Molecular mapping and candidate gene analysis of the semi-dominant gene Vestigial glume1 in maize

The glume is an organ of the maize spikelet and plays important roles in anther and kernel development.Vestigial glume1(Vg1)is a classic mutant associated with ligule and glume development.Here we report the phenotypic characterization,fine mapping,and candidate gene analysis of the Vg1 mutant.Vg1 is a semi-dominant and pleiotropic gene,and also affects plant height,ear height,and tassel length.Vg1 ligule degeneration begins at the first leaf,and the Vg1 tassel and ear can be distinguished from those of wild-type plants when their lengths reach respectively 55 mm and 51 mm.Using a BC3 mapping population of 11,445 plants,we delimited the Vg1 functional site to an interval of 7.4 kb,flanked by the markers InDelLM and CRM6.A putative cyclopropane fatty-acid synthase gene(ZmCPA-FAS1)was hypothesized to underlie the mutant phenotype.We detected a Helitron insertion in the sixth intron of ZmCPAFAS1.Its presence caused abnormal alternative splicing of ZmCPA-FAS1 that conferred new characteristics on the Vg1 mutant.These findings are a basis for further discovery of the molecularmechanism underlying glume development and a potential guide formaize breeding of small-glume varieties,especially sweet corn breeding.

Fosmid library construction and screening for the maize mutant gene Vestigial glume 1

The maize mutant gene Vestigial glume 1(Vg1) has been fine-mapped to a narrow region by map-based cloning and the candidate gene for Vg1 spanned 19.5 kb. Here we report Vg1 genomic fosmid library construction and screening. The fosmid library of Vg1 consisted of574,000 clones with an average insert size of 36.4 kb, representing 7.9-fold coverage of the maize genome. Fosmid stability assays indicated that clones were stable during propagation in the fosmid system. Using Vg1 candidate gene-specific primers, a positive clone was successfully identified. This discovery will pave the way for identifying the function of Vg1 in maize development.

Phenotypic characterization of a rice mutant Oryza sativa extraordinary glume 1(Oseg 1)and its genetic analysis

A rice mutant with Yaponica 9522 cultivar background Oryza sativa extraordinary glume 1 （Oseg 1） was identified from the M2 mutant pool mutagenized by ^60Co γ-ray. Compared with wild type plants, Oseg 1 developed longer empty glumes and rudimentary glumes. In some Oseg 1 mutants, the number of stamens of flowers was reduced and leaf-like lodicules occurred, and excessive lemma/palea-like organ could be observed in some mutant spikelets. This indicated that OsEG1 could regulate the development of rudimentary glumes, empty glumes, lemma/palea, lodicules, and stamens. Genetic analysis indicated that Oseg 1 came from a single recessive genetic locus. To clone OsEG1 gene, F2 population was constructed by a cross between Oseg 1 （Japonica） and Guangluai4 （Indica）. Using map-based cloning approach, OsEG1 was mapped on chromosome 4, between INDEL marker OS407 and WHM0466 with genetic distance of 2.0 cm and 1.0 cm, respectively. These results are useful for further cloning and functional analysis of the OsEG1 gene.

Morphogenesis,Anatomical Observation and Primary Genetic Analysis of a Multi-glume Floral Organ Mutant in Rice

A multi-glume （mg） mutant was obtained by screening the T-DNA inserted mutant pool. Anatomical observation revealed that the florets of the mutant showed elongated leafy paleas/lemmas and palea/lemma-like structures, just like multi-glumes. Among the 215 observed florets of the mutant, 14.27% were failed to produce pistil and stamens, 23.72% showed extra floret generated on the same rachilla, while 62.01% consisted of one to nine stamens and one to three pistils in a single floret. On the other hand, in some cases the transparent bulged vesile-like tissue could be observed at the basis of filament. The mutant showed glumaceous Iodicules, which prevented the florets from opening in natural conditions, while the absolute male and female sterility was an obvious character of the current mutant. Observation on the process of floral organ morphogenesis by a scanning electron microscopy （SEM） indicated that no phenotype difference in floret primordia was found between the wild-type and the mutant. Meanwhile, for the mutant, the beginning of stamen and pistil primordial differentiation was later than the wild type and the palea/lemma-like structure continued to differentiate after the formation of normal palea and lemma. Furthermore, in the mutant the asymmetrical division of floral primordial caused variation in the number of stamens and pistils. Therefore, the genetic analyses indicated that the mutation phenotype was a recessive trait controlled by a single gene and co-segregated with the T-DNA. Based on the phenotypic characteristics, it could be deduced that the mutant was the result of homeotic conversion from the function of the class E genes in ABCD model.

Rice AGL1 determines grain size and sterile lemma identity

The grass spikelet is a unique inflorescence structure that determines grain size.Although many genetic factors have been well characterized for grain size and glume development,the underlying molecular mechanisms in rice are far from established.Here,we isolated rice gene,AGL1 that controlled grain size and determines the fate of the sterile lemma.Loss of function of AGL1 produced larger grains and reduced the size of the sterile lemma.Larger grains in the agl1 mutant were caused by a larger number of cells that were longer and wider than in the wild type.The sterile lemma in the mutant spikelet was converted to a rudimentary glume-like organ.Our findings showed that the AGL1(also named LAX1)protein positively regulated G1 expression,and negatively regulated NSG1 expression,thereby affecting the fate of the sterile lemma.Taken together,our results revealed that AGL1 played a key role in negative regulation of grain size by controlling cell proliferation and expansion,and supported the opinion that rudimentary glume and sterile lemma in rice are homologous organs.

BS型小麦光温敏雄性不育系开颖规律研究

BS型小麦光温敏雄性不育系的异交性能直接影响杂交小麦的制种产量,而开花习性是衡量不育系异交性能的重要指标。为了明确开颖率、开颖角度对不育系异交结实率的影响,本研究以15个不育系为材料,同时选取3个配合力好、制种性状优良的恢复系作为父本,分析了不育系在开花过程中开颖率和开颖角度的变化规律及其与异交结实率的相关性。结果表明,在开花过程中,不育系开颖率基本都呈先升后降的变化趋势,且中部和上部穗位开颖较早,但在盛花期,下部穗位开颖角度较大且开颖主要集中在中部穗位,开颖性状较好的不育系为BS278-2、BS185、BS126、BS163、BS136。相关性分析表明,开颖率与开颖小花异交结实率呈极显著正相关,开颖角度与异交结实率呈显著正相关。这说明开颖性状是影响不育系异交结实率的重要因素之一,选育开颖性状好的不育系对提高杂交小麦制种产量具有重要意义。

水稻颖壳异常突变体ah1的鉴定与候选基因分析

【目的】水稻是世界性的粮食作物,其籽粒形态直接影响水稻的最终产量和营养品质,进而影响其经济价值。此外,水稻花发育与籽粒形态又具有复杂的相关性,探究新的水稻花发育调控基因及其分子调控机制可以为培育籽粒更大、更饱满的水稻品种奠定基础。【方法】在利用甲基磺酸乙酯(ethyl methyl sulfonate,EMS)诱变西大1B(籼稻保持系)得到的突变体库中鉴定到一个颖壳和浆片发育异常且矮化的突变体abnormal hull 1(ah1);观察并统计野生型和突变体的农艺性状;选取各个时期的小穗,对突变体的组织学、形态学变化进行分析;采用ah1和籼稻温敏不育系56S构建F2分离群体,并将其用于遗传分析和基因定位;提取野生型和突变体的幼穗RNA并将其反转录为cDNA,对花发育调控基因和ABA合成关键基因的表达量进行RT-qPCR分析。【结果】农艺性状分析表明,突变体ah1各节间的大幅缩短直接导致其矮化;同时,突变体的小穗畸形严重,结实率极低。组织学和形态学分析发现,ah1小穗主要表现为内外稃、浆片和雄蕊等花器官发生了不同程度的退化,部分严重的小穗出现了花器官特征和花分生组织确定性的改变,且伴随大面积的白化,根据其退化程度的不同可分为一般和严重2种类型。遗传分析发现分离群体中野生型和突变体的比值为3:1,表明ah1突变性状受1个隐性基因控制。AH1定位于第1染色体上的分子标记RM6716和RM128之间,物理距离近8 Mb,对突变体进行重测序分析后发现该区间内的LOC_Os01g53450和LOC_Os01g51860在野生型和突变体之间出现了变异,因此,将这两个基因暂定为候选基因。RT-qPCR分析表明,在突变体幼穗早期发育过程中,各花器官发育调控基因的表达量发生了显著的变化;同时,ABA合成关键基因OsNCED1/OsNCED2/OsNCED3/OsNCED4/OsNCED5的表达严重受阻。【结论】AH1对于维持水稻内外稃等花器官的形态建成起到至关重要的作用,将LOC_Os01g53450和LOC_Os01g51860暂定为候选基因。

ELE restrains empty glumes from developing into lemmas

Although there is evident homology among reproductive organs when comparing Poaceae （grass） and eudicots, the identity of grass specific organs, such as lodicules, palea, lemma, and glumes has been the subject of a vast and largely inconclusive discussion. Here we provide some direct evidence to support the idea that the empty glumes of rice （Oryza sativa） are counterparts of lemmas. We show that the development of empty glumes is regulated by ELE （elongated empty glume）, which belongs to a plant specific novel gene family. Mutations at the ELE locus cause elongated empty glumes, which mimic the lemmas and have the epidermal morphology of lemmas with four or five vascular bundles. As a nuclear-localized gene, ELE is specifically expressed at the empty glumes of immature spikelets, and its ectopic expression causes many floral development defects, including lemma-like palea, extra palea-like structures, elongated lodicules, extra stamens and stigmas. Our result suggests that empty glumes are lemmas of the sterile florets located at the lateral side of the rice spikelet, and ELE acts as a regulator restraining its growth to maintain its small size in wild-type plants.

Ectopic expression of VRT-A2 underlies the origin of Triticum polonicum and Triticum petropavlovskyi with long outer glumes and grains

Polish wheat (Triticum polonicum) is a unique tetraploid wheat species characterized by an elongated outer glume. The genetic control of the long-glume trait by a single semi-dominant locus, P1 (from Polish wheat), was established more than 100 years ago, but the underlying causal gene and molecular nature remain elusive. Here, we report the isolation of VRT-A2, encoding an SVP-clade MADS-box transcription factor, as the P1 candidate gene. Genetic evidence suggests that in T. polonicum, a naturally occurring sequence rearrangement in the intron-1 region of VRT-A2 leads to ectopic expression of VRT-A2 in floral organs where the long-glume phenotype appears. Interestingly, we found that the intron-1 region is a key ON/OFF molecular switch for VRT-A2 expression, not only because it recruits transcriptional repressors, but also because it confers intron-mediated transcriptional enhancement. Genotypic analyses using wheat accessions indicated that the P1 locus is likely derived from a single natural mutation in tetraploid wheat, which was subsequently inherited by hexaploid T. petropavlovskyi. Taken together, our findings highlight the promoter-proximal intron variation as a molecular basis for phenotypic differentiation, and thus species formation in Triticum plants.

The C2H2 zinc-finger protein LACKING RUDIMENTARY GLUME 1 regulates spikelet development in rice

Rice(Oryza sativa)spikelets are a unique inflorescence structure and their development directly determines grain size and yield.Although many genes related to spikelet development have been reported,the molecular mechanisms underlying this process have not been fully elucidated.In this study,we identified a new recessive rice mutant,lacking rudimentary glume 1(lrg1).The lrg1 spikelets only formed one rudimentary glume,which,along with the sterile lemmas,was homeotically transformed into lemma-like organs and acquired lemma identity.The transition from the spikelet to the floral meristem was delayed in the lrg1 mutant,resulting in the formation of an ectopic lemma-like organ between the sterile lemma and the terminal floret.In addition,we found that the abnormal lrg1 grain phenotype resulted from the alteration of cell numbers and the hull size.LRG1 encodes a ZOS4-06-C2H2 zinc-finger protein with the typical EAR motifs,and is expressed in all organs and tissues.LRG1 localizes to the nucleus and can interact with the TOPLESS-RELATED PROTEINs(TPRs)to repress the expressions of their downstream target genes.Taken together,our results reveal that LRG1 plays an important role in the regulation of spikelet organ identity and grain size.

孕穗期高温对水稻颖壳发育的影响

[目的]揭示孕穗期高温是影响颖壳发育的关键时期,为北方水稻生产中预防高温热害提供理论依据。[方法]以千重浪2号为试验材料,在盆栽条件下研究孕穗期高温胁迫对颖壳发育的影响及颖壳大小与粒重的关系。[结果]孕穗期高温显著地降低了颖壳的长度、宽度和面积。强、中势粒降低幅度最大的处理时期是叶龄余数为1.0;而弱势粒降低幅度最大的时期是叶龄余数为0即剑叶完全展开时。相关分析表明,颖壳大小与粒重的关系更为密切,随着颖壳长、宽和面积的增加,粒重也明显增加。[结论]在水稻生产上,在孕穗期要特别注意预防叶龄余数接近1.0时的热害,根据天气状况,采取相应的预防措施。

2个小麦AL型细胞质雄性不育系的异交特性分析

提高不育系的异交结实率是杂交小麦制种产量取得突破的重要途径。为明确小麦AL型雄性不育系的异交结实特性,连续3年测定了2个小麦AL型不育系36A和3235A的柱头活力、柱头外露率、颖间距和自然异交结实率。结果表明,36A和3235A的平均自然异交结实率分别为113.02%和44.65%,柱头活力分别为104.96%和76.7%,差异均达到显著水平;二者的柱头外露率和颖间距无显著差异。相关分析表明,授粉天数与柱头活力呈显著负相关,柱头活力随授粉天数的延长而降低;柱头活力与柱头外露率和颖间距、柱头外露率与颖间距间均呈显著正相关。随授粉进程推移,柱头活力、柱头外露率和颖间距均呈先增后减趋势,36A的3个指标均高于3235A,综合表现为异交结实率高。花后第4~11天套袋饱和授粉异交结实率最高,授粉后第6~12天,柱头外露率和颖间距均达到最大。因此,小麦AL型不育系的恢复系扬花始期比不育系推迟2~3 d,小花陆续开放、柱头活力也较强,并持续开颖,可保持较高结实率,为最佳授粉时间。

黑稻颖壳和籽粒的营养成分与饲用价值分析

为研究黑稻颖壳和籽粒的饲用价值,通过对黑稻颖壳和籽粒的常规营养成分、花色苷、总黄酮、氨基酸和脂肪酸等组分进行测定,结合有机物质消化率、相对饲料价值等指标对颖壳和籽粒的饲用价值进行综合评价。结果显示,黑稻颖壳和籽粒均富含花色苷与黄酮等生理活性物质。黑稻颖壳由于粗纤维、中性洗涤纤维、酸性洗涤纤维含量较高,导致其有机物质消化率仅为26.35%;颖壳虽然氨基酸含量不高,且未检测出蛋氨酸和胱氨酸,但其脂肪酸种类较为丰富,包含亚油酸、亚麻酸和花生五烯酸EPA在内的17种脂肪酸,且其相对饲料价值RFV为103.26,达到饲料二级标准。黑稻籽粒的粗蛋白质含量达到8.73%,必需氨基酸模式与FAO模式相当。籽粒的油酸与亚油酸的比例为1.09∶1,基本符合国际卫生组织推荐的1∶1的摄入比例;而且饱和脂肪酸(SFA):单不饱和脂肪酸(MUFA):多不饱和脂肪酸(PUFA)的比值较为接近中国营养协会推荐的“<1∶1∶1”的比例。黑稻籽粒由于粗纤维、中性洗涤纤维、酸性洗涤纤维含量极低,故其有机物质消化率极高,且相对饲料价值RFV为1626.98,达到饲料特级标准。综上所述,黑稻籽粒可直接作为优质饲料资源使用。

水稻小粒突变体smg2的表型鉴定和候选基因分析

水稻产量是由单位面积有效穗数、每穗粒数以及千粒质量决定的,籽粒形态可以决定千粒质量进而影响水稻产量.报道了一个与水稻籽粒形态发育相关的突变体,来源于籼稻保持系西大1B的甲基磺酸乙酯(ethyl methane sulfonate,EMS)诱变群体.该突变体表现为由细胞扩展和细胞增殖异常导致的籽粒变小,暂命名为水稻小粒突变体(small grain 2,smg2).遗传分析表明:smg 2突变体性状受1对隐性单基因控制,利用群体分离分析法(bulked segregation analysis,BSA)将SMG 2基因定位在第1染色体IN/DEL标记A-0.85和A-1.05之间,物理距离200 kb,在定位区间内含注释基因28个.PCR测序发现其中编号为LOC_Os01g02890的基因中第1个外显子的第153位碱基发生了单碱基缺失,造成编码框移码突变,从而导致蛋白翻译提前终止.因此,将LOC_Os 01 g 02890暂定为SMG 2的候选基因.

水稻短宽粒基因SWG1的图位克隆

【目的】水稻产量由单位面积有效穗数、每穗粒数和粒重3个因素构成,其中,粒重主要由水稻的籽粒形态决定。筛选和鉴定新的粒型突变材料和基因,可为产量性状的分子设计育种奠定基础。【方法】在籼稻保持系西大1B(XD1B)的甲基磺酸乙酯(EMS)诱变群体中鉴定到一个短宽粒突变体short and widen grain 1(swg1);分析籽粒形态和其他农艺性状,并对颖壳进行组织细胞学观察分析;运用BSA法进行基因定位;通过遗传互补试验确定候选基因;采用qRT-PCR分析该基因的表达模式及其他粒型相关基因和细胞发育基因的表达水平。【结果】农艺性状分析发现,与野生型相比,swg1突变体粒长显著降低,粒宽显著增加,表现出短宽粒的表型;进一步组织和细胞学分析,发现突变体颖壳纵向细胞变短是粒长变短的主要原因,而粒宽增加是由于颖壳横向细胞数目和细胞大小同时增加。遗传分析结果表明,该突变性状受隐性单基因控制,通过图位克隆与遗传互补验证,确定候选基因为LOC_Os07g42410,编码一个植物特异转录因子。qRT-PCR分析发现该基因表达无明显的组织特异性,在茎、叶、幼穗中表达强烈。通过对已知粒型相关基因、细胞周期和细胞扩展相关基因进行分析,发现通过正向调控颖壳横向细胞数目和(或)细胞大小决定粒宽的GS5和GW8在突变体中上调明显,而正向调控纵向细胞数目和大小并负向调控横向细胞数目和大小的GW7/GL7在突变体明显下调;另外,部分与细胞周期和细胞扩展相关的基因也在突变体和野生型之间的表达也呈现显著差异。【结论】SWG1编码一个植物特异的转录因子,通过调控粒型基因(GS5、GW8和GW7/GL7等)影响颖壳的细胞增殖和细胞扩展,从而决定水稻籽粒长度和宽度。

小麦黑色颖壳兼抗赤霉病新种质KD522的选育及其鉴定

通过人工杂交选育出一个具有黑色颖壳(黑颖)、抗赤霉病的高代小麦新品系KD522,将其与黄白色颖壳(白颖)品系2073配置杂交组合,对杂交后代的颖壳颜色进行遗传分析和遗传效应评价。抽穗后,KD522的颖壳与麦芒均为正常绿色,灌浆后期—蜡熟期颖色加深,完熟期变成黑色。KD522与2073杂交后代F_(1)均为黑颖,F_(2)颖色出现分离,经χ^(2)检测,黑颖与白颖的比例为3∶1,黑颖由单显性基因控制。黑颖单株株高、穗长、小穗数、穗粒数、旗叶叶绿素含量均与白颖单株无显著差异。KD522中抗赤霉病,可作为抗赤霉病的种质资源。研究结果可为小麦颖壳颜色与赤霉病抗性的遗传改良研究提供基础和依据。

Research Progress on Mechanized Mixed Sowing Seed Production Technology of Hybrid Rice

Hybrid rice planting has been widely popularized and applied in the world. However, the high cost of seed production and the complicated procedures have become a bottleneck in the development of hybrid rice. The research progress on mixed sowing seed production techniques of hybrid rice was introduced from the aspects of rice resources creation, breeding, sowing seed technology research and cost benefit analysis. The production technology of the new mixed seeding combina- tion ＂Xinhunyou 6＂ was investigated, including the research and validation of benta- zon treatment period and dosage, mixing ratio of male and female parents, and the comparative test of different different sowing methods, which revealed that the mechanization technology of seed production of hybrid rice was mature and feasible and would be one of the most important development trend of technological devel- opment of hybrid rice production.

稻谷颖壳开裂特点及其影响的初步研究

调查同一组合2个典型株系共10个植株穗粒常规稻颖壳开裂特点及其影响。结果表明:稻谷颖壳开裂粒间表现有轻重之分;品质不同株系籽粒裂颖率存在较大差异,稻穗中部或下部及二次枝梗上的稻谷颖壳开裂现象严重;裂颖稻粒形成的糙米与正常稻粒形成的糙米相比,斑点米、裂纹米、有色米比率及厚度有所增加,而千粒重、长度、宽度减小。

高温胁迫对国稻6号开花结实习性的影响

为研究杂交稻对开花结实期高温胁迫的生理生态适应性,选用具代表性的主栽杂交稻组合国稻6号、两优培九等,人工设计极值高温40℃,自始穗后第15天每天6h进行热害处理,以自然气候条件为对照,测评结实的热害指数,并从中选择对开花结实期高温胁迫反应差异较大的国稻6号与协优46,观察高温胁迫对其开花结实习性的影响。结果表明,国稻6号具较高耐热性似与其"被动避热"适应性有关,主要表现在开花结实习性的变化,首先是花期缩短,花期集中,"粒间顶端优势"有所减弱;其次是花时分散,开花峰期峰值下降,虽未见花时提前,但见日初花量增加与花时向后延长2h。

水分胁迫对燕麦穗颖渗透调节和抗氧化能力的影响

以抗旱性不同的燕麦品种‘蒙燕1号’（抗旱性强）和‘坝莜3号’（水分敏感）为试验材料,采用盆栽方式研究了抽穗期和灌浆期水分胁迫对燕麦穗颖渗透调节和抗氧化能力的影响.结果表明：（1）水分胁迫处理均显著促进了不同抗旱性品种穗颖渗透调节物质（游离脯氨酸和可溶性蛋白）含量增加,并以抗旱品种累积水平高于水敏感品种,且两种渗透调节物质对抽穗期胁迫的反应比灌浆期胁迫更敏感.（2）两时期的水分胁迫处理均能降低不同抗旱性品种穗颖SOD和POD活性,抗旱品种的保护酶活性要高于水敏感品种,抗旱品种的SOD活性降低幅度明显低于水敏感品种,而POD活性降低幅度在两品种间差异不明显.（3）水分胁迫导致2个品种穗颖丙二醛（MDA）含量和相对电导率显著增加,细胞膜结构受到严重伤害,且水敏感品种受害程度大于抗旱品种.（4）水分胁迫使2个品种单株籽粒产量下降,且在中度胁迫和重度胁迫下,抗旱品种的减产幅度要低于同期水敏感品种;水分胁迫下,水敏感品种‘坝莜3号’减产4.54％～30.29％,抗旱品种‘蒙燕1号’减产6.69％～23.54％.可见,抗旱性强的燕麦品种在受到水分胁迫的条件下能通过增强穗颖渗透调节和抗膜质过氧化能力、减弱穗颖细胞质膜损伤程度来适应干旱胁迫,最大限度减少水分胁迫对穗颖的伤害,有利于稳产.