磷转运蛋白基因TaPHT2;1在染色体上定位 及对小麦磷素吸收和利用效率的影响

【目的】以中国春遗传背景的整套B染色体双端体为材料,鉴定磷转运蛋白基因TaPHT2;1的染色体定位特征。解析不同供磷水平下,上述材料和不同磷利用效率小麦品种该基因的表达特征及其与植株干物质生产能力和磷效率特征的联系。【方法】采用溶液培养法水培中国春（CS）及该品种遗传背景的整套B染色体组双端体和不同磷效率小麦品种材料。以B染色体组供试端体为材料进行TaPHT2;1 PCR扩增,鉴定TaPHT2;1在染色体上的定位。采用半定量RT-PCR及qRT-PCR技术分析B染色体组供试端体和小麦品种TaPHT2;1的表达水平。采用常规分析技术,测定供试材料单株干重和磷吸收参数。【结果】①PCR检测发现,在CS及所有供试B染色体组双端体材料中,除缺失1B长臂的1BS外,其它所有材料均能特异扩增出目标基因TaPHT2;1,表明TaPHT2;1定位在1B长臂。②丰、缺磷条件下,TaPHT2;1在CS及除1BS外双端体根、叶中的表达均表现为叶片优势表达特征,且在叶片中表达受到低磷胁迫的诱导。TaPHT2;1在根系中的表达不受低磷逆境调控。表明TaPHT2;1在介导丰磷下磷素吸收、转运及增强低磷下植株体内磷素再度调运中可能发挥重要功能。③丰磷条件下,与CS相比,1BS的单株干重和全磷含量显著降低;缺磷条件下,1BS的单株干重与CS相比也显著下降,但全磷含量增加。表明位于1B染色体长臂后的磷转运蛋白基因TaPHT2;1,对丰、缺磷条件下的植株磷素吸收、转运具有较大影响,进一步对不同供磷水平下的植株干重产生重要调控效应。④丰磷条件下,与CS相比,1BS的单株磷累积量显著增加,磷利用效率没有改变;缺磷条件下,与CS相比,1BS的单株磷累积量没有变化,磷利用效率显著降低。不同磷利用效率品种相比,丰磷条件下,随着品种磷利用效率提高,叶片中TaPHT2;1的表达水平、单株干重、全磷含量和单株磷累积量也随着增加;缺磷条件下,随着小麦品种磷利用效率提高,叶片中TaPHT2;1的表达水平、单株干重和磷利用效率也随之增高,但全磷含量呈下降趋势,单株磷累积量品种间差异较小。因此,TaPHT2;1的表达水平与小麦品种丰、缺磷条件下的磷素吸收、利用和干物质积累能力具有紧密联系。【结论】小麦磷转运蛋白基因TaPHT2;1位于1B染色体长臂。该基因通过其特定对外界供磷水平产生应答,在较大程度上调控植株的磷素吸收和利用能力,对不同供磷水平下的植株干重产生重要影响。TaPHT2;1在调控植株丰磷下磷素吸收和低磷下磷素利用中发挥重要作用,可作为鉴定小麦品种磷效率的评价指标。     

Expression and functional analyses of the mitogen-activated protein kinase(MPK) cascade genes in response to phytohormones in wheat (Triticum aestivum L.)

Mitogen-activated protein kinase(MPK) cascades consist of a set of kinase types(MPKKKs,MPKKs,MPKs) to establish conserved signal-transducing modules mediating plant growth,development as well as responses to internal and external cues.In this study,the expression patterns of six MPKKK,two MPKK,and 11 MPK genes in wheat in responses to external treatments of phytohormones,including naphthylacetic acid(NAA),abscisic acid(ABA),6-benzyladenine(6-BA),gibberellin(GA_3),salisylic acid(SA),jasmonic acid(JA),and ethylene(ETH),were investigated.Expression analysis revealed that several of the MPK cascade genes are responses to the external phytohormone signaling.Of which,TaMPKKKA;3is induced by 6-BA and NAA while TaMPK4 repressed by ETH,GA_3,SA,and JA;TaMPKKKA,TaMPKKKA;3 and TaMPK1 are down-regulated by ETH and GA_3whereas TaMPK9 and TaMPK12 repressed by ETH and JA in addition that TaMPK12 also repressed by GA_3;TaMPK12;1 is down-regulated by ABA,GA_3 and SA while TaMPK17 repressed by all exogenous phytonormones examined.TaMPK4,a MPK type gene previously characterized to mediate tolerance to phosphate(Pi)deprivation,was functionally evaluated for its role in mediation of responses of plants to exogenous GA_3,ETH,SA,and JA.Results indicated that overexpression and antisense expression of TaMPK4 in tobacco dramatically modify the growth of seedlings upon treatments of GA_3,SA and JA,in which the overexpressors behaved deteriorated growth feature whereas the seedlings with antisense expression of TaMPK4 exhibited improved seedling phenotype.The growth behaviors in lines overexpressing or antisensely expressing TaMPK4 are closely associated with the biomass and the corresponding hormone-associated parameters.These results demonstrated that TaMPK4 acts as a critical player in mediating the phytohormone signaling.Our findings have identified the phytohormone-responsive MPK cascade genes in wheat and provided a connection between the phytohormone-mediated responses and the MPK cascade pathways.     

植物低磷胁迫信号转导与适应性反应研究进展

磷是植物生长发育所必须的大量元素之一,据调查世界范围内大量耕地有效磷缺乏,植物生长发育和作物产量因此受到限制。在长期进化过程中,耐低磷植物在早期响应基因(主要与低磷信号感受和转导有关)和后期响应基因(主要与形态、生理生化和代谢有关)的协同作用下形成了多种低磷适应机制。从低磷信号的感受和转导,形态、生理生化和代谢适应性机制方面综述了目前植物耐低磷方面的研究进展。以期为植物低磷信号转导和适应机制研究及耐低磷作物品种的培育提供依据。     

植物低磷胁迫信号转导与适应性反应研究进展

磷是植物生长发育所必须的大量元素之一,据调查世界范围内大量耕地有效磷缺乏,植物生长发育和作物产量因此受到限制。在长期进化过程中,耐低磷植物在早期响应基因（主要与低磷信号感受和转导有关）和后期响应基因（主要与形态、生理生化和代谢有关）的协同作用下形成了多种低磷适应机制。从低磷信号的感受和转导,形态、生理生化和代谢适应性机制方面综述了目前植物耐低磷方面的研究进展。以期为植物低磷信号转导和适应机制研究及耐低磷作物品种的培育提供依据。     

响应低磷胁迫的小麦MDP激酶基因TaMPK1a-1的克隆和表达分析

 【目的】蛋白磷酸化在介导非生物逆境信号的转导中具有重要作用。以笔者在低磷胁迫下鉴定的小麦促分裂原活化蛋白激酶（MAP激酶）基因TaMPK1a-1为基础,开展该基因应答低磷逆境分子特征的研究。【方法】利用cDNA-AFLP技术,鉴定特异上调表达的MAP激酶基因TaMPK1a-1。采用生物信息学技术研究基因结构和编码蛋白特征,采用半定量RT-PCR技术研究TaMPK1a-1应答低磷胁迫逆境的分子特征。【结果】TaMPK1a-1 cDNA长度为2 170 bp,开放阅读框为1 737 bp,编码578个氨基酸残基。TaMPK1a-1含有2个参与双重磷酸化作用的TEY和TDY基序。在正常供磷条件下,磷高效品种石新828和磷低效品种冀7369根叶中均检测不到TaMPK1a-1的转录本;低磷处理下,TaMPK1a-1的表达在上述品种的根叶中均受到明显诱导。与冀7369相比,低磷条件下石新828根叶中TaMPK1a-1的转录本明显增多。【结论】TaMPK1a-1级联转导途径不仅影响着小麦对低磷信号的响应,而且对于增强小麦适应低磷胁迫的能力中也可能具有重要作用。     

磷胁迫下羊草的响应及磷响应相关基因表达分析

【目的】磷是植物生长发育所必需的大量营养元素,研究无机磷酸盐（inorganic phosphate,Pi）胁迫下羊草的响应,筛选不同浓度Pi胁迫下的内参基因,并分析Pi响应相关基因的表达,为羊草Pi胁迫响应的分子机制研究提供基础数据。【方法】以羊草幼苗为材料,进行不同浓度Pi处理培养,对羊草的株高和根长进行检测,利用钒钼黄比色法检测羊草植株中Pi的含量。根据羊草转录组数据选取7个候选内参基因,从NCBI核酸序列数据库选取1个候选内参基因,对羊草不同处理材料进行总RNA提取和cDNA合成,利用qRT-PCR对候选内参基因的表达进行检测,并通过geNorm、NormFinder和Bestkeeper软件对其稳定性进行评估。根据筛选获得的较稳定内参基因,对Pi响应基因的qRT-PCR检测数据进行相对定量分析。【结果】表型观测结果显示,无论是低Pi还是高Pi胁迫都使得羊草的生长减缓;株高对低Pi或缺Pi胁迫较为敏感,根长对高Pi胁迫较为敏感;并随着处理浓度的增加羊草中Pi的含量也增加。qRT-PCR溶解曲线分析显示8个候选内参基因均具有单一的溶解峰,基因表达谱分析表明8个候选内参基因的CT值范围是17.16—26.61,其中,LcGAPDH的表达丰度最高为17.16—20.22,Lc18SrRNA的表达丰度最低为23.28—26.61,CT值变异系数最小的为LcARPT（2.09%）,最大的为LcTUA（6.8%）。综合geNorm、NormFinder和Bestkeeper稳定性排名,通过计算几何平均数,获得8个候选内参基因综合稳定性排名,其中排名靠前的3个基因分别为Lc18SrRNA、LcCAP和LcEF1α,排名最后的2个基因为LcTUA和LcTUB。分别以Lc18SrRNA、LcCAP和LcEF1α作为内参基因,qRT-PCR相对定量表达分析结果显示,与对照相比,LcPHO1-2的表达受低Pi或者缺Pi诱导;LcPAP2的表达受高Pi诱导;而LcPAP27的表达同时受低Pi和高Pi下调。【结论】低Pi和高Pi胁迫均使羊草生长受阻,羊草地上组织与地下组织对Pi胁迫响应的模式不同。筛选出3个表达较为稳定的内参基因Lc18SrRNA、LcCAP和LcEF1α。LcPHO1-2和LcPAP2分别参与了羊草对低Pi和高Pi的应答响应,LcPAP27同时参与了羊草对低Pi和高Pi的响应进程。     

湖南稻瘟病菌生理小种的组成及其致病性

利用来自日本、国际水稻研究所(IRRI)和中国的10个稻瘟病菌单基因鉴别品种,将59个湖南稻瘟病菌株分别划分为20个生理小种,优势小种为H531.1、H511.1和H501.1,对抗性基因Pi–CO39、Pita、Pi3、Pi12、Pia具有致病性。各个稻区生理小种组成和优势小种存在差异,其中,湘东、湘中稻区稻瘟病菌生理小种多样性指数较高,优势小种的变化最为明显,说明这2个稻区的抗源基因正逐渐变得匮乏,甚至丧失。鉴别品种C101LAC(Pi1)和C101A51(Pi2)对59个湖南稻瘟病菌菌株抗性频率均为100.0%,说明这些抗性品种(基因)可作为湖南稻区稻瘟病的抗源材料(基因),Pi1、Pi2可直接作为分子标记辅助选择育种的供体抗源基因。品种关东51(Pik)、C105TTP–4L23(Pi4–b)和C101PKT(Pita)在湖南各稻区的抗性水平表现存在差异,可以作为选择性抗源材料(基因)加以应用。     

Postulation of seedling leaf rust resistance genes in 84 Chinese winter wheat cultivars

Wheat leaf rust(caused by Puccinia triticina) is one of the most important fungal diseases in China. There are tens of winter wheat cultivars which are approved to be released by the government at a national level and more than 100 wheat cultivars at the provincial level. But there is no information about leaf rust(Lr) genes in these cultivars, which makes it difficult for farmers and breeders to select which cultivars they should plant in their fields and use in their breeding programs. The objective of this paper was to identify the leaf rust resistant genes at seedling stage present in the 84 commercial wheat cultivars from China that have been released in the past few years. A set of 20 near isogenic lines with Thatcher background and 6 lines with known Lr genes were used to test the virulence of 12 races of P. triticina(Pt). By comparing the infection types(ITs) produced on the 84 cultivars by the 12 Pt races with the ITs on the differential sets, the Lr genes were postulated. In addition, 8 molecular markers of Lr genes such as Lr9, Lr10, Lr19, Lr20, Lr21, Lr24, Lr26 and Lr29, which are closely linked to or co-segregated with the Lr gene, were used for further validation of the genes in the 84 Chinese winter wheat cultivars. Twelve Lr genes, including Lr1, Lr3,(Lr3bg),(Lr3ka), Lr11, Lr13, Lr14 a, Lr16, Lr26, Lr27, Lr30 and Lr31 were postulated to be present either singly or in combinations in these Chinese wheat cultivars. Lr3 and Lr26 were detected most often in the tested cultivars, with frequencies of 51.2 and 38.1%, respectively. No wheat Lr genes were detected in 16 cultivars, and 4 cultivars may carry unknown Lr genes other than those used in this study. Lr9, Lr20, Lr21, Lr24, Lr25 and Lr29 were not present in any of the 84 tested accessions.     

不同基因型小麦在水培体系下的磷胁迫反应研究

采用改进的Hoagland培养基,优化建立了小麦温室水培体系,对不同基因型小麦在水培条件下对磷饥饿胁迫的生理反应进行了研究.结果表明:1)在研究建立的小麦水培体系中,小麦植株生长正常并对磷饥饿胁迫条件做出发育形态建成水平的反应;半定量RT-PCR分析显示在此体系中小麦磷响应基因也在转录水平上产生显著变化;2)在水培条件下不同基因型小麦对磷饥饿胁迫的反应总体上表现在根轴长度和根冠比有所提高,但不同基因型小麦的反应特点有差异;对这些反应特征的分子机理的深入分析可为有效筛选磷高效小麦新种质以及发掘磷高效基因资源提供线索.     

济麦22及其转TaPHR1基因植株的磷胁迫反应研究

采用营养液培养体系,研究不同供磷处理条件下,受体济麦22及其转磷高效基因TaPHR1株系的生长反应、根系形态特征及对磷的吸收、分配和利用能力的差异。结果表明:1)通过农杆菌介导方法获得的TaPHR1基因过量表达的转基因植株后代有更强的耐低磷胁迫的能力;2)在水培条件下,小麦对磷饥饿胁迫的反应总体上表现在生物量降低、根冠比提高、根长及表面积增大、根半径减小,磷吸收和利用效率改变,不同基因型间变化趋势相同,但变化幅度存在差异;3)对这些反应特征的分子机理的深入分析可为有效筛选磷高效小麦新种质以及发掘磷高效基因资源提供参考依据。     

冬小麦膨胀素基因TaEXPB8、TaEXPB10的克隆及低温表达分析

以冬小麦强抗寒品种东农冬麦1号为试验材料,从其根中克隆膨胀素基因TaEXPB8、TaEXPB10,并分析其在低温条件下的表达情况,为研究膨胀素基因与植物抗寒性之间的关系奠定基础。结果表明,在冬小麦强抗寒品种东农冬麦1号根中克隆得到膨胀素基因TaEXPB8、TaEXPB10,二者分别编码273、271个氨基酸,二者序列与Gen Bank上公布的序列同源性分别达到99.2%、99.6%。4℃、-10℃和-20℃的低温胁迫处理均可诱导TaEXPB8、TaEXPB10基因在冬小麦强抗寒品种东农冬麦1号根中表达,尤其是TaEXPB10基因,其受诱导程度高于TaEXPB8;两基因在低抗寒品种济麦22号中总体上不受诱导,甚至受到抑制。综合比较发现,4℃、-10℃和-20℃低温胁迫下,TaEXPB8、TaEXPB10基因在强抗寒品种东农冬麦1号中的表达量均明显高于低抗寒品种济麦22号。据此推测膨胀素基因TaEXPB8、TaEXPB10与冬小麦根系的低温胁迫耐受能力具有重要的相关性,有助于进一步深入了解冬小麦根系的抗寒机制。     

宁夏春小麦矮秆基因分子标记检测分析

[目的]分析宁夏春小麦育种材料常用矮秆基因的分布,为本地小麦改良株高性状提供理论依据。[方法]以宁夏大学小麦遗传育种实验室212个(其中54个材料由中国农业科学院引进)春小麦育种亲本为材料,调查其株高、穗长、穗下茎节等农艺性状,利用分子标记检测分析材料中常用矮秆基因Rht-B1b(Rht1)、Rht-D1b(Rht2)和Rht8的分布。[结果]中国农业科学院引进材料的株高、穗长、穗茎节的平均值(分别为69.2、7.8、28.5 cm)明显低于现有亲本材料(分别为83.6、11.1、32.0 cm)。矮秆基因检测发现166份材料携带Rht-B1b基因,占总检测材料的75.1%;88份材料含有Rht-D1b基因,占39.8%;191份材料含Rht8基因,占总检测材料的86.4%。携带Rht-D1b基因的9个材料同时含有Rht-B1b和Rht8基因。其中,引进材料中Rht8、Rht-B1b、Rht-D1b分别占85.1%、72.0%、33.3%,与整体材料中各矮秆基因分布相似。[结论]目前宁夏春小麦育种材料中普遍含有矮秆基因Rht8,其次是Rht-B1b,但主栽品种和主推品种以及核心育种亲本材料均含有Rht-D1b,即现有亲本材料中含Rht8+Rht-B1b的材料所占比例较Rht8+Rht-D1b多,但新培育的品种以含有Rht8+Rht-D1b为主。这可能是主栽品种宁春4号(Rht8+Rht-D1b)在当地长期种植,并作为育种骨干亲本被长期利用的结果。     

Breeding wheat for drought tolerance: Progress and technologies

Recurrent drought associated with climate change is among the principal constraints to global productivity of wheat(Triticum aestivum(L.) and T. turgidum(L.)). Numerous efforts to mitigate drought through breeding resilient varieties are underway across the world. Progress is, however, hampered because drought tolerance is a complex trait that is controlled by many genes and its full expression is affected by the environment. Furthermore, wheat has a structurally intricate and large genome. Consequently, breeding for drought tolerance requires the integration of various knowledge systems and methodologies from multiple disciplines in plant sciences. This review summarizes the progress made in dry land wheat improvement, advances in knowledge, complementary methodologies, and perspectives towards breeding for drought tolerance in the crop to create a coherent overview. Phenotypic, biochemical and genomics-assisted selection methodologies are discussed as leading research components used to exploit genetic variation. Advances in phenomic and genomic technologies are highlighted as options to circumvent existing bottlenecks in phenotypic and genomic selection, and gene transfer. The prospects of further integration of these technologies with other omics technologies are also provided.