棉花叶皱小突变体的遗传分析及激素含量研究

在海岛棉(Gossypium barbadense L.)品种海7124的基因组总DNA导入陆地棉(G.hirsutum L.)品种石远321的后代中,发现了一个形态性状发生变异的突变体。与正常棉株相比,突变体叶片皱小、茎杆细弱,铃小但种子发育正常。叶表皮显微观察发现,突变体叶表皮细胞增大,突变体叶片变小是由于叶片细胞数目的减少。遗传分析表明,突变体为杂合基因型,突变性状受显性基因控制,并可能具有纯合致死效应。内源激素含量测定显示,突变体茎尖中IAA和ZR含量显著高于正常棉株,推测突变体的叶片皱缩变小可能与主茎顶芽中IAA和ZR含量的异常有关。     

小麦穗部发育多效基因的遗传分析与基因定位

在小麦育种材料中首次发现一种穗部发育萎缩且花器官明显退化,但茎、叶等其他器官发育正常的突变体sda1(spike development atrophy 1)。用显微镜观察突变体sda1的花器官,用碘-碘化钾鉴定其小孢子育性;以‘陕麦94’为父本,突变材料sda1为母本构建F2群体,调查各主要农艺性状,灌浆期测定穗部及穗下茎可溶性糖含量、旗叶光合性能(净光合速率、气孔导度、胞间CO2浓度、蒸腾速率),对该突变体进行遗传分析;利用SSR微卫星标记,通过混合分离分析(BSA)和群体连锁分析进行基因定位,进一步探索该基因功能。结果表明:(1)小麦突变体sda1雄蕊发育畸形,雌蕊发育萎缩,小孢子几乎全部丧失育性。(2)对突变体sda1原株系中表型正常植株的后代分离统计分析结果证明,该突变性状由1对隐性核基因控制,并命名该基因为SDA1。(3)在F2群体中,突变株抽穗期较正常株延迟4d;穗部及穗下茎可溶性糖含量分别显著高于正常株30.6%和11.0%,但突变株与正常株的抽穗持续时间(均为8d)和光合性能无显著差异。(4)经基因定位分析初步确定SDA1位于小麦6B染色体WMC398和BARC136标记之间,与两标记的遗传距离分别为2.2cM和2.1cM。推测认为,SDA1是一个控制抽穗期与器官发育的多效基因,且该基因突变影响植株的糖分转化与利用。     

辣椒矮化及果皮皱缩突变体鉴定RAPD初步分析

从‘怀椒六号'60Co-γ射线辐射后代中选得一株突变体,该突变体株高较矮,侧枝较少.野生型果皮光滑,而突变体果皮皱缩.经连续两年种植比较发现,该突变性状能稳定遗传,经RAPD分子标记比较发现,该突变性状是基因突变引起,并且从F1代表型分析发现,决定突变性状的基因为隐性基因.野生型与突变体花朵性状、果长、叶长、坐果能力、单果重、果肉厚、心室数等性状方面无显著性差异.     

三个陆地棉芽黄突变体的遗传及育种利用研究

棉花芽黄性状是一种优良的指示性状,在棉花杂种优势研究中具有重要的利用价值。本研究以3个芽黄突变体71-7、62-17和115-23为试验材料,配制了33个杂交组合,研究芽黄性状分离规律、芽黄性状的回交转育和优势组合选配。结果表明:71-7和115-23两个突变体的芽黄基因为单基因控制的隐性性状,62-17材料为单基因控制的不完全显性性状;通过回交转育方法,创制了9份带有芽黄标记性状的陆地棉三系种质资源;通过配制的12个优势组合综合性状分析,筛选出组合Y34和Y26具有较高的籽棉产量和皮棉产量,并可作为早熟杂交棉新品系选送新疆维吾尔自治区早熟陆地棉组进行品种参试。本研究结果可为新疆棉花种质创制、新基因挖掘及新品种培育提供新的亲本及基因来源。     

烟草黄绿叶突变体的遗传分析与基因定位

以G117为父本、RG13为母本,杂交获得F1群体。系谱法常规选择过程中,在F3株系内发现黄绿自然隐性突变株。该突变体的叶色在旺长期前呈现正常绿色,进入旺长期后,叶色逐渐发黄,叶脉呈乳白色,与正常烟株差别明显。遗传分析表明该突变体性状受1对隐性基因控制。从来源于一个连续自交单株的分离群体中分别选取10份隐性纯合烟株和10份显性烟株,利用430K烟草高密度SNP芯片进行基因型分析,快速确定了与目标性状相关联的标记。进而利用该分离群体验证相关分子标记,将该基因定位在烟草第5号染色体M7和M18之间,并与M7标记共分离。相关研究为进一步克隆该基因奠定了基础,同时也为烟草其他重要性状的定位提供了一种有效、快速的方法。     

小麦矮秆圆粒突变体的鉴定与分析

小麦的子粒形态性状和株高与小麦的产量密切相关,是育种的重要选择目标性状。本研究通过对小麦品种偃展1号(YZ1)EMS突变体W98的农艺性状的调查与分析,发现W98的株高只有24 cm,而野生型YZ1的株高是73 cm。突变体株高的变异是由每个节间长度变短造成的,而非节间数目减少所致。相关分析表明矮秆与圆粒性状呈显著相关。利用高秆长粒的墨西哥品种10th12与突变体W98配制杂交组合,获得1544个F2∶3单株(株系),通过对分离群体的遗传分析,发现圆粒性状是由1对不完全显性基因控制的。赤霉素(GA)与油菜素内酯(BR)激素敏感性试验表明:野生型和突变体都对GA处理不敏感;不同浓度BR的展叶试验表明野生型对BR不敏感,而突变体W98对BR敏感。     

转双价抗虫基因棉花的主要农艺性状的遗传变异

以11个转外源基因(CrylAc+API-B)系为材料,受体亲本陆地棉品种鄂抗9号为对照,研究其在抗棉铃虫性、产量和纤维品质性状上的差异.结果表明:11个转基因系的抗棉铃虫效果明显,达到中抗和高抗水平;转基因系及其受体、转基因系之间在单铃重等10个性状上均存在极显著性差异;和受体亲本相比,各个转基因系变异方向比较一致的性状有:衣指、衣分降低;而株高、单株结铃数、单铃重、子指、皮棉产量、纤维长度、断裂比强度、马克隆值、伸长率等性状的变化方向不定.     

拟南芥株型突变体zpr1的性状特征与基因鉴定分析

对本研究室经T-DNA插入法获得的拟南芥株型突变株系——隐性突变体zpr1植株进行植物学性状调查和遗传分析,并对该突变基因进行鉴定、表达定位和调控元件分析。结果显示:(1)性状分析表明,与野生型拟南芥Ws-2相比,突变体zpr1的茎生叶分枝数量增加,茎生叶分枝发生于拟南芥顶端花序部位;野生型拟南芥茎生叶为披针形,而突变体zpr1没有出现分枝的茎生叶呈倒卵形,出现分枝的茎生叶呈披针型;突变体zpr1的主花序高度、株高、分枝高度和分枝长度都高于野生型,且分枝数多于野生型。(2)利用质粒挽救和反向PCR法(IPCR)确定了ZPR1基因突变发生位置是该基因起始密码子上游426bp处,证明T-DNA插入破坏了ZPR1基因的启动子区域,导致该基因在拟南芥内不能正常表达。(3)基因转录调控区域的顺式作用元件分析发现在ZPR1基因的转录调控区有多个与植物激素相关的调控元件,还有与光周期调节相关的调控元件。(4)亚细胞定位发现,ZPR1基因在所有细胞中的细胞膜中表达,而在部分细胞的细胞膜、细胞质和细胞核中均有表达。研究表明,ZPR1基因的表达对植物株型发育有重要的调控作用,该基因的表达水平受植物激素和光照的调节,最终导致了植物株型的变化。     

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。     

棉花矮化突变体的遗传分析

陆地棉科遗２号×中棉完紫的种间杂交衍生后代群体中分离出一株矮秆小叶突变体,经多年选择育成了矮早棉１号。在北京气候条件下,矮早棉１号成熟时,株高只有４５ｃｍ,不到正常陆地棉的１／２。遗传分析揭示矮早棉１号的矮化早熟特性系由两对隐性基因控制,其基因符号定名为ｄ_１和ｄ_２,矮早棉１号为双隐性纯合子,基因型为ｄ_１ｄ_１ｄ_２ｄ_２。正常陆地棉ＴＭ－１、中棉所１２及中棉所１６均为显性纯合子,基因型为Ｄ_１Ｄ_１Ｄ_２Ｄ_２。控制棉花株高的两对基因Ｄ_１／ｄ_１和Ｄ_２／ｄ_２间表现重叠作用。矮早棉１号在棉花早熟育种中有重要价值。     

Ethyl methanesulfonate mutant library construction in Gossypium hirsutum L. for allotetraploid functional genomics and germplasm innovation

As the gene pool is exposed to both strain on land resources and a lack of diversity in elite allotetraploid cotton, the acquisition and identification of novel alleles has taken on epic importance in facilitating cotton genetic improvement and functional genomics research. Ethyl methanesulfonate (EMS) is an excellent mutagen that induces genome‐wide efficient mutations to activate the mutagenic potential of plants with many advantages. The present study established, determined and verified the experimental procedure suitable for EMS‐based mutant library construction as the general reference guide in allotetraploid upland cotton. This optimized method and procedure are efficient, and abundant EMS mutant libraries (approximately 12 000) in allotetraploid cotton were successfully obtained. More than 20 mutant phenotypes were observed and screened, including phenotypes of the leaf, flower, fruit, fiber and plant architecture. Through the plants mutant library, high‐throughput and high‐resolution melting technology‐based variation evaluation detected the EMS‐induced site mutation. Additionally, based on overall genome‐wide mutation analyses by re‐sequencing and mutant library assessment, the examination results demonstrated the ideal quality of the cotton EMS‐treated mutant library constructed in this study with appropriate high mutation density and saturated genome. What is more, the collection is composed of a broad repertoire of mutants, which is the valuable resource for basic genetic research and functional genomics underlying complex allotetraploid traits, as well as cotton breeding.     

New red flower germplasm lines of cotton selected from hybrid ofGossypium hirsutum XG. bickii

By means of dropping GA₃(50 ppm) and NAA (40 ppm) on the hybrid boll-embryo culturein vitro, one F₁ plant ofG. hirsutum × G. bickii was obtained; when F₁ branches were grafted on upland cotton and then back-crossed with upland cotton under short-day and cooler-night condition, some BC₁ seeds could be harvested. The characteristic segregation was very violent in early generation. Through 3 times of back-crossing and selecting, ten stable hybrid lines with the character of both male parent (viz. red petal-purple spot and strong fibre) and female parent (plant type, earliness, white fibre, lint length, etc.) were established. These lines were assigned as HB red flower lines (HBRL). Transference of character ofG. bickii to upland cotton was proved to be successful for the first time. These new germplasms may play an important role in both the genetic research and new cotton variety breeding.     

New red flower germplasm lines of cotton selected from hybrid of Gossypium hirsutum × G. bickii

By means of dropping GA₃(50 ppm) and NAA (40 ppm) on the hybrid boll-embryo culturein vitro, one F₁ plant ofG. hirsutum × G. bickii was obtained; when F₁ branches were grafted on upland cotton and then back-crossed with upland cotton under short-day and cooler-night condition, some BC₁ seeds could be harvested. The characteristic segregation was very violent in early generation. Through 3 times of back-crossing and selecting, ten stable hybrid lines with the character of both male parent (viz. red petal-purple spot and strong fibre) and female parent (plant type, earliness, white fibre, lint length, etc.) were established. These lines were assigned as HB red flower lines (HBRL). Transference of character ofG. bickii to upland cotton was proved to be successful for the first time. These new germplasms may play an important role in both the genetic research and new cotton variety breeding.     

一年生陆地棉细胞核雄性不育系洞A的宿生栽培效应

植物细胞核雄性不育系具有易于恢复但保持困难的特点。根据广西南部冬季无霜冻的气候特点,在广西南宁进行了一年生陆地棉细胞核雄性不育系洞A的三年露地栽培。结果表明:二、三年生洞A在5月上旬开花、6月下旬吐絮,这是一年生洞A在相同环境条件下难以达到的;与一年生洞A相比,二、三年生洞A的子指、单株铃数、产量显著增加,但单铃重、衣分显著下降,僵瓣率显著增加;纤维品质基本没有显著变化。说明利用陆地棉细胞核雄性不育系在南亚热带的宿生栽培进行良种繁育具有较好的前景。     

Genome- Wide Analysis and Characterization of the TRX Gene Family in Upland Cotton

Thioredoxins (TRX) are small molecules of proteins that are present in all organisms. TRXs play an important role in diverse functions of plant growth and development. In this study, we performed genome-wide, characterization and expression levels of TRX gene family in cotton. A total of 150 GhTRX proteins were identified in upland cotton and classified into five subfamilies based on their domain compositions. Phylogenetic tree analysis divided TRX genes into seven subgroups. GhTRX genes covered all upland cotton chromosomes, with duplicated gene events. Ka/Ks ratio of three gene pairs was less than 1, suggesting purifying selection. The functions of GhTRX genes were studied using gene ontology, protein localization, and promoter analysis. Furthermore, six GhTRX genes were randomly selected to examine their expression level in cotton development and under various exogenous treatments. The genes showed high expressions in various tissues and at different stages of leaf senescence, also showed high expression under abscisic acid, ethylene, drought, and salinity. This study reveals the first report of TRX family genes in upland cotton. However further studies are needed to elucidate their specific functions in cotton plant.     

Viewing the difference between the diploid and the polyploid in the light of the upland cotton aneuploid

The aneuploidy of Gossypium hirsutum L. (upland cotton) aneusomatics were obtained by induced parthenogenesis. These aneuploids could grow and set seeds normally. In the process of meiosis there appeared large quantities of heteromorphic pairs and multivalent chromosomes and many cases of cytomixis and multisperm fertilization occurred. The aneuploids produced offsprings through sexual propagation. We explored penetratingly the questions how and why these aneuploids could survive. Through this research, we found that the upland cotton possessed an immense latent capacity to adapt to adverse environments. More importantly, in the case of the upland cotton, we discovered that the genetic pattern of the polyploid differs in some respects from that of the diploid.     

海岛棉与陆地棉叶绿素含量变化的差异研究

以5份陆地棉和4份海岛棉为材料,研究了整个发育进程中陆地棉和海岛棉叶绿素的变化规律之间的差异,结果显示：陆地棉与海岛棉的叶绿素a、叶绿素b和叶绿素a＋b各有不同动态变化规律,其叶绿素a/b在整个发育进程中的变化也有差异。海岛棉与陆地棉的叶绿素a变化趋势有较大差别,陆地棉的叶绿素a含量在6月18日（苗期）最高,峰值出现在8月6日,海岛棉峰值、最大值均在7月4日（现蕾期）;叶绿素b含量表现为陆地棉、海岛棉峰值均在8月6日（花铃期）,但海岛棉平均值高于陆地棉;陆地棉的叶绿素a＋b和叶绿素a曲线相似,海岛棉的叶绿素a＋b和叶绿素b曲线相似;陆地棉叶绿素a/b值表现苗期最大,然后迅速下降,海岛棉a/b值表现普遍低于陆地棉,其变化趋势为前期在7月4日出现峰值,然后下降再升高。实验说明陆地棉和海岛棉叶绿素的合成机制、调控机理可能不同。     

Genome-wide association study identified genetic variations and candidate genes for plant architecture component traits in Chinese upland cotton

Key message

Thirty significant associations between 22 SNPs and five plant architecture component traits in Chinese upland cotton were identified via GWAS. Four peak SNP loci located on chromosome D03 were simultaneously associated with more plant architecture component traits. A candidate gene, Gh_D03G0922, might be responsible for plant height in upland cotton.

Abstract

A compact plant architecture is increasingly required for mechanized harvesting processes in China. Therefore, cotton plant architecture is an important trait, and its components, such as plant height, fruit branch length and fruit branch angle, affect the suitability of a cultivar for mechanized harvesting. To determine the genetic basis of cotton plant architecture, a genome-wide association study (GWAS) was performed using a panel composed of 355 accessions and 93,250 single nucleotide polymorphisms (SNPs) identified using the specific-locus amplified fragment sequencing method. Thirty significant associations between 22 SNPs and five plant architecture component traits were identified via GWAS. Most importantly, four peak SNP loci located on chromosome D03 were simultaneously associated with more plant architecture component traits, and these SNPs were harbored in one linkage disequilibrium block. Furthermore, 21 candidate genes for plant architecture were predicted in a 0.95-Mb region including the four peak SNPs. One of these genes (Gh_D03G0922) was near the significant SNP D03_31584163 (8.40 kb), and its Arabidopsis homologs contain MADS-box domains that might be involved in plant growth and development. qRT-PCR showed that the expression of Gh_D03G0922 was upregulated in the apical buds and young leaves of the short and compact cotton varieties, and virus-induced gene silencing (VIGS) proved that the silenced plants exhibited increased PH. These results indicate that Gh_D03G0922 is likely the candidate gene for PH in cotton. The genetic variations and candidate genes identified in this study lay a foundation for cultivating moderately short and compact varieties in future Chinese cotton-breeding programs.

     

Generation of ESTs for flowering gene discovery and SSR marker development in upland cotton

Background

Upland cotton, Gossypium hirsutum L., is one of the world''s most important economic crops. In the absence of the entire genomic sequence, a large number of expressed sequence tag (EST) resources of upland cotton have been generated and used in several studies. However, information about the flower development of this species is rare.

Methodology/Principal Findings

To clarify the molecular mechanism of flower development in upland cotton, 22,915 high-quality ESTs were generated and assembled into 14,373 unique sequences consisting of 4,563 contigs and 9,810 singletons from a normalized and full-length cDNA library constructed from pooled RNA isolated from shoot apexes, squares, and flowers. Comparative analysis indicated that 5,352 unique sequences had no high-degree matches to the cotton public database. Functional annotation showed that several upland cotton homologs with flowering-related genes were identified in our library. The majority of these genes were specifically expressed in flowering-related tissues. Three GhSEP (G. hirsutum L. SEPALLATA) genes determining floral organ development were cloned, and quantitative real-time PCR (qRT-PCR) revealed that these genes were expressed preferentially in squares or flowers. Furthermore, 670 new putative microsatellites with flanking sequences sufficient for primer design were identified from the 645 unigenes. Twenty-five EST–simple sequence repeats were randomly selected for validation and transferability testing in 17 Gossypium species. Of these, 23 were identified as true-to-type simple sequence repeat loci and were highly transferable among Gossypium species.

Conclusions/Significance

A high-quality, normalized, full-length cDNA library with a total of 14,373 unique ESTs was generated to provide sequence information for gene discovery and marker development related to upland cotton flower development. These EST resources form a valuable foundation for gene expression profiling analysis, functional analysis of newly discovered genes, genetic linkage, and quantitative trait loci analysis.