利用SSR定位籼稻品种Kaharamana中抗褐飞虱基因Bph9

褐飞虱是危害水稻生产最重要的害虫之一,利用寄主抗性被认为是防治褐飞虱最经济而有效的方法。斯里兰卡水稻品种Kaharamana对东亚和东南亚的褐飞虱种群均表现抗虫性,利用分子遗传学的方法对其携带的Bph9基因进行了SSR定位。所用的遗传群体为来源于Kaharamana和02428的含有180个单株的F2分离群体,每个F2单株套袋自交获得F2：3家系。利用苗期集团鉴定埘F2：3家系进行抗褐飞虱鉴定,以推测相应F2单株的基因型。连锁分析表明,Bph9位于第12染色体上的两个SSR标记RM463和RM5341之间,分别与之相距6．8cM和9．7cM。该标记有助十将Bph9用于分子标记辅助选择育种研究。     

水稻褐飞虱综合治理研究与示范——农业公益性行业专项“水稻褐飞虱综合防控技术研究”进展

针对我国褐飞虱Nilaparvata lugens(Stl)近年来严重为害及其对当家农药品种抗性急剧上升的现状,本项目在华中、华南、华东地区等代表性地区开展褐飞虱灾变规律、抗虫品种培育、抗药性监测及复配农药新剂型开发、生态治理新技术研究、预测预报技术、可持续治理技术集成与示范研究。结果表明:越南、老挝等境外虫源地及我国华南、长江中下游等地田间褐飞虱种群仍以2型为主,3型次之。田间小气候是褐飞虱逃避高温的关键因素,褐飞虱在上午气温升高时大量向温度较低的水稻基部20cm范围内转移以逃避高温,将高温天气造成的不利影响降至最低。不同栽插方式对褐飞虱发生量有明显影响,手栽稻田褐飞虱发生最重,机插稻次之,直播稻最轻。不同水稻品种、N肥施用水平对褐飞虱的发生有明显的影响:超级杂交稻褐飞虱虫口密度明显高于常规杂交稻,高N肥施用量促进褐飞虱的发生,且水稻品种与N肥施用量对褐飞虱发生量的影响有明显交互作用。抗药性监测结果表明:我国褐飞虱对吡虫啉有极高水平抗性(168.1～561.5倍),对噻嗪酮为低到中等水平抗性(4.2～33.1倍),对氟虫腈为敏感性降低到高水平抗性(2.7～67.7倍),对烯啶虫胺与毒死蜱为低水平抗性。筛选出噻虫嗪、吡蚜酮、唏啶虫胺和仲丁威4种高效低毒的防治单剂。研制出3种农药复配新制剂,其中1种新制剂已获得农业部药检所正式登记,且规模化生产,2种新制剂已进入农药登记程序。精细定位水稻抗褐飞虱基因Bph6、Bph7、Bph9、Bph15、BG1222,并找到了它们的近距离共分离分子标记。培育出高产、优质、熟期适宜、含有抗稻飞虱基因Bph14的水稻新品种广两优476。储备了一批聚合多抗稻飞虱基因的水稻亲本材料。研发出在田埂种植芝麻、大豆等天敌诱集植物,增加褐飞虱卵期寄生蜂和捕食性天敌蜘蛛的种群数量的轻简化生态调控技术;研发出对褐飞虱成、若虫取食及雌成虫产卵均有驱避作用的植物提取物混配剂3种;研发出显著提高稻虱缨小蜂对田间褐飞虱卵寄生率的引诱剂1种。建立了褐飞虱环境友好防控技术集成体系,并在湖南、湖北、广东、广西、浙江、江西6省区示范应用,取得了良好的经济、生态、社会效益。     

利用普通野生稻褐飞虱抗性改良水稻恢复系的研究

褐飞虱是我国水稻主要害虫之一,挖掘和利用稻飞虱抗性基因,选育抗稻飞虱的水稻品种是目前防治褐飞虱最科学、安全的一种措施。本研究利用具有高抗褐飞虱显性基因的抗源材料HS204作为抗源供体,以恢复系明恢65、582、MR等作为轮回亲本,通过回交,苗期群体鉴定和成株期农艺性状选择,成功地将高抗褐飞虱基因转育到轮回亲本中,获得45份抗性纯合株系,同时其配合力、米质等其他农艺性状也得到了相应改善。     

利用回交重组自交群体检测水稻抗褐飞虱数量性状基因座

利用由98个系组成的Nipponbare/Kasalath//Nipponbare回交重组自交系（Backcross Inbred lines,BILs）作图群体（BC1F9）,进行水稻抗褐飞虱数量性状基因座（Quantative Trait Locus,QTL）的检测和遗传效应分析。采用苗期集团鉴定方法,并以死苗率作为抗褐飞虱表型值,分析亲本和98个BILs抗褐飞虱表现。共检测到3个苗期抗褐飞虱QTL,分别位于2、10和12染色体上,各QTL的LOD值为2.01～2.33,贡献率为10.4%～16.6%,可解释群体总表型变异的39.0%。这3个数量性状基因座对褐飞虱的抗性均来自抗虫亲本Kasalath。与这些数量性状基因座连锁的分子标记可望应用于聚合多个抗性基因,培育对褐飞虱具有水平抗性水稻新品种的育种实践中。     

水稻OsLecRK1基因介导的抗褐飞虱功能分析

褐飞虱Nilaparvata lugens St(a)l是对水稻最具破坏性的害虫之一,OsLecRK1是水稻Bph3基因簇中对褐飞虱抗性贡献最大的基因.本文对RHTd(含Bph3)等材料进行了褐飞虱抗性评价,克隆并构建了OsLecRK1过量表达突变体水稻,利用该突变体分析了OsLecRK1基因对褐飞虱若虫存活率、若虫发育历期等生物学参数的影响.结果 表明,含Bph3基因水稻RHTd对褐飞虱的抗性明显地强于含Bph1基因水稻Mudgo和bph2基因水稻ASD7,RHTd水稻的褐飞虱受害指数仅为Mudgo和ASD7水稻的53.5％和24.1％.过量表达OsLecRK1基因能显著地增加水稻对褐飞虱的驱避性和抗生性,褐飞虱雌成虫偏好于在野生型水稻上产卵;突变体水稻上的褐飞虱若虫存活率显著地降低,仅为野生型水稻上若虫存活率的75.2％ ～81.8％,且若虫发育历期显著地延长,羽化率和初羽化雌成虫体重均显著地降低;此外,褐飞虱在突变体水稻上取食分泌的蜜露量只有野生型上的40.3％ ～ 60.9％,褐飞虱单雌产卵量只为野生型51％ ～61.2％,卵孵化率只有野生型的52.2％～56.7％,均显著地减少.结果 表明,含Bph3基因水稻RHTd对褐飞虱的抗性明显地高于分别含Bph1、bph2的水稻Mudgo和ASD7;水稻Bph3基因座的OsLecRK1单个基因过量表达即可显著增加水稻对褐飞虱的抗性,OsLecRK1协同影响褐飞虱的多个生物学参数降低褐飞虱的适合度.     

云南野生稻抗褐飞虱评价及其抗性基因鉴定

褐飞虱是水稻生产中最严重的害虫之一,从野生稻中发掘抗虫基因,有利于培育具有抗虫能力强的水稻新品种。该研究通过对云南野生稻进行温室和大田抗虫鉴定以及9个已知抗褐飞虱基因的PCR鉴定,发现云南野生稻对褐飞虱表现出不同程度的抗性,尤其疣粒野生稻和药用野生稻对褐飞虱表现出高抗,可作为抗虫基因发掘的优良抗源材料;不同褐飞虱抗性的云南野生稻中含有的抗褐飞虱基因差异很大,3种野生稻中均不含Bph1和Bph18(t)抗病基因,景洪普通野生稻和元江普通野生稻可能含bph2基因,东乡普通野生稻可能含bph2、Bph15和Bph27(t)基因,疣粒野生稻中可能含bph2和bph19(t)基因,药用野生稻和药用野生稻(宽叶型)中可能含bph2和Bph6基因,药用野生稻F1中可能含bph2、Bph14和bph20(t)基因,药用野生稻F2中可能含bph2和Bph27(t)基因或者其同源基因。该研究为快速发掘利用云南野生稻中的抗虫基因奠定了理论基础。     

水稻抗稻飞虱基因遗传与定位研究进展

褐飞虱(Nilaparvata lugens St(a)l)、白背飞虱(Sogatella furcifera Horvath)和灰飞虱(Laodelphax striatellus Fallén)是水稻生产上的重要害虫,给我国水稻生产造成了严重的经济损失.培育和利用抗虫品种是防治稻飞虱经济有效的措施.抗性遗传和抗性基因研究是进行抗虫育种的基础.目前,有关水稻抗褐飞虱基因的遗传与定位研究取得了较大进展,包括21个主基因、50余个QTLs和许多褐飞虱抗性相关基因被发掘、定位与克隆,而白背飞虱和灰飞虱抗性基因尚有待进一步发掘和鉴定.此外,今后应加强稻飞虱抗性基因在生产上的应用.     

基因聚合改良杂交稻组合的稻飞虱田间抗性表现

褐飞虱Nilaparvata lugens(Stl)是杂交水稻的重要虫害,培育高抗褐飞虱的杂交水稻新组合被认为是目前防治飞虱最有效、经济的方法。本研究以分子标记辅助选择培育的抗褐飞虱杂交水稻为材料,通过苗期和成株期的全生育期田间抗性鉴定,对基因聚合改良杂交稻组合的褐飞虱抗性进行综合评价,结果表明,基因聚合改良杂交稻组合的苗期抗性介于抗和中抗水平,田间虫量在7次调查中均低于800头/百丛,远低于2000头/百丛的防治标准,而对照扬两优6号、两优培九和当地农家品种两优63在8月29日到9月10日飞虱发生高峰期的百丛虫量分别最高达到5400头、2030头和10700头,高于或远高于2000头/百丛的防治标准。改良组合川香29A/华恢1462的产量在通城比对照扬两优6号增产11.8%。上述结果表明抗褐飞虱改良杂交稻新组合能够有效抑制稻田褐飞虱种群数量,节约人工和农药成本,增加水稻产量。     

抗褐飞虱基因的发掘、鉴定与利用

水稻是我国最重要的粮食作物,而褐飞虱是水稻生产中的主要害虫,培育与利用抗褐飞虱的水稻品种是综合防治褐飞虱的基础。现从水稻抗褐飞虱种质资源、水稻抗褐飞虱基因的定位与克隆、水稻抗褐飞虱的分子机理,以及抗褐飞虱水稻的培育等方面,综述水稻抗褐飞虱基因研究的最新成果。     

十六个水稻品种(系)对褐飞虱的抗虫性评价

本文通过苗期抗性鉴定、田间抗虫性鉴定、稻田节肢动物功能团优势度比较及产量测定的方法来评价16个水稻品种(系)对褐飞虱Nilaparvata lugens(Stl)的抗性。结果表明,有12个品种(系)对褐飞虱表现为抗级,2个品系表现为中抗。这14个抗虫-中抗的品种(系)在成株期也表现一定的抗虫性,且与对照TN1之间差异显著。本试验条件下,广占63-3S/华恢15、广两优106、Y58S/华15、广两优476、广占63-4S/华恢15这5个品种(系)不仅抗虫性好,而且产量也较高,分别比TN1和主栽品种扬两优6号高63.49%～66.57%和3.47%～11.57%,具有广泛的开发应用前景。     

High-resolution mapping of a new brown planthopper (BPH) resistance gene, Bph18(t), and marker-assisted selection for BPH resistance in rice (Oryza sativa L.)

Brown planthopper (BPH) is a destructive insect pest of rice in Asia. Identification and the incorporation of new BPH resistance genes into modern rice cultivars are important breeding strategies to control the damage caused by new biotypes of BPH. In this study, a major resistance gene, Bph18(t), has been identified in an introgression line (IR65482-7-216-1-2) that has inherited the gene from the wild species Oryza australiensis. Genetic analysis revealed the dominant nature of the Bph18(t) gene and identified it as non-allelic to another gene, Bph10 that was earlier introgressed from O. australiensis. After linkage analysis using MapMaker followed by single-locus ANOVA on quantitatively expressed resistance levels of the progenies from an F₂ mapping population identified with marker allele types, the Bph18(t) gene was initially located on the subterminal region of the long arm of chromosome 12 flanked by the SSR marker RM463 and the STS marker S15552. The corresponding physical region was identified in the Nipponbare genome pseudomolecule 3 through electronic chromosome landing (e-landing), in which 15 BAC clones covered 1.612 Mb. Eleven DNA markers tagging the BAC clones were used to construct a high-resolution genetic map of the target region. The Bph18(t) locus was further localized within a 0.843-Mb physical interval that includes three BAC clones between the markers R10289S and RM6869 by means of single-locus ANOVA of resistance levels of mapping population and marker-gene association analysis on 86 susceptible F₂ progenies based on six time-point phenotyping. Using gene annotation information of TIGR, a putative resistance gene was identified in the BAC clone OSJNBa0028L05 and the sequence information was used to generate STS marker 7312.T4A. The marker allele of 1,078 bp completely co-segregated with the BPH resistance phenotype. STS marker 7312.T4A was validated using BC₂F₂ progenies derived from two temperate japonica backgrounds. Some 97 resistant BC₂F₂ individuals out of 433 screened completely co-segregated with the resistance-specific marker allele (1,078 bp) in either homozygous or heterozygous state. This further confirmed a major gene-controlled resistance to the BPH biotype of Korea. Identification of Bph18(t) enlarges the BPH resistance gene pool to help develop improved rice cultivars, and the PCR marker (7312.T4A) for the Bph18(t) gene should be readily applicable for marker-assisted selection (MAS). K. K. Jena and J. U. Jeung contributed equally to this study.     

Pyramiding and evaluation of the brown planthopper resistance genes Bph14 and Bph15 in hybrid rice

The brown planthopper (BPH) is the most devastating insect pest in rice-producing areas. Shanyou 63 has become a widely cultivated hybrid in China over the last two decades; however, this line has become increasingly susceptible to bacterial blight (BB), blast, and BPH, resulting in a rapid decline in its use in rice production. In this study, a molecular marker-assisted selection (MAS) introgression of Bph14 and Bph15 was performed to improve the BPH resistance of Minghui 63 and its derived hybrids such as Shanyou 63. The effect of pyramiding genes was then comprehensively evaluated using three tests that comprised seedbox screening, feeding rate, and antixenosis for settling in the field. The results showed that the improved hybrids containing a single BPH resistance gene showed enhanced resistance (lower resistance score, honeydew weight and number of BPH settling) compared to conventional hybrids, while pyramiding two genes provided even higher resistance. Moreover, both Bph14 and Bph15 are partial dominance genes, and have a strong dosage effect on the resistance to BPH in the hybrid background, which is useful for breeding BPH-resistant hybrids. Field trial data demonstrated that yields of improved hybrid rice were higher than or similar to the control (Shanyou 63) under natural field conditions. These improved versions could be used in breeding programs for “green super rice.”     

Fine mapping of the rice Bph1 gene, which confers resistance to the brown planthopper (Nilaparvata lugens stal), and development of STS markers for marker-assisted selection

The brown planthopper (BPH) is a major insect pest in rice, and damages these plants by sucking phloem-sap and transmitting viral diseases. Many BPH resistance genes have been identified in indica varieties and wild rice accessions, but none has yet been cloned. In the present study we report fine mapping of the region containing the Bph1 locus, which enabled us to perform marker-aided selection (MAS). We used 273 F8 recombinant inbred lines (RILs) derived from a cross between Cheongcheongbyeo, an indica type variety harboring Bph1 from Mudgo, and Hwayeongbyeo, a BPH susceptible japonica variety. By random amplification of polymorphic DNA (RAPD) analysis using 656 random 10-mer primers, three RAPD markers (OPH09, OPA10 and OPA15) linked to Bph1 were identified and converted to SCAR (sequence characterized amplified region) markers. These markers were found to be contained in two BAC clones derived from chromosome 12: OPH09 on OSJNBa0011B18, and both OPA10 and OPA15 on OSJNBa0040E10. By sequence analysis of ten additional BAC clones evenly distributed between OSJNBa0011B18 and OSJNBa0040E10, we developed 15 STS markers. Of these, pBPH4 and pBPH14 flanked Bph1 at distances of 0.2 cM and 0.8 cM, respectively. The STS markers pBPH9, pBPH19, pBPH20, and pBPH21 co-segregated with Bph1. These markers were shown to be very useful for marker-assisted selection (MAS) in breeding populations of 32 F6 RILs from a cross between Andabyeo and IR71190, and 32 F5 RILs from a cross between Andabyeo and Suwon452.     

Mapping of a broad-spectrum brown planthopper resistance gene, <Emphasis Type="Italic">Bph3</Emphasis>, on rice chromosome 6

The brown planthopper (BPH) is one of the most destructive insect pests of rice in Thailand. We performed a cluster analysis that revealed the existence of four groups corresponding to the variation of virulence against BPH resistance genes in 45 BPH populations collected in Thailand. Rice cultivars Rathu Heenati and PTB33, which carry Bph3, showed a broad-spectrum resistance against all BPH populations used in this study. The resistant gene Bph3 has been extensively studied and used in rice breeding programs against BPH; however, the chromosomal location of Bph3 in the rice genome has not yet been determined. In this study, a simple sequence repeat (SSR) analysis was performed to identify and localize the Bph3 gene derived from cvs. Rathu Heenati and PTB33. For mapping of the Bph3 locus, we developed two backcross populations, BC₁F₂ and BC₃F₂, from crosses of PTB33 × RD6 and Rathu Heenati × KDML105, respectively, and evaluated these for BPH resistance. Thirty-six polymorphic SSR markers on chromosomes 4, 6 and 10 were used to survey 15 resistant (R) and 15 susceptible (S) individuals from the backcross populations. One SSR marker, RM190, on chromosome 6 was associated with resistance and susceptibility in both backcross populations. Additional SSR markers surrounding the RM190 locus were also examined to define the location of Bph3. Based on the linkage analysis of 208 BC₁F₂ and 333 BC₃F₂ individuals, we were able to map the Bph3 locus between two flanking SSR markers, RM589 and RM588, on the short arm of chromosome 6 within 0.9 and 1.4 cM, respectively. This study confirms both the location of Bph3 and the allelic relationship between Bph3 and bph4 on chromosome 6 that have been previously reported. The tightly linked SSR markers will facilitate marker-assisted gene pyramiding and provide the basis for map-based cloning of the resistant gene.