^[14]的方法提取水稻全基因组DNA。20 μL PCR反应体系, 包括MgCl₂ (25 mmol L^-1) 2.0 μL, buffer (×10) 2.0 μL, 引物(上下游引物, 3.084 mmol L^-1) 1.5 μL, dNTP (10 mmol L^-1) 0.4 μL, Taq DNA聚合酶(5 U μL^-1) 0.2 μL, 模板DNA 2.0 μL; ddH₂O 11.9 μL。PCR条件为94℃预变性5 min; 94℃变性50 s, 55℃复性50 s, 72℃延伸1 min, 36个循环; 72℃充分延伸10 min。将PCR产物上样于含溴化乙锭的3%琼脂糖凝胶, 电泳分离后于BIORAD凝胶成像仪下拍摄并观察读数。供体亲本带型、受体亲本带型和双带分别赋值1、3和2, 缺带赋值0。

1.4 遗传背景回复率检测

结合http://www.gramene.org/网站上公布的水稻SSR分子标记的信息和本实验室开发的标记, 采用目标基因区间以外相对均匀分布的132个分子标记, 针对供、受体双亲进行多态性筛选, 在2个亲本之间具有多态性的标记确定为用于背景回复率检测的中选标记, 要求每条染色体至少获得3个以上的多态性标记且分布相对均匀。参照Hospital等^[15]的方法计算株系背景回复率, G_g = ( L + X_g )/2 L , 其中, G_g 为在 g 代的遗传背景回复率; X_g 为在回交 g 代表现为轮回亲本带型的分子标记数量; L 为检测的分子标记总数量。

1.5 病原菌的分离与培养

从广东、江苏、海南、浙江和湖北5省重发病区田间采集自然发生的稻瘟病病穗, 参考周益军等的方法^[16]培养病菌, 单孢分离后在PDA培养基上培养5~7 d, 转于产孢培养基上培养5~7 d, 用无菌水洗去培养基表面菌丝后于黑光灯下照射培养72 h (培养温度均为26℃)。筛选获得204个产孢量大的单孢菌株, 洗脱稻瘟菌孢子, 调节浓度至2×10⁵个 mL^-1, 作为鉴定所用的接种体。

1.6 抗性鉴定与评价

1.6.1 苗瘟鉴定与评价将各株系种子浸种催芽后播于塑料秧盘内, 2次重复, 利用上述培养的204个菌株孢子悬浮液, 待秧苗长至三至四叶期时移至玻璃箱内喷雾接种, 于26~28℃下黑暗保湿24 h (相对湿度>95%), 然后将秧盘移至可控温室内, 26~28℃、相对湿度>95%的高湿环境下生长。7~10 d后调查病情, 依据国家农业行业标准(水稻稻瘟病鉴定技术规范)记载病情, 以发病最重的稻株作为该品种的抗性级别, 每个重复中只要有2株以上感病即记为感(S), 低于2株记为抗(R)。抗性频率=(接种后表现抗的菌株数/接种的总菌株数) × 100%; 抗谱重叠度=(具有相同抗性表型的菌株数/接种的总菌株数) × 100%。

1.6.2 穗瘟鉴定与评价试验于江苏里下河地区农业科学研究所万福基地进行, 5月10日播种各试验材料, 苗床育苗30 d后于6月10日移栽, 单本栽插, 每区5行, 每行12株, 株行距13.3 cm × 25.0 cm, 2次重复, 常规水肥管理。孕穗期选用来自广东、湖北、海南、浙江和江苏的5个代表性的强毒力菌株GD18-3 (B1, 致病率65.6%)、ES9-5-1 (B15, 致病率75.0%)、hn1-1 (C1, 致病率59.4%)、ZJ13-4 (A15, 致病率63.3%)和XH8-4-4 (G1, 致病率37.5%), 扩大培养后等比例混合5个菌株的孢子液。选择每个株系的第2行, 用注射器将接种液注入稻苞内, 每穗注射1 mL, 每重复接种10穗, 并对接种的稻穗作好标记。穗瘟鉴定评价标准参见罗楚平等^[17]的方法。

1.6.3 病圃自然诱发鉴定自然诱发鉴定病圃建于武陵山区湖北省恩施的两河地区, 常年3月至9月份相对湿度超过90%, 非常适宜稻瘟病的发生。3月25日播种试验材料, 5月上旬移栽, 每个株系栽插100穴, 2次重复。齐穗后30 d调查每个株系的穗发病情况, 按照国家稻瘟病测报调查规范(GB/T 15790-2009)划分抗感类型和病级。

1.7 基本农艺性状的考察

对用于田间穗颈瘟人工接种鉴定的试验材料, 同时进行基本农艺性状考察。调查记录各试验小区的播始历期和株高。成熟时, 每重复取中间第3、第4两行各8株共16株, 考察有效穗数、每穗粒数、结实率、千粒重、单株产量等性状。

2 结果与分析

2.1 携带 Pigm 、 Pi2 的近等基因系构建

以广占63S、97S、R084和R609为母本, 与供体亲本W1、谷梅4号分别杂交获得F₁, 然后以广占63S、97S、R084和R609分别为轮回亲本连续回交, 从BC₁F₁世代开始分别利用目标基因 Pi2 、 Pigm 紧密连锁标记AP22、ZJ58.7检测基因型, 选择标记为杂合带型且农艺性状与轮回亲本接近的单株回交, 经过连续4次回交获得BC₄F₁代后(江苏扬州、海南一年两季加代)自交, 利用单个鉴别菌株对BC₄F₂群体接种鉴定, 发现AP22、ZJ58.7的选择效率高达93%以上, 与前人研究结果一致^[12]。BC₄F₂分离世代选择标记为纯合抗性基因型且农艺性状优良的单株(图1-A), 针对每个轮回亲本的单个目标基因, 分别获得优良单株10个以上。

将BC₄F₃世代种植成株系, 进行以下三方面选择: (1)遗传背景回复率, 针对广占63S、97S、R084和R609等4个轮回亲本与供体亲本, 利用132个SSR标记进行多态性分析, 筛选发现W1与4个亲本间的多态性标记个数依次为47、46、49、50, 谷梅4号与4个亲本间的多态性标记个数依次为44、47、52、53, 且均匀分布于水稻12条染色体上(每条染色体上均有3个以上的多态性标记), 利用这些多态性标记对每个株系进行遗传背景回复率检测, 选择背景回复率>95%的株系; (2)目标基因选择的准确性, 为避免分子标记与目标基因之间由于交换重组等导致的假阳性选择, 对每个株系收获的种子, 利用不同地区分离的15个代表性强毒力菌株开展苗瘟接种鉴定, 选择抗性频率较轮回亲本显著提高的株系(图1-B, C, D), 确保目标基因被准确选择; (3)农艺性状, 在群体水平上选择与轮回亲本最为相似的株系。最后, 综合上述三方面选择情况, 针对每个轮回亲本的单个目标基因, 确定一个株系用于后续的抗谱分析。

2.2 不同遗传背景下目标基因的抗谱分析

2.2.1 Pigm 与 Pi2 的抗谱差异不同地区分离的204个稻瘟病菌株的苗期接种结果显示, 携带 Pigm 与 Pi2 基因的近等基因系, 在不同遗传背景的情况下表现基本相似的抗性频率(图2-A), 而与其各自的轮回亲本(对照)相比, 抗性频率显著提高, 均超过70%, 表明 Pigm 与 Pi2 均是抗谱较广的稻瘟病抗性基因。

Pigm 与 Pi2 的抗谱重叠程度, 在不同遗传背景下介于54.4%~65.7%之间(图2-B), 表明两者虽然均有较宽的抗谱, 但抗谱差异较大, 虽然可能为等位或紧密连锁基因, 但在进化上已表现出明显分化。

2.2.2 Pigm 、 Pi2 与 Pi1 的互作效应有研究表明 Pi2 与另一广谱稻瘟病抗性基因 Pi1 具有良好的互补效应^[11,12,13]。对广占63S、97S与W1杂交的后代, 针对 Pi1 基因, 通过上述同样流程, 构建广占63S、97S为轮回亲本的携带 Pi1 的近等基因系, 携带目标基因株系的抗性频率分别为62.79%和60.26%, 表现出相对较宽的抗谱。对 Pigm 、 Pi2 分别与 Pi1 的抗谱比较显示, 广占63S背景下, Pigm 和 Pi1 的抗谱重叠度是59.49%, Pi2 和 Pi1 是58.23%; 97S背景下, Pigm 和 Pi1 的抗谱重叠度是50.63%, Pi2 和 Pi1 是46.84% (图2-C)。表明 Pigm 与 Pi2 相似, 均与 Pi1 基因之间具有良好的互补性。

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图1 目标基因近等基因系的构建A: 目标基因BC₄F₂世代的选择, 绿色箭头示目标个体(M: DL2000 DNA标准分子量; P1: 受体亲本; P2: 供体亲本; L1~17: BC₄F₂世代不同基因型个体); B: 标记选择为阳性的株系进行苗瘟接种验证, 红色箭头示携带目标基因的抗性株系, 白色箭头示感病轮回亲本; C: 携带目标基因的株系叶片呈现过敏性坏死病班(抗性表型); D: 轮回亲本叶片呈现感病表型。Fig. 1 Construction of NILs carrying target genesA: choosing of target genes in BC₄F₂ generation, and green arrows indicate target individuals (M: DL2000 DNA marker; P1: donor parent; P2: recipient parent; L1-17: Individuals with different genotypes in BC₄F₂ generation). B: positive lines were inoculated on seedling stage, and red arrows show the resistant lines carrying target genes, while white arrows indicate susceptible parents. C: line carrying the target gene shows resistant phenotype. D: recurrent parent shows susceptible phenotype.

利用携带 Pi1 的广占63S、97S的近等基因系作母本, 携带 Pigm 、 Pi2 的R084、R609近等基因系作父本, 配制杂交组合, 同时用相应原轮回亲本配制杂交组合作对照, 研究 Pigm 、 Pi2 分别与 Pi1 的聚合效应。苗瘟人工接种结果显示, 与 Pi2 相似, Pigm 与 Pi1 有良好的聚合效应, 聚合后其抗性频率在不同遗传背景下均达到90%以上(图2-D)。

2.2.3 目标基因的穗瘟抗性及病圃自然诱发鉴定

由于穗瘟对水稻的产量影响更为重要, 而关于苗瘟与穗瘟抗性是否相关, 针对不同基因以及不同的研究材料, 一直有不同的研究结论^[18,19,20]。为了解 Pigm 基因的穗瘟抗性效果, 选择不同生态区分离的代表性强毒力菌株, 对不同近等基因系穗瘟人工接种鉴定表明, Pigm 与 Pi2 对穗瘟均有良好的抗性效果, 不同遗传背景下, 均表现中抗(MR)或中感(MS)表型; 而与 Pi1 分别聚合后, 则表现抗(R)或高抗(HR)表型, 而原轮回亲本(组合)均表现高感(HS)表型, 与苗瘟抗性表现明显相关性(表2和图3)。

此外, 为了解上述材料在自然种植、病原菌强胁迫条件下的发病情况, 将其种植于武陵山区湖北恩施两河地区的病圃。该区域雨量丰沛、光照少、田间湿度大, 是我国稻瘟病重发病区, 且国家长江中下游水稻区试稻瘟病病圃鉴定也是设于该地。自然诱发鉴定结果显示(表2), 与穗瘟人工鉴定结果基本一致, 携带 Pigm 、 Pi2 的近等基因系苗瘟和穗瘟表现MR或MS表型, 与 Pi1 分别聚合后, 表现R或HR抗性表型。表明 Pigm 在全生育期均能稳定表达抗性, 具有重要育种价值。

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图2 Pigm 与 Pi2 的抗谱比较及与 Pi1 的互作A: Pigm 与 Pi2 的抗性频率差异; B: Pigm 与 Pi2 的抗谱重叠度; C: Pigm 、 Pi2 分别与 Pi1 的抗谱重叠度; D: Pigm 、 Pi2 分别与 Pi1 聚合后的互作效应。Fig. 2 Comparison of resistance spectrum between Pigm and Pi2, and pyramiding effect of Pigm/Pi1 and Pi2/Pi1A: difference of resistance frequency between Pigm and Pi2 ; B: overlapping degree of resistance spectrum between Pigm and Pi2 ; C: overlapping degree of resistance spectrum between Pigm and Pi1 or Pi2 and Pi1 ; D: pyramiding effect of Pigm/ Pi1 and Pi2 /Pi1 .

表2 不同近等基因系穗瘟人工接种及病圃自然诱发鉴定情况 Table 2 Results of artificial inoculation on heading stage and natural induction in disease nursery for NILs

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图3 部分近等基因系的穗瘟人工接种表现A: R084背景下携带 Pi2 与 Pigm 的近等基因系的穗瘟人工接种表现; B: 聚合有 Pi1 / Pi2 和 Pi1 / Pigm 的近等基因系杂种“GZ63S/R084”穗瘟人工接种表现; C: R609背景下携带 Pi2 与 Pigm 的近等基因系的穗瘟人工接种表现; D: 聚合有 Pi1 / Pi2 和 Pi1 / Pigm 的近等基因系杂种“GZ63S/R609”穗瘟人工接种表现。Fig. 3 Phenotype of partial NILs carrying target genes with inoculation for panicle blastA: phenotype of NILs carrying Pi2 and Pigm inoculated panicle blast under R084 background; B: phenotype of NILs hybrid combination from “GZ63S/R084”pyramiding Pi1/Pi2 or Pi1/Pigm under inoculation for panicle blast; C: phenotype of NILs carrying Pi2 and Pigm inoculated panicle blast under R609 background; D: phenotype of NILs hybrid combination from “GZ63S/R609” pyramiding Pi1/Pi2 or Pi1/Pigm inoculated panicle blast.

2.3 目标近等基因系的农艺性状

对携带 Pigm 、 Pi2 近等基因系农艺性状的考察结果显示(表3), 经过分离世代农艺性状及背景检测的选择, 与4个轮回亲本相比, 各近等基因系基本农艺性状与之相似, 只有少数株系在个别性状上与对照存在差异, 但产量构成性状尤其是单株产量方面没有明显差异, 表明 Pi2 和 Pigm 位点附近基本不存在与产量性状相关的累赘连锁, 通过常规筛选结合分子标记辅助选择可以达到选育广谱抗瘟基因的优良品种的目标。

3 讨论

植物中的抗性基因往往具有成簇分布于特定的染色体区域、共同组成紧密连锁的复合抗病基因座的特性^[21,22,23]。水稻中已定位的77个稻瘟病抗性基因, 超过半数以基因簇形式存在, 位于第6染色体短臂的 Pi2 、 Pi9 、 Piz 、 Piz-t 等构成一个复合抗性基因座, 不同种质间在该位点包含有数目不等的NBS-LRR类抗病基因。种质间单元型分析表明, 直系同源基因之间在序列和大小上具有极高的相似性, 而旁系同源基因的序列歧化显著。已克隆的 Pi2 、 Piz-t、Pi9 位点均包含有9个NBS-LRR类抗病基因簇, 但在分子水平上, Pi2 与 Piz-t 为直系同源基因, 8个氨基酸的差异造成了其抗性专化性的差异, 它们是真正的等位基因, 而 Pi9 是 Pi2 、 Piz-t 的旁系同源基因, 是其紧密连锁基因^[24,25]。 Pigm 是谷梅4号中新鉴定的稻瘟病抗性基因, 初步的作图分析表明其可能与 Pi2 、 Pi9 等位或连锁^[8]。本文构建了广占63S、97S、R609和R084四个遗传背景下 Pigm 、 Pi2 两个基因的系列近等基因系, 苗期接种鉴定结果显示 Pigm 与 Pi2 对不同生态区分离的204个菌株均具有较高的抗性频率(>70%), 而两者抗谱重叠仅为54.4%~65.7%, 且穗瘟人工接种和重发病区病圃的自然诱发鉴定结果与苗期接种趋势基本一致, 表明 Pigm 是一个与 Pi2 抗谱差异明显的、等位或紧密连锁的广谱抗性基因, 对稻瘟病抗性育种具有重要应用价值。

表3 近等基因系的基本农艺性状 Table 3 Basic agronomic traits of NILs carrying target genes

聚合抗性基因是获得广谱抗性的重要途径, 但随着聚合基因数目的增加, 不良性状的连锁累赘也可能增加。杨健源等^[26]提出选择目标基因时, 用少数的抗性基因获得广谱抗性是抗稻瘟病育种中必须考虑的问题。已有部分研究表明聚合 Pi1 、 Pi2 基因对我国南方稻区稻瘟病抗性有良好效果^[11,12,13], 本文利用携有 Pi1 的不育系, 与携有 Pigm、Pi2 的恢复系配组, 研究 Pi1 与 Pigm 的聚合效应, 发现聚合有 Pi1 / Pigm 和 Pi1 / Pi2 的杂交稻, 其抗性频率均超过90%, 表明利用 Pigm 、 Pi2 与 Pi1 等少数广谱稻瘟病抗性基因进行聚合, 可达到有效改良抗性的目的。

本文针对 Pigm 、 Pi2 构建的近等基因系, 在每一个回交分离世代都选择与轮回亲本最相似的个体进行回交, 最后获得的4个遗传背景下的系列近等基因系, 其产量等基本农艺性状与轮回亲本绝大多数无显著差异, 表明回交过程中表型选择的有效性, 同时也表明 Pigm 、 Pi2 基因位点附近存在较少的与产量性状相关累赘连锁, 较易实现丰产基础上的抗性改良。但需要提及的是, 谷梅4号中 Pigm 附近存在一个与其连锁的迟抽穗期基因, 在自然长日照条件下(如江苏扬州)会显著延迟抽穗; 在自然短日条件下(如海南)表现明显的感光特性而提早抽穗。迟抽穗期基因与 Pigm 的重组率约3%。本文获得系列近等基因系, 均是通过第一个分离回交世代大群体选择、打破这种连锁后而成功构建的, 因而表现出与轮回亲本生育期的基本一致性。水稻第6染色体短臂上分布有 Hd1 等生育期基因, 且拥有不同效应的等位基因型^[27]。谷梅4号中与 Pigm 连锁的迟抽穗期基因, 是否是 Hd1 位点的强效应等位基因, 亦或是其他新的生育期基因, 有待后续研究进一步验证。

4 结论

Pigm 是与 Pi2 抗谱有显著差异的广谱稻瘟病抗性基因, 并与 Pi1 有良好的聚合效应, 对我国水稻稻瘟病抗性改良具有重要应用价值。

The authors have declared that no competing interests exist.

作者已声明无竞争性利益关系。The authors have declared that no competing interests exist.

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2002

2.635

0.0

Mol Genet Genomics. 2002, 267(4):472 - 480 DOI:10.1007/s00438-002-0677-2

Two broad-spectrum blast resistance genes, Pi9(t) and Pi2(t), are physically linked on rice chromosome 6

1.Department of Plant Pathology, The Ohio State University, 201 Kottman Hall, 2021 Coffey Road, Columbus, OH 43210, USA

... 其中, 第6染色体短臂近着丝粒的附近分布有Pi2 、Pi9 、Pi22 、Pi25 、Pi26 、Pi40 、Pi42 、Pi50、Piz 、Piz-t 等至少10个抗性基因, 这些基因在抗病基因簇内呈等位或紧密连锁关系, 并有研究结果显示其抗谱存在明显差异^{[1,2,3,4,5,6,7]} ...

2000

1.554

0.0

Euphytica. 2000, 116(1):17 - 22 DOI:10.1023/A:1004045812357

Molecular mapping of a new gene for resistance to rice blast (Pyricularia grisea Sacc.)

Sang-Nag Ahn(1) Yeon-Kyu Kim(1) Ha-Cheol Hong(1) Seong-Sook Han(2) Soo-Jin Kwon(1) Hae-Chune Choi(1) Huhn-Pal Moon(1) Susan R. McCouch(3)

1.Rural Development Administration Rice Breeding Division, National Crop Experiment Station Suweon 441-100 Korea Suweon 441-100 Korea
2.Rural Development Administration National Institute of Agricultural Science and Technology Suweon 441-100 Korea Suweon 441-100 Korea
3.Cornell University Department of Plant Breeding & Biometry Ithaca NY 14853 U.S.A. Ithaca NY 14853 U.S.A.

A single dominant blast resistance gene conferring resistance to a Korean rice blast isolate was identified in rice variety `Suweon 365'. We report the chromosomal localization and molecular mapping of this blast resistance gene designated asPi-18, which confers resistance to Korean isolate `KI-313' of the blast pathogen. To know whether there is a relationship among genes conditioning resistance to location-specific isolates of the blast pathogen and thereby to identify linked markers to resistance gene for isolate KI-313 collected in Korea, RFLP markers previously reported to be linked to major blast resistance genes in different rice germplasm and other markers mapped to nearby regions were surveyed for polymorphism between a resistant (`Suweon 365') and a susceptible (`Chucheongbyeo') parent. Linkage associations of the RFLP markers with the resistance gene were verified using an F2 and F3 segregating population of known blast reaction. RFLP analysis showed thatPi-18 was located near the end of chromosome 11, linked to a single copy clone RZ536 at a distance of 5.4 centiMorgans (cM) and that this gene was different fromPi-1(t). An allelism test revealed that this gene was also different fromPi-k. Currently, a combination of RAPD and microsatellite primers is being employed to find additional markers in this region. Tightly linked DNA markers will facilitate selection for resistant genotypes in breeding programs and provide the basis for map based cloning of this new blast resistance gene.

2005

3.297

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Theor Appl Genet. 2005, 111(1):50 - 56 DOI:10.1007/s00122-005-1971-2

Genetic control of rice blast resistance in the durably resistant cultivar Gumei?2 against multiple isolates

J.-L. Wu(13) Y.-Y. Fan(1) D.-B. Li(2) K.-L. Zheng(1) H. Leung(3) J.-Y. Zhuang(1)

1.Chinese National Centre for Rice Improvement and State Key Laboratory of Rice Biology China National Rice Research Institute Hangzhou 310006 China
2.Institute of Biotechnology Zhejiang University Hangzhou 310029 China
3.Entomology and Plant Pathology Division International Rice Research Institute DAPO P.O. Box 7777 Metro Manila Philippines

To further our understanding of the genetic control of blast resistance in rice cultivar Gumei 2 and, consequently, to facilitate the utilization of this durably blast-resistant cultivar, we studied 304 recombinant inbred lines of indica rice cross Zhong 156/Gumei 2 and a linkage map comprising 181 markers. An analysis of segregation for resistance against five isolates of rice blast suggested that one gene cluster and three additional major genes that are independently inherited are responsible for the complete resistance of Gumei 2. The gene cluster was located to chromosome 6 and includes two genes mapped previously,Pi25(t), against Chinese rice blast isolate 92-183 (race ZC15) andPi26(t) against Philippine rice blast isolate Ca89 (lineage 4), and a gene for resistance against Philippine rice blast isolate 92330-5 (lineage 17). Of the two genes conferring resistance against the Philippine isolates V86013 (lineage 15) and C923-39 (lineage 46), we identified one asPi26(t) and mapped the other onto the distal end of chromosome 2 wherePib is located. We used three components of partial blast resistance, percentage diseased leaf area (DLA), lesion number and lesion size, all measured in the greenhouse, to measure the degree of susceptibility to isolates Ca89 and C923-39 and subsequently identified nine and eight quantitative trait loci (QTLs), respectively. Epistasis was determined to play an important role in partial resistance against Ca89. Using DLA measured on lines susceptible in a blast nursery, we detected six QTLs. While different QTLs were detected for partial resistance to Ca89 and C923-39, respectively, most were involved in the partial resistance in the field. Our results suggest that the blast resistance in Gumei 2 is controlled by multiple major genes and minor genes with epistatic effects.

2007

3.297

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Theor Appl Genet. 2007, 115(8):1163 - 1177 DOI:10.1007/s00122-007-0642-x

A novel gene,Pi40(t), linked to the DNA markers derived from NBS-LRR motifs confers broad spectrum of blast resistance in rice

J. U. Jeung(123) B. R. Kim(4) Y. C. Cho(3) S. S. Han(4) H. P. Moon(3) Y. T. Lee(3) K. K. Jena(12)

1.International Rice Research Institute Plant Breeding, Genetics, and Biotechnology Division Los Ba?os Laguna Philippines
2.National Institute of Crop Science, RDA IRRI-Korea Office Suwon 441-857 Republic of Korea
3.National Institute of Crop Science, RDA Genetics and Breeding Division Suwon 441-857 Republic of Korea
4.National Institute of Agricultural Science and Technology, RDA Plant Pathology Division Suwon 441-707 Republic of Korea

Rice blast disease caused byMagnaporthe grisea is a continuous threat to stable rice production worldwide. In a modernized agricultural system, the development of varieties with broad-spectrum and durable resistance to blast disease is essential for increased rice production and sustainability. In this study, a new gene is identified in the introgression line IR65482-4-136-2-2 that has inherited the resistance gene from an EE genome wildOryza species,O. australiensis (Acc. 100882). Genetic and molecular analysis localized a major resistance gene,Pi40(t), on the short arm of chromosome 6, where four blast resistance genes (Piz, Piz-5, Piz-t, andPi9) were also identified, flanked by the markers S2539 and RM3330. Throughe-Landing, 14 BAC/PAC clones within the 1.81-Mb equivalent virtual contig were identified on Rice Pseudomolecule3. Highly stringent primer sets designed for 6 NBS-LRR motifs located within PAC clone P0649C11 facilitated high-resolution mapping of the new resistance gene,Pi40(t). Following association analysis and detailed haplotyping approaches, a DNA marker, 9871.T7E2b, was identified to be linked to thePi40(t) gene at the 70 Kb chromosomal region, and differentiated thePi40(t) gene from the LTH monogenic differential lines possessing genesPiz, Piz-5, Piz-t, andPi-9.Pi40(t) was validated using the most virulent isolates of Korea as well as the Philippines, suggesting a broad spectrum for the resistance gene. Marker-assisted selection (MAS) and pathotyping of BC progenies having two japonica cultivar genetic backgrounds further supported the potential of the resistance gene in rice breeding. Our study based on new gene identification strategies provides insight into novel genetic resources for blast resistance as well as future studies on cloning and functional analysis of a blast resistance gene useful for rice improvement.

2012

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Theor. Appl. Genet.. 2012, 124(7):1295-304 DOI:10.1007/s00122-012-1787-9

The identification of Pi50(t), a new member of the rice blast resistance Pi2/Pi9 multigene family.

Zhu X, Chen S, Yang J, Zhou S, Zeng L, Han J, Su J, Wang L, Pan Q

Laboratory of Plant Resistance and Genetics, College of Resources and Environmental Sciences, South China Agricultural University, Guangzhou 510642, China. zhuxy@gdppri.com

The deployment of broad-spectrum resistance genes is the most effective and economic means of controlling blast in rice. The cultivar Er-Ba-Zhan (EBZ) is a widely used donor of blast resistance in South China, with many cultivars derived from it displaying broad-spectrum resistance against blast. Mapping in a set of recombinant inbred lines bred from the cross between EBZ and the highly blast-susceptible cultivar Liangjiangxintuanheigu (LTH) identified in EBZ a blast resistance gene on each of chromosomes 1 (Pish), 6 (Pi2/Pi9) and 12 (Pita/Pita-2). The resistance spectrum and race specificity of the allele at Pi2/Pi9 were both different from those present in other known Pi2/Pi9 carriers. Fine-scale mapping based on a large number of susceptible EBZ × LTH F(2) and EBZ × LTH BC(1)F(2) segregants placed the gene within a 53-kb segment, which includes Pi2/Pi9. Sequence comparisons of the LRR motifs of the four functional NBS-LRR genes within Pi2/Pi9 revealed that the EBZ allele is distinct from other known Pi2/Pi9 alleles. As a result, the gene has been given the designation Pi50(t).

PMID: 22270148

2006

3.297

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Theor Appl Genet. 2006, 113(2):251 - 260 DOI:10.1007/s00122-006-0290-6

Development of PCR-based allele-specific and InDel marker sets for nine rice blast resistance genes

K. Hayashi(1) H. Yoshida(1) I. Ashikawa(2)

1.National Agricultural Research Center, 1-2-1 Inada, Joetsu, 943-0193 Japan
2.National Institute of Crop Science, 2-1-18 Kannondai, Tsukuba, 305-8518 Japan

Blast resistance is one of the most important traits in rice breeding, and application of molecular markers for blast resistance breeding is likely to allow the rapid screening for the trait during early growth stages, without the need for inoculation of pathogen and phenotyping. Allele-specific PCR markers and insertion/deletion (InDel) markers, which genotype single-nucleotide polymorphisms and InDel polymorphisms, respectively, are useful tools for marker-assisted selections. We developed sets of allele-specific PCR and InDel markers for nine rice blast resistance genes—Piz, Piz-t, Pit, Pik, Pik-m, Pik-p, Pita, Pita-2, andPib—which are commonly used in Japanese blast resistance rice breeding programs. For each resistance gene, we used the segregation information from thousands of progeny in several crosses or published gene locations to generate a marker that cosegregated with the gene and markers that closely flanked the gene on either side. The developed cosegregating markers uniquely discriminated among each of the lines with the individual resistance genes (except forPita andPita-2). Therefore, these markers will likely facilitate the development of multiline cultivars carrying one or a combination of these nine blast resistance genes. In addition, the systems we developed may be valuable tools in the quality control of seed production from blast-resistant multiline cultivars.

2006

4.007

0.0

2006

3.297

0.0

Theor Appl Genet. 2006, 113(4):705 - 713 DOI:10.1007/s00122-006-0338-7

Genetic characterization and fine mapping of the blast resistance locusPigm(t) tightly linked toPi2 andPi9 in a broad-spectrum resistant Chinese variety

Yiwen Deng(1) Xudong Zhu(2) Ying Shen(2) Zuhua He(1)

1.National Key Laboratory of Plant Molecular Genetics, Institute of Plant Physiology and Ecology, Shanghai Institutes for Biological Sciences, The Chinese Academy of Sciences (CAS), 300 Fenglin Road, Shanghai, 200032 China
2.State Key Laboratory of Rice Biology, China National Rice Research Institute, Hangzhou, 310006 China

The identification and utilization of broad-spectrum resistance genes have been proven the most effective and economical approach to control rice blast disease. To understand the molecular mechanism of broad-spectrum resistance to rice blast, we conducted genetic and fine mapping analysis of the blast resistance gene in a Chinese rice variety: Gumei 4 (GM4) identified with broad-spectrum resistance and used in rice breeding for blast resistance for more than 20 years. Genetic and mapping analysis indicated that blast resistance to nine isolates of different Chinese races in GM4 was controlled by the same dominant locus designated asPigm(t) that was finely mapped to an approximately 70-kb interval between markers C5483 and C0428 on chromosome 6, which contains five candidate NBS--LRR disease resistance genes. The allelism test showed thatPigm(t) was either tightly linked or allelic toPi2 andPi9, two known blast resistance genes. Mapping information also indicated that another blast resistance genePi26(t) might also be located at the same region. Candidate genes were identified by sequence analysis of the Nipponbare andPi9 locus and the corresponding region in GM4. Sequence divergence of candidate genes was observed between GM4 and model varieties Nipponbare and 9311, andPi9. Our current study provides essential information and new genetic resource for the cloning of functional resistance gene(s) and for marker-assisted selection in rice breeding for broad-spectrum blast resistance.

... Pigm 是从持久抗稻瘟病品种谷梅4号中鉴定的一个新的广谱稻瘟病抗性基因, 定位及等位性测验结果显示其可能与第6染色体的Pi2 和Pi9 等位或连锁^[8] ...

... Pigm 是谷梅4号中新鉴定的稻瘟病抗性基因, 初步的作图分析表明其可能与Pi2 、Pi9 等位或连锁^[8] ...

2005

0.0

0.619

Journal of Fujian Agriculture and Forestry University(Natural Science Edition). 2005, (1):74 - 77

The screening of molecular markers closely linked to rice blast resistant gene Pi-1 and their application

稻瘟病抗性基因Pi-1连锁SSR标记的筛选和应用

CHENZhi-Wei;GUANHua-zhong;WUWei-ren;ZHOUYuan-chang;HANQing-dian

陈志伟;官华忠;吴为人;周元昌;韩庆典

根据稻瘟病抗性基因Pi-1在水稻物理图谱和水稻微卫星(SSR)遗传图谱上的位置信息,获得在抗性供体亲本(LAC23)与受体亲本(珍汕97B、金山B-1和金山S-1)间扩增产物表现出多态性的SSR标记MGR4766,它与Pi-1的遗传距离仅为1.3 cM;分子标记辅助选择选择准确性验证表明:RM224对Pi-1选择的准确率可达90%以上,而MRG4766对Pi-1选择的准确率为98%-100%;同时利用RM224和MRG4766对Pi-1选择的准确率高达100%.

... 3 cM处, 现已经被成功克隆^[9,10], 并有研究显示其与Pi2 有良好的互补效应^[11,12,13] ...

... 1 供试材料的选择Pi1 和Pi2 基因的供体亲本为W1, 是以金23B为轮回亲本导入Pi1 和Pi2 基因的中间材料^[9] ...

2012

3.297

0.0

Theor. Appl. Genet.. 2012, 125(5):1047-55 DOI:10.1007/s00122-012-1894-7

The isolation of Pi1, an allele at the Pik locus which confers broad spectrum resistance to rice blast.

Hua L, Wu J, Chen C, Wu W, He X, Lin F, Wang L, Ashikawa I, Matsumoto T, Wang L, Pan Q

State Key Laboratory for Conservation and Utilization of Subtropic Agrobioresources, Laboratory of Plant Resistance and Genetics, College of Natural Resources and Environmental Sciences, South China Agricultural University, Guangzhou 510642, China.

We report the isolation of Pi1, a gene conferring broad-spectrum resistance to rice blast (Magnaporthe oryzae). Using loss- and gain-of-function approaches, we demonstrate that Pi1 is an allele at the Pik locus. Like other alleles at this locus, Pi1 consists of two genes. A functional nucleotide polymorphism (FNP) was identified that allows differentiation of Pi1 from other Pik alleles and other non-Pik genes. A extensive germplasm survey using this FNP reveals that Pi1 is a rare allele in germplasm collections and one that has conferred durable resistance to a broad spectrum of pathogen isolates.

PMID: 22643901

... 3 cM处, 现已经被成功克隆^[9,10], 并有研究显示其与Pi2 有良好的互补效应^[11,12,13] ...

2008

0.0

1.4083

Chin J Rice Sci. 2008, 22(1):23 - 27

Pyramiding Three Genes with Resistance to Blast by MarkerAssisted Selection to Improve Rice Blast Resistance of Jin 23B

利用分子标记技术聚合3个稻瘟病基因改良金23B的稻瘟病抗性

CHEN Hong-qi , CHEN Zong-xiang , NI Shen , ZUO Shi-min , PAN Xue-biao , ZHU Xu-dong

陈红旗1;陈宗祥2;倪深1;左示敏2;潘学彪2，*;朱旭东1，*

Using rice lines C101LAC and C101A51 as the donors of the resistance genes to rice blast, three resistance genes (Pi1, Pi2 and Pi33) were introgressed into an elite variety Jin 23B by crossing, backcrossing and multicrossing. Six nearisogenic lines of Jin 23B with the three resistance genes were obtained through traditional breeding method combined with markerassisted selection method, and the resistance frequency was 96.7%. The results indicated that the resistance frequency of the lines with the three resistance genes were higher than that of single resistance gene line. It could be concluded that gene pyramiding is an effective approach for the improvement of rice varieties with durable resistance to rice blast.

以C101LAC和C101A51为稻瘟病抗性基因的供体亲本，金23B为受体亲本，通过杂交、复交及一次回交，在分离世代，利用分子标记辅助选择技术结合特异稻瘟病菌株接种鉴定和农艺性状筛选，获得6个导入Pi1、Pi2和Pi33基因的金23B导入系，其中导入系W1对稻瘟病的抗病频率为96.7%，明显高于携带单个基因的C104LAC（Pi1）、C101A51（Pi2）和北京糯（Pi33）。基因聚合后抗病频率提高，说明基因聚合是培育稻瘟病持久抗性的有效方法之一。

... 3 cM处, 现已经被成功克隆^[9,10], 并有研究显示其与Pi2 有良好的互补效应^[11,12,13] ...

... 2 Pigm 、Pi2 与Pi1 的互作效应有研究表明Pi2 与另一广谱稻瘟病抗性基因Pi1 具有良好的互补效应^[11,12,13] ...

... 已有部分研究表明聚合Pi1 、Pi2 基因对我国南方稻区稻瘟病抗性有良好效果^[11,12,13], 本文利用携有Pi1 的不育系, 与携有Pigm、Pi2 的恢复系配组, 研究Pi1 与Pigm 的聚合效应, 发现聚合有Pi1 /Pigm 和Pi1 /Pi2 的杂交稻, 其抗性频率均超过90%, 表明利用Pigm 、Pi2 与Pi1 等少数广谱稻瘟病抗性基因进行聚合, 可达到有效改良抗性的目的 ...

2008

0.0

1.8267

Acta Agron Sin. 2008, 34(7):1128 - 1136

Improvement of Rice Blast Resistance in TGMS Line by Pyramiding ofPi-1andPi-2 through Molecular Marker-Assisted Selection

利用分子标记辅助选择聚合Pi-1和Pi-2基因改良两系不育系稻瘟病抗性

LIU Wu-Ge¹,WANG Feng^1*,JIN Su-Juan¹²,ZHU Xiao-Yuan³,LI Jin-Hua¹,LIU Zhen-Rong¹,LIAO Yi-Long¹,ZHU Man-Shan¹,HUANG Hui-Jun¹, FU Fu-Hong¹,LIU Yi-Bai²

柳武革¹;王丰^1,*;金素娟^1,2;朱小源³;李金华¹;刘振荣¹;廖亦龙¹;朱满山¹;黄慧君¹;符福鸿¹;刘宜柏²

1 Rice Research Institute, Guangdong Academy of Agricultural Sciences, Guangzhou 510640, Guangdong; 2 Jiangxi Agricultural University, Nanchang 330045, Jiangxi; 3 Plant Protection Institute, Guangdong Academy of Agricultural Sciences, Guangzhou 510640, Guangdong, China

Blast is one of the most serious diseases of rice worldwide, which usually leads to a sharp decline in rice production and even results in no yield. Breeding blast-resistant varieties is one of the most effective and economical ways to control the disease. In this study, the BL122 contained two blast resistance genesPi-1 andPi-2 was used as gene donor to be crossed and then backcrossed with the TGMS line GD-7S. Five improved TGMS lines withPi-1 andPi-2 homozygous genotype were successfully developed through molecular marker-assisted selection. The resistance frequency of the improved TGMS lines ranged from 94.12% to 97.06%, much higher than that of the check GD-7S, which was only 17.65%, by artificially inoculating 34 representa-tive blast isolates collected in Guangdong Province. The improved TGMS lines also displayed high resistance to leaf and neck blast in the epidemic areas. The critical temperature of fertility transformation for the improved TGMS lines from fertile to sterile was preliminarily identified in the phytotron with the regime of 12.5 h/23℃ and natural conditions. The results showed that the critical temperature point for the improved TGMS lines was likely consistent with that of the check GD-7S. The statistical analysis showed there was no significant difference in the agronomic traits between improved TGMS lines and GD-7S except the flag leaf length and number of panicles per plant. Moreover, there was also no significant difference in most of the agronomic traits be-tween the F1s derived from the crosses of the same restorer line L38 with both the check GD-7S and the improved TGMS lines, respectively. All these indicated that the improved TGMS lines retained almost the same agronomic traits and combining ability as the check.

以含广谱稻瘟病抗性基因Pi-1和Pi-2的BL122为供体, 温敏核不育系GD-7S为受体, 通过杂交、回交和自交并结合分子标记辅助选择, 将Pi-1、Pi-2基因导入温敏核不育系GD-7S中, 获得5个携带两个抗性基因的纯合改良不育株系。利用34个广东代表性稻瘟病菌株接种鉴定, 5个改良株系的抗性频率为94.12%~97.06%, 而对照GD-7S抗性频率仅为17.65%; 自然病圃诱发鉴定5个改良株系的叶瘟和穗颈瘟均为0级, 表现高抗。经自然条件和人工气候箱育性鉴定, 改良株系与对照均为无或少花粉败育类型, 自交结实率为0, 说明不育起点温度与对照基本一致。统计分析表明, 除剑叶长和每株穗数外, 改良株系与对照在其他农艺性状方面均无显著差异。与恢复系L38杂交, 改良株系的杂种F1与对照的F1大多农艺性状无显著差异, 说明改良株系基本保持了GD-7S的农艺性状和配合力。

... 3 cM处, 现已经被成功克隆^[9,10], 并有研究显示其与Pi2 有良好的互补效应^[11,12,13] ...

... 7的选择效率高达93%以上, 与前人研究结果一致^[12] ...

... 2 Pigm 、Pi2 与Pi1 的互作效应有研究表明Pi2 与另一广谱稻瘟病抗性基因Pi1 具有良好的互补效应^[11,12,13] ...

2012

0.0

... 3 cM处, 现已经被成功克隆^[9,10], 并有研究显示其与Pi2 有良好的互补效应^[11,12,13] ...

... 2 Pigm 、Pi2 与Pi1 的互作效应有研究表明Pi2 与另一广谱稻瘟病抗性基因Pi1 具有良好的互补效应^[11,12,13] ...

1992

0.0

1.4083

Chin J Rice Sci. 1992, 6(1):47 - 48

A Simple Method for Isolation of Rice DNA

提取水稻DNA的一种简易方法

Lu Yangjiang; Zheng Kangle

卢扬江1; 郑康乐2

... 3 标记检测参照卢扬江等^[14]的方法提取水稻全基因组DNA ...

1992

4.007

0.0

... 参照Hospital等^[15]的方法计算株系背景回复率, G_g = (L + X_g )/2L , 其中, G_g 为在g 代的遗传背景回复率 ...

2009

0.0

... 5 病原菌的分离与培养从广东、江苏、海南、浙江和湖北5省重发病区田间采集自然发生的稻瘟病病穗, 参考周益军等的方法^[16]培养病菌, 单孢分离后在PDA培养基上培养5~7 d, 转于产孢培养基上培养5~7 d, 用无菌水洗去培养基表面菌丝后于黑光灯下照射培养72 h (培养温度均为26℃) ...

2010

0.0

1.0171

Jiangsu Journal of Agricultural Sciences. 2010, (5):948 - 953

Resistances of Four Sets of Rice Varieties to 19 Strains of Magnaporthe griseas in Jiangsu Area

4套水稻稻瘟病鉴别品种对江苏地区主要稻瘟病菌株的抗性分析

ZHANGLei^[1];ZHANGQi-jun^[2];CHENGZhao-bang^[3];YANGHao^[3];ZHOUYi-jun^[3];L(U)Chuan-gen^[2]

张磊^[1];张启军^[2];程兆榜^[3];杨豪^[3];周益军^[3];吕川根^[2]

为构建适于目前江苏地区稻瘟病的水稻鉴别品种,区分不同菌株的毒力,该研究用江苏地区19个主要稻瘟病菌群的菌株,分别在水稻苗期和孕穗期以人工接种的方法,对水稻稻瘟病的中国鉴别品种(CRDS)、日本清泽单基因系鉴别品种(TMGLs)、中国农业科学院构建的丽江新团黑谷近等基因系鉴别品种(LTH NILs)和国际水稻研究所构建的C039系列近等基因系(C039 NILS)进行鉴定.病情调查和统计结果表明,苗瘟与穗颈瘟之间存在极显著的正相关(r=0.599,P<0.01).清泽单基因系和丽江新团黑谷近等基因系对各菌株的抗性差异都较明显,可作为较好的鉴别品种.江苏地区目前毒力较强的稻瘟病菌群是ZD1和ZF1生理小种,尤其是ZF1菌群,这4套稻瘟病鉴别品种对该小种的4个菌株均表现较高的感病率.根据目前江苏地区的水稻品种和稻瘟病菌群特征,将LTH NILs和TMGLs的爱知旭、K3、福锦、K59搭配使用,较适宜作为江苏地区的水稻稻瘟病鉴别品种.

... 穗瘟鉴定评价标准参见罗楚平等^[17]的方法 ...

2006

0.0

... 由于穗瘟对水稻的产量影响更为重要, 而关于苗瘟与穗瘟抗性是否相关, 针对不同基因以及不同的研究材料, 一直有不同的研究结论^[18,19,20] ...

2010

0.0

1.0251

J Agric Sci Tech China. 2010, (1):100 - 105

Applications of Pi-z、Pi-ta2 Genes in Japonica Rice Breeding with Blast Resistance in Yunnan

Pi-z、Pi-ta2等基因在云南粳稻稻瘟病抗性育种中的应用

ZHANGXue-tang;LIAOXin-hua;ZHUZhen-hua;SUZhen-xi;SHIRong;ZHAOGuo-zhen;JIANGCong;DAILu-yuan

张学堂;廖新华;朱振华;苏振喜;世荣;赵国珍;蒋聪;戴陆园

采用与育种同步的抗性鉴定方法,利用日本鉴别菌系作为鉴别标准品种,分析了Pi-z、Pi-ta2等基因2002-2007年在云南水稻稻瘟病抗性育种实践中的表现.结果表明,截至2007年,Pi-z、Pi-ta2、Pi-zt在云南稻区仍然有利用价值;Pi-ta2叶/穗瘟抗性反应表现明显不一致;Pi-z、Pi-zt在各种育种材料中叶/穗瘟抗性表现一致;在不同抗性组合中,发现Pi-z对育种材料的抗性改良效果更明显.

... 由于穗瘟对水稻的产量影响更为重要, 而关于苗瘟与穗瘟抗性是否相关, 针对不同基因以及不同的研究材料, 一直有不同的研究结论^[18,19,20] ...

1991

1.554

0.0

... 由于穗瘟对水稻的产量影响更为重要, 而关于苗瘟与穗瘟抗性是否相关, 针对不同基因以及不同的研究材料, 一直有不同的研究结论^[18,19,20] ...

1995

9.737

0.0

... 3 讨论植物中的抗性基因往往具有成簇分布于特定的染色体区域、共同组成紧密连锁的复合抗病基因座的特性^[21,22,23] ...

2003

3.297

0.0

Theor Appl Genet. 2003, 106(8):1467 - 1472 DOI:10.1007/s00122-003-1205-4

Genetic and physical mapping of a new gene for bacterial blight resistance in rice

1., National Key Laboratory of Crop Genetic Improvement, National Center of Crop Molecular Breeding, Huazhong Agricultural University, Wuhan 430070, China

... 3 讨论植物中的抗性基因往往具有成簇分布于特定的染色体区域、共同组成紧密连锁的复合抗病基因座的特性^[21,22,23] ...

2006

4.431

0.0

Mol. Plant Microbe Interact.. 2006, 19(11):1216-28

The eight amino-acid differences within three leucine-rich repeats between Pi2 and Piz-t resistance proteins determine the resistance specificity to Magnaporthe grisea.

Zhou B, Qu S, Liu G, Dolan M, Sakai H, Lu G, Bellizzi M, Wang GL

Department of Plant Pathology, Ohio State University, Columbus, OH 43210, USA.

The rice blast resistance (R) genes Pi2 and Piz-t confer broad-spectrum resistance against different sets of Magnaporthe grisea isolates. We first identified the Pi2 gene using a map-based cloning strategy. The Pi2 gene is a member of a gene cluster comprising nine gene members (named Nbs1-Pi2 to Nbs9-Pi2) and encodes a protein with a nucleotide-binding site and leucine-rich repeat (LRR) domain. Fine genetic mapping, molecular characterization of the Pi2 susceptible mutants, and complementation tests indicated that Nbs4-Pi2 is the Pi2 gene. The Piz-t gene, a Pi2 allele in the rice cultivar Toride 1, was isolated based on the Pi2 sequence information. Complementation tests confirmed that the family member Nbs4-Piz-t is Piz-t. Sequence comparison revealed that only eight amino-acid changes, which are confined within three consecutive LRR, differentiate Piz-t from Pi2. Of the eight variants, only one locates within the xxLxLxx motif. A reciprocal exchange of the single amino acid between Pi2 and Piz-t did not convert the resistance specificity to each other but, rather, abolished the function of both resistance proteins. These results indicate that the single amino acid in the xxLxLxx motif may be critical for maintaining the recognition surface of Pi2 and Piz-t to their respective avirulence proteins.

PMID: 17073304

... 3 讨论植物中的抗性基因往往具有成簇分布于特定的染色体区域、共同组成紧密连锁的复合抗病基因座的特性^[21,22,23] ...

2010

3.297

0.0

Theor Appl Genet. 2010, 121(2):295 - 309 DOI:10.1007/s00122-010-1310-0

Genomic structure and evolution of thePi2/9 locus in wild rice species

Liangying Dai(1) Jun Wu(12) Xunbo Li(2) Xuejun Wang(1) Xionglun Liu(1) Chatchawan Jantasuriyarat(3) Dave Kudrna(5) Yeisoo Yu(5) Rod A. Wing(5) Bin Han(2) Bo Zhou(24) Guo-Liang Wang(13)

1.Hunan Provincial Key Laboratory of Crop Germplasm Innovation and Utilization, Hunan Agricultural University, Changsha, 410128 China
2.National Center For Gene Research, Chinese Academy of Sciences, Shanghai, 200233 China
3.Department of Plant Pathology, Ohio State University, Columbus, OH 43210, USA
4.Institute of Biotechnology, Zhejiang University, Hangzhou, 310029 China
5.Arizona Genomics Institute, School of Plant Sciences and BIO5 Institute for Collaborative Research, University of Arizona, Tucson, AZ 85721, USA

Rice blast, caused by the fungal pathogenMagnaporthe oryzae, is a devastating disease of rice worldwide. Among the 85 mapped resistance (R) genes against blast, 13 have been cloned and characterized. However, how these genes originated and how they evolved in theOryza genus remains unclear. We previously cloned the rice blast R-genesPi2, Pi9, andPiz-t, and analyzed their genomic structure and evolution in cultivated rice. In this study, we determined the genomic sequences of thePi2/9 locus in four wildOryza species representing three genomes (AA, BB and CC). The number ofPi2/9 family members in the four wild species ranges from two copies to 12 copies. Although these genes are conserved in structure and categorized into the same subfamily, sequence duplications and subsequent inversions or uneven crossing overs were observed, suggesting that the locus in different wild species has undergone dynamic changes. Positive selection was found in the leucine-rich repeat region of most members, especially in the largest clade wherePi9 is included. We also provide evidence that thePi9 gene is more related to its homologues in the recurrent line and other rice cultivars than to those in its alleged donor speciesO. minuta, indicating a possible origin of thePi9 gene fromO. sativa. Comparative sequence analysis between the four wildOryza species and the previously established reference sequences in cultivated rice species at thePi2/9 locus has provided extensive and unique information on the genomic structure and evolution of a complex R-gene cluster in theOryza genus.

... 已克隆的Pi2 、Piz-t、Pi9 位点均包含有9个NBS-LRR类抗病基因簇, 但在分子水平上, Pi2 与Piz-t 为直系同源基因, 8个氨基酸的差异造成了其抗性专化性的差异, 它们是真正的等位基因, 而Pi9 是Pi2 、Piz-t 的旁系同源基因, 是其紧密连锁基因^[24,25] ...

2008

0.0

1.4083

Chin J Rice Sci. 2008, 22(2):190 - 196

Evaluation on Resistance of Major Rice Blast Resistance Genes toMagnaporthe grisea Isolates Collected from indica Rice in Guangdong Province,China

稻瘟病主效抗性基因对广东省籼稻稻瘟病菌的抗性评价

YANG Jian-yuan , CHEN Shen , ZENG Lie-xian , LI Yi-long , CHEN Zhen , ZHU Xiao-yuan

杨健源;陈深;曾列先;李亦龙;陈珍;朱小源*

Race specific resistance and field resistance of 30 monogenic rice lines resistant to rice blast,derived from different resources were evaluated.The resistance spectrum(RS) of the tested monogenic lines ranged from 0.6% to 89.6%.Most of the monogenic lines showed narrow RS and high susceptibility in the blast zone,but the lines with Pikh and Pi1(t) showed good and stable blast resistance in field with the RSs of 89.6% and 82.2%,respectively.According to the cluster analysis of specific resistance to 163 blast isolates collected from indica rice in Guangdong Province,China,the 30 monogenic lines were divided into 15 groups,and based on the principal component analysis,9 kinds of race-specific resistance were identified.Pik,Piz5,Pi9,Pish could be used as candidate resistance genes for future rice breeding since their specific resistance differed from those of the key parents in Guangdong.Pyramiding of Pikh[orPi1(t)],Pi9(orPiz5) andPish(orPita2) will be an effective measure to obtain broad spectrum blast resistance in local rice breeding program.The strategy for studying rice blast resistance genes and its application was discussed.

对来自不同抗源的30个抗稻瘟病单基因系进行了小种专化抗性及田间抗性的分析和评价。30个单基因系对163个广东省稻瘟病菌株的抗谱介于0.6%~89.6%，大多数单基因系对测试的病菌表现出较窄的抗谱，在病区表现为高度感病，但含有Pikh 和 Pi1(t)的单基因系表现出广谱抗性，抗谱分别为89.6% 和 82.2%，在病区表现出较好的田间抗性。基于抗性基因与163个稻瘟病菌的抗感互作聚类分析，将30个单基因系的小种专化抗性划分为15个类型。主成分因子分析结果表明，其中9个类型的小种专化抗性表现出较大差异，可解释总变异的81.3%。含有Pik、Piz5、Pi9、Pish的单基因系在小种专化抗性上有别于广东省骨干亲本，这些基因在今后抗性育种中可重点引入。Pikh\[或 Pi1（t）\]、Pi9（或 Piz5）及Pish（或 Pita2）的基因聚合有利于提高广东省水稻品种对该地区稻瘟病菌的抗谱。并就稻瘟病抗性基因的研究及利用策略进行了讨论。

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... 杨健源等^[26]提出选择目标基因时, 用少数的抗性基因获得广谱抗性是抗稻瘟病育种中必须考虑的问题 ...