金针菇菌柄发育的转录组与蛋白组分析

菌柄是金针菇等食用菌的主要商品部位,但其生长机制仍不明确。本研究对金针菇伸长期和成熟期菌柄进行了转录组联合蛋白组分析,结果显示,两样本显著性差异表达基因和蛋白分别为721个和61个,均以上调表达为主。GO（gene ontology）功能聚类分析表明：有72.41%的差异表达基因富集在催化活性（catalytic activity）条目下。细胞组分（cell part）和绑定结合（binding）条目同时富集了较多的差异表达基因和蛋白。KEGG通路富集分析显示：碳水化合物代谢通路（carbohydrate metabolism）和氨基酸代谢通路（amino acid metabolism）富集的差异表达基因较多。差异表达蛋白富集较多的通路是单环菌素生物合成（monobactam biosynthesis,ko00261）、链霉素生物合成（streptomycin biosynthesis,ko00521）和有机含硒化合物代谢（selenocompound metabolism,ko00450）等。内质网蛋白质加工（protein processing in endoplasmic reticulum,ko04141）和MAPK信号通路（MAPK signaling pathway-yeast,ko04011）在转录组和蛋白组的KEGG富集分析中均为差异通路。本研究联合转录组和蛋白组数据筛选了40个金针菇菌柄发育中差异表达基因,为深入研究揭示食用菌菌柄发育过程提供候选基因。     

一个玉米矮秆突变体K123d的遗传鉴定

矮秆已被广泛用于改良作物的抗倒伏性状,培育理想株型,从而提高作物产量。玉米矮秆突变体K123d由自交系K123自然突变产生。本研究比较该突变体与野生型主要农艺性状差异及其对赤霉素的敏感性;用K123d与株高不同的3个自交系分别构建F1、BC和F2群体,分析矮秆性状的遗传模式;以K169/K123d-F2为定位群体,采用集团分离分析法(BSA),运用SSR标记定位矮秆基因d123;参照br-2序列信息分段设计特异引物,同源克隆d123。结果表明,与野生型相比K123d株高降低35.59%,穗位高降低、节间缩短、叶片较直立,但结实率差,对赤霉素敏感;在F2群体和BC1群体中,正常植株与矮秆植株分离比例分别符合3∶1和1∶1,说明矮秆性状受1对隐性基因控制;其矮秆基因d123定位于第一条染色体上SSR标记umc1278和bnlg1564之间,遗传距离分别为12.8 c M和7.3 c M;同源克隆显示d123与br-2存在12个碱基替换,其中第4个外显子编码的一个谷氨酸被替换为赖氨酸。由此可见,矮秆突变体K123d为br-2的一个突变类型,对矮化育种具有进一步研究利用价值。     

蒺藜苜蓿株高控制基因COSTIN的克隆及功能研究

株高是影响植物株型建成的重要农艺性状之一,直接决定作物的倒伏性和生物产量,但目前关于苜蓿等豆科牧草株高性状形成的分子调控机制尚不清楚。通过定向筛选豆科模式植物蒺藜苜蓿Tnt1逆转座子插入突变体库,分离鉴定了一个蒺藜苜蓿矮化突变体compact stalk internodes(costin),该突变体的矮化表型是由于茎节伸长受到抑制所致。通过基因表型连锁分析成功克隆了COSTIN基因,该基因编码一个钙离子交换蛋白,与拟南芥的CALCIUM EXCHANGER 7(CAX7) 基因高度同源。qRT-PCR检测发现COSTIN基因在茎、叶和果荚等组织中有较高的表达。进一步研究发现在costin突变体中赤霉素合成途径关键基因MtCPS、MtKAO1、MtGA20ox4、MtGA20ox7和MtGA3ox1表达下调;外施赤霉素GA3可以恢复costin突变体的矮化表型。上述研究表明COSTIN基因通过影响植物激素赤霉素的生物合成来调控蒺藜苜蓿的茎节伸长。     

陆地棉矮化突变体Ari1327茎尖的转录组分析

为了从分子水平上研究陆地棉矮秆突变体Ari1327的矮化机理,本研究以矮秆突变体Ari1327、野生型Ari971和高秆突变体Ari3697的茎尖为材料,建立3个cDNA文库,用Illumina HiSeqTM2000系统对3个材料的茎尖cDNA进行转录组测序。3个文库测序共得4.9 G数据量,拼接得到Unigene 70877个。通过矮化突变体Ari1327与野生型Ari971和高秆突变体Ari3697两个文库的差异筛选,得到13919个与矮化相关的差异表达基因,其中5406个表现上调,8513个表现下调。GO功能和KEGG通路富集发现,差异基因在植物激素信号转导途径显著富集。通过实时荧光定量PCR(qRT-PCR)验证,推测Ari1327的矮化可能与赤霉素和生长素2种激素的信号转导及互作有关。转录组测序得到的大量差异基因,为深入研究棉花的矮化机理具有重要参考价值,同时为棉花的矮化育种工作奠定了基础。     

转基因金叶银中杨叶色及生长变异分析

转PaGLK基因银中杨抑制表达株系叶绿素含量显著降低,叶片呈现黄色（命名为金叶银中杨）,以转PaGLK基因的银中杨为材料,测定其叶色参数和叶绿素含量的时序变化规律、分析生长特性。结果显示,转PaGLK基因的银中杨使叶片颜色发生改变,抑制表达株系整个生长期叶绿素含量显著低于WT（P<0.05）,叶色亮度显著高于WT（P<0.05）,并且在生长发育期叶片一直呈现深黄绿色。抑制表达株系中的Y2速生期内苗高日生长量（GD）高于对照株系,苗期株高不受影响。而过表达转基因银中杨的当年高生长都显著低于对照株系 （P<0.05）,其速生期内苗高日生长量均值（GD）也低于对照株系,其均值为对照株系的22.19%。PaGLK抑制表达株系在城市园林绿化具有潜在的应用价值。     

小麦主胚根生长及主要性状全基因组关联分析

为解析小麦初生根系建成的遗传机制,本研究以黄淮麦区的198份小麦自然群体为材料,对在室内人工气候箱内水培21 d的小麦主胚根的一级分枝根数、分枝密度、长度、表面积、体积和平均直径6个性状进行调查分析,结合660K基因芯片用Q+K混合线性模型对主胚根性状进行全基因组关联分析,并对显著且稳定的关联位点进行功能注释和候选基因挖掘。结果表明,主胚根不同性状呈正态或近似正态分布,变异系数为5.56%～22.10%。通过全基因组关联分析,共检测到136个显著关联位点,这些位点分布在除7B以外的染色体上,可解释5.10%～13.60%的表型变异,同时检测到13个显著的多效位点,挖掘到TraesCS4A01G023100、TraesCS1B01G294400、TraesCS4A01G006200等16个可能与主胚根生长相关的候选基因,这些基因可能通过调控DNA拓扑结构异构酶、泛素结合酶E2、磷酸肌醇磷酸酶家族蛋白等参与小麦主胚根系的建成。本研究结果为小麦根系调控网络构建,以及优化根系构型和发挥根系功能提供了参考。     

毛竹茎秆伸长过程中赤霉素生物合成、降解和信号转导关键基因的鉴定及表达分析

赤霉素(Gibberellin)是一类非常重要的植物激素,在高等植物生命活动的整个周期都起着重要的调控作用。从毛竹Phyllostachys edulis基因组中共鉴定出23个赤霉素途径基因,包括赤霉素生物合成相关的8个GA20ox和1个GA3ox基因、降解相关的8个GA2ox基因、参与赤霉素感知的2个GID1基因以及信号转导的2个GID2基因和2个DELLA基因。拟南芥、水稻和毛竹的系统进化树和保守基序分析显示赤霉素的合成代谢与信号转导在这些物种中是高度保守的。利用外源赤霉素处理毛竹种子和幼苗,发现赤霉素能显著提高种子的萌发率和幼苗的茎秆伸长,并且有着最佳的作用浓度。在GA3处理后,毛竹体内赤霉素生物合成基因GA20ox和GA3ox表达量均下调而降解活性赤霉素的GA2ox基因表达量上调;赤霉素受体GID1和正调控基因GID2的转录水平显著提高而负调控基因DELLA的表达受到抑制。这些基因在竹笋茎秆的不同形态学位置表达差异明显,大部分赤霉素生物合成与降解的相关基因GA20ox、GA3ox和GA2ox以及赤霉素受体GID1和正调控基因GID2都在竹笋的形态学上端大量表达,而赤霉素信号转导的阻遏基因DELLA在笋体形态学底端大量积累而顶端基本不表达。     

莱茵衣藻丝/苏氨酸蛋白激酶突变株蓝光响应的表征

为研究莱茵衣藻丝/苏氨酸蛋白激酶(silk/threonine protein kinase, STK)介导藻细胞蓝光响应的分子机制，本文对蓝光胁迫下莱茵衣藻STK突变株系crstk11(AphvIII盒反向插入stk11基因编码区)进行表型鉴定及转录组分析。表型鉴定显示，正常光(白光)下，野生型株CC5325与突变株crstk11的生长和色素含量差异较小；蓝光抑制了crstk11藻细胞生长和叶绿素合成，但显著促进类胡萝卜素积累。转录组分析显示，蓝光处理4 d，突变株(STK4) vs.野生型(wild type, WT4)共检测到差异表达基因(differential expression genes, DEGs) 860条(559个上调，301个下调)。高蓝光处理8 d,STK8 vs. WT8共获得1 088个DEGs (468个上调，620个下调)。KEGG富集分析发现，与CC5325相比，crstk11蓝光响应基因主要参与胞内光合作用催化活性、碳代谢和色素合成等。其中，上调基因包括psaA、psaB和psaC,psbA、psbB、psbC、psbD、psbH和psbL,pet...     

小麦耐低磷相关性状的全基因组关联分析

通过对小麦耐低磷相关性状进行全基因组关联分析(GWAS,genome-wide association study),挖掘与小麦耐低磷性显著相关的单核苷酸多态性标记(SNP,single nucleotide polymorphism)位点及候选基因,为小麦耐低磷性状的遗传基础和分子机制研究提供理论参考。本试验以198份黄淮麦区小麦品种(系)为试验材料,设置低磷和正常磷营养液水培试验,利用小麦35K芯片对分布于小麦全基因组的11896个SNP,采用Q+K关联模型对小麦耐低磷性相关性状进行关联分析。结果表明,小麦耐低磷性状表现出广泛的表型变异,变异系数为15.65%~26.59%,多态性信息含量(PIC,polymorphic information content)为0.095~0.500。群体结构分析表明,试验所用自然群体可分为2个亚群,GWAS共检测到67个与小麦耐低磷相关性状显著关联的SNP位点(P≤0.001),这些位点分布在除3A、3B和3D以外的18条染色体上,单个SNP位点可解释5.826%~9.552%的表型变异。在这些显著位点中有4个SNP位点同时关联到了2个不同的耐低磷性状。对67个SNP位点进行发掘,筛选到7个可能与小麦耐低磷性有关的候选基因。TraesCS6A02G001000和TraesCS6A02G001100在锌指合成中有重要作用;TraesCS6A02G118100可能为低磷胁迫诱导基因;TraesCS5D02G536400、TraesCS1B02G154200和TraesCS5D02G536500与低磷胁迫相关酶类基因家族有关;TraesCS1D02G231200与植物DUF 538结构域蛋白有关,是植物胁迫相关调控蛋白候选基因。     

太空诱变玉米核不育突变体矮化的遗传及外施赤霉素分析

为了解太空诱变玉米核不育突变体矮化的遗传规律和原因,该研究以不育突变体为母本,自交系178、478为父本,对测交 F1、F2群体进行育性鉴定和株高分析,对 F2可育株进行基因型和株高分析,对姊妹交后代分离群体进行育性鉴定和株高、雄穗长度、节间数、节间长度分析,同时,还对姊妹交后代分离群体进行施赤霉素处理,调查育性和株高的变化。结果表明：178和478背景下的 F1表现出与测交母本一样的极显著差异;在178和478核背景下的 F2中,不育株株高极显著矮于可育株,两核背景下的不育株间株高差异不显著,而可育株间株高差异极显著;F2中纯合和杂合可育株的株高差异不显著;姊妹交后代分离群体中不育株株高、雄穗长度、节间数和节间长度极显著小于可育株;外施赤霉素的不育株在苗期表现出对赤霉素一定的敏感性,但株高最终未恢复正常高度。因此,得出该突变体矮化表现稳定,与不育性状并存,且不受细胞核背景的影响;核不育基因对植株株高的矮化无剂量效应;突变体的矮化与雄穗长度、节间数和节间长度有关;突变体不完全属于赤霉素不敏感型,其矮化并不是单一缺乏赤霉素而引起。该研究结果为认识太空诱变玉米核不育突变体矮化的遗传和生理机制提供了参考。     

Genetic analysis and fine mapping of a novel semidominant dwarfing gene LB4D in rice

A dwarf mutant, designated LB4D, was obtained among the progeny of backcrosses to a wild rice introgression line. Genetic analysis of LB4D indicated that the dwarf phenotype was controlled by a single semidominant dwarfing gene, which was named LB4D. The mutants were categorized as dn-type dwarf mutants according to the pattern of internode reduction. In addition, gibberellin (GA) response tests showed that LB4D plants were neither deficient nor insensitive to GA. This study found that tiller formation by LB4D plants was decreased by 40% compared with the wild type, in contrast to other dominant dwarf mutants that have been identified, indicating that a different dwarfing mechanism might be involved in the LB4D dominant mutant. The reduction of plant height in F(1) plants ranged from 27.9% to 38.1% in different genetic backgrounds, showing that LB4D exerted a stronger dominant dwarfing effect. Using large F(2) and F(3) populations derived from a cross between heterozygous LB4D and the japonica cultivar Nipponbare, the LB4D gene was localized to a 46 kb region between the markers Indel 4 and Indel G on the short arm of chromosome 11, and four predicted genes were identified as candidates in the target region.     

Analysis of the rice mutant dwarf and gladius leaf 1. Aberrant katanin-mediated microtubule organization causes up-regulation of gibberellin biosynthetic genes independently of gibberellin signaling

Molecular genetic studies of plant dwarf mutants have indicated that gibberellin (GA) and brassinosteroid (BR) are two major factors that determine plant height; dwarf mutants that are caused by other defects are relatively rare, especially in monocot species. Here, we report a rice (Oryza sativa) dwarf mutant, dwarf and gladius leaf 1 (dgl1), which exhibits only minimal response to GA and BR. In addition to the dwarf phenotype, dgl1 produces leaves with abnormally rounded tip regions. Positional cloning of DGL1 revealed that it encodes a 60-kD microtubule-severing katanin-like protein. The protein was found to be important in cell elongation and division, based on the observed cell phenotypes. GA biosynthetic genes are up-regulated in dgl1, but the expression of BR biosynthetic genes is not enhanced. The enhanced expression of GA biosynthetic genes in dgl1 is not caused by inappropriate GA signaling because the expression of these genes was repressed by GA3 treatment, and degradation of the rice DELLA protein SLR1 was triggered by GA3 in this mutant. Instead, aberrant microtubule organization caused by the loss of the microtubule-severing function of DGL1 may result in enhanced expression of GA biosynthetic genes in that enhanced expression was also observed in a BR-deficient mutant with aberrant microtubule organization. These results suggest that the function of DGL1 is important for cell and organ elongation in rice, and aberrant DGL1-mediated microtubule organization causes up-regulation of gibberellin biosynthetic genes independently of gibberellin signaling.     

Illuminating the role of the Gα heterotrimeric G protein subunit,RGA1, in regulating photoprotection and photoavoidance in rice

We studied physiological mechanisms of photoavoidance and photoprotection of a dwarf rice mutant with erect leaves, d1, in which the RGA1 gene, which encodes the Gα subunit of the heterotrimeric G protein, is non‐functional. Leaves of d1 exhibit lower leaf temperature and higher photochemical reflectance index relative to wild type (WT), indicative of increased photoavoidance and more efficient light harvesting. RNA sequencing analysis of flag leaves revealed that messenger RNA levels of genes encoding heat shock proteins, enzymes associated with chlorophyll breakdown, and ROS scavengers were down‐regulated in d1. By contrast, genes encoding proteins associated with light harvesting, Photosystem II, cyclic electron transport, Photosystem I, and chlorophyll biosynthesis were up‐regulated in d1. Consistent with these observations, when WT and d1 plants were experimentally subjected to the same light intensity, d1 plants exhibited a greater capacity to dissipate excess irradiance (increased nonphotochemical quenching) relative to WT. The increased capacity in d1 for both photoavoidance and photoprotection reduced sustained photoinhibitory damage, as revealed by a higher F_v/F_m. We therefore propose RGA1 as a regulator of photoavoidance and photoprotection mechanisms in rice and highlight the prospect of exploiting modulation of heterotrimeric G protein signalling to increase these characteristics and improve the yield of cereals in the event of abiotic stress.     

矮化香蕉及其野生型GA3ox基因的结构特点和表达分析

香蕉的矮化突变是香蕉无性繁殖后代最常见的表型变异之一,但其变异的分子调控机理目前尚未研究清楚; 而内源赤霉素是影响植物株高的重要激素之一,GA3-氧化酶是赤霉素生物合成后期的关键酶。为探究GA3-氧化酶编码基因对香蕉矮化的分子调控机理,该研究以威廉斯B6矮化突变体及其野生型亲本为材料,通过RT-PCR技术克隆得到矮化香蕉及其野生型亲本GA3ox基因的全长cDNA序列,并对其推测的氨基酸序列进行比对分析,同时利用qRT-PCR技术对GA3ox基因在不同组织中的表达水平差异进行分析。结果表明:(1)矮化香蕉GA3ox-A和野生型香蕉GA3ox-G的ORF长度均为864 bp,均编码287个氨基酸,经序列比对分析发现两条氨基酸序列之间存在5个位点的差异,从而产生具有不同性质的蛋白质。(2)氨基酸序列同源性分析表明,矮化香蕉GA3ox的氨基酸序列与油棕、海枣、椰子的同源性最高。(3)qRT-PCR显示,GA3ox基因在矮化香蕉叶片和茎秆中的表达水平整体上低于野生型,其中GA3ox在野生型茎秆中的表达水平是矮化植株的2.2～32倍。综上推测,GA3ox基因可能对香蕉茎杆的矮化变异具有重要的调控作用。该研究结果为揭示香蕉矮化突变的分子机制与筛选优良矮化香蕉株系奠定了基础。     

Quantitative Proteomics Analysis Identifies the Potential Mechanism Underlying Yellow-Green Leave Mutant in Wheat

Enhancing photosynthesis efficiency is considered as one of the most crucial targets during wheat breeding. However, the molecular basis underlying high photosynthesis efficiency is not well understood up to now. In this study, we investigated the protein expression profile of wheat Jimai5265yg mutant, which is a yellow-green mutant with chlorophylls b deficiency but high photosynthesis efficiency. Though TMT-labeling quantitative proteomics analysis, a total of 72 differential expressed proteins (DEPs) were obtained between the mutant and wild type (WT). GO analysis found that they significantly enriched in thylakoid membrane, pigment binding, magnesium chelatase activity and response to light intensity. KEGG analysis showed that they involved in photosynthesis-antenna protein as well as porphyrin and chlorophyll metabolism. Finally, 118 RNA editing events were found between mutant and WT genotype. The A to C editing in the 3-UTR of TraesCS6D02G401500 lead to its high expression in mutant through removing the inhibition of tae-miR9781, which might have vital role in regulating the yellow-green mutant. This study provided some useful clues about the molecular basis of Jimai5265yg mutant as well as chlorophylls metabolism in wheat.     

矮秆基因对小麦部分农艺性状的效应

以中国主要麦区的124份小麦品种为材料,利用分子标记和系谱分析相结合,对其按照所含的矮秆基因Rht-B1b、Rht-D1b和Rht8进行分类,结合田间株高、旗叶长、小穗数和穗粒数以及室内苗期根系长度等农艺形状的调查,分析不同矮秆基因对小麦农艺性状的效应.结果显示:(1)参试的124份小麦品种(系)中23份含有Rht-B1b,7份含有Rht-D1b,22份含有Rht8基因,34份同时含有Rht-B1b和Rht8,16份同时含有Rht-D1b和Rht8,可分为6组.(2)Rht-B1b和Rht-D1b在降低株高的同时也缩短了旗叶的长度和苗期叶长,Rht8对株高的影响较弱,对旗叶和苗期叶长的影响也较小;3个矮秆基因对苗期根系长度、小穗数没有显著影响;Rht-D1b和Rht8显著增加穗粒数.研究表明,矮秆基因Rht8对小麦株高以及其他农艺性状的影响均较小,但能够显著增加穗粒数,是小麦矮化育种中比较理想的矮秆基因.     

Physiological genetics of the dominant gibberellin-nonresponsive maize dwarfs,Dwarf8 and Dwarf9

Maize (Zea mays L.) Dwarf8-1 (D8-1) is an andromonoecious dwarf mutant proposed to be involved in gibberellin (GA) reception (Fujioka et al. 1988b; Harberd and Freeling 1989). The mutant D8-1 is dominant and GA-nonresponsive (Phinney 1956). We show by map position and similarity of phenotype that five additional dwarf mutants are D8 alleles. We show by map position and similarity of phenotype that a second andromonoecious dwarf mutant, D9-1, defines a duplicate gene. Maize D9-1 and each dominant D8 allele specify a different plant stature, from very mild to very severe dwarfism. Plants of D9-1 and all dominant D8 alleles, except D8-1591, were GA-nonresponsive when treated with 7500 nmol GA₃. The behavior of the mild dwarf D8-1591 was unique in that a small but significant growth response was detected (37% for D8-1591 vs. 130% for the wild type) when treated with 7500 nmol GA₃. These results establish that all dwarf genotypes, except D8-1591, in one dose set a maximum limit on plant growth and block the normal response to GA. When treated with the GA-synthesis inhibitor paclobutrazol, plants of all dwarf genotypes and wild-type siblings were severely dwarfed. Plants of all dwarf genotypes treated with the GA-synthesis inhibitor paclobutrazol and GA₃ were returned to their normal dwarf phenotype. Dominant dwarfing, delayed flowering, increased tillering, and anther development in the ear are characteristic features of D9-1 and all D8 alleles. The GA-synthesis-deficient dwarfs also have these characteristic features. We discuss the function of the wild-type gene product in the context of the observed results.Abbreviations D8 Dwarf8 - D9 Dwarf9 - GA_(n) gibberellin A_(n) - GA₃ gibberellic acid - MNL Maize Genetics Cooperation Newsletter - NIL near-isogenic lines - RFLP restriction fragment length polymorphism - WT wild type This work was supported, in part, by a National Science Foundation Plant Postdoctoral Fellowship to R.G.W., by grants from NIH and ICI Seeds to M.F., the NSF Center for Plant Developmental Biology and the California Agriculture Experiment Station. Much of the work was done in the laboratory of Tim Helentjaris and was supported by a grant from Pioneer Hi-Bred Int'l. The generous gifts of the dominant dwarfing mutants from M.G. Neuffer and O.E. Nelson Jr. are gratefully acknowledged.     

Loss of function of 3-hydroxy-3-methylglutaryl coenzyme A reductase 1 (HMG1) in Arabidopsis leads to dwarfing, early senescence and male sterility, and reduced sterol levels

3-Hydroxy-3-methylglutaryl-CoA reductase (HMGR) catalyzes the first committed step in the cytosolic isoprenoid biosynthesis pathway in higher plants. To understand the contribution of HMGR to plant development, we isolated T-DNA insertion mutants for HMG1 and HMG2. The hmg1 and hmg2 mutants were both more sensitive than the wild type (WT) to lovastatin, an inhibitor of HMGR. The hmg2 mutant showed no visible phenotype under normal growth conditions. In contrast, the hmg1 mutant exhibited dwarfing, early senescence, and sterility. Expression of senescence-associated genes 12 (SAG12), a marker gene for senescence, was induced in the hmg1 mutant at an earlier stage than in the WT. Levels of trans-cytokinins--hormones known to inhibit senescence--were not lower in hmg1. The mutant did not have the typical appearance of brassinosteroid (BR)-deficient mutants, except for a dwarf phenotype, because of the suppression of cell elongation. The expression of several genes involved in cell elongation was suppressed in hmg1. WT plants treated exogenously with inhibitors of sterol biosynthesis had similar gene expression and sterility characteristics as the hmg1 mutants. Pleiotropic phenotypes were rescued by feeding with squalene, the precursor of sterols and triterpenoids. The sterol levels in hmg1 mutants were lower than in the WT. These findings suggest that HMG1 plays a critical role in triterpene biosynthesis, and that sterols and/or triterpenoids contribute to cell elongation, senescence, and fertility.