三个水稻联乙烯还原酶基因突变体的比较研究

 【Objective】 The present study was aimed at investigating the differences of mutant phenotypes resulting from different mutations in base sequence of rice divinyl reductase gene OsDVR (Os03g22780), and evaluating application potential of this gene in rice breeding as a leaf color marker gene. 【Method】The yellow-green leaf mutants isolated from Indica rice variety G46B and japonica rice variety Nipponbare via ethyl methanesulfonate mutagenesis were detected using high-performance liquid chromatography (HPLC) to screen the mutants accumulating divinyl chlorophyll. Then genetic analysis, gene mapping and allelism test of the target mutants were performed. Subsequently, DNA sequencing of OsDVR gene in these mutants and alignment of the deduced amino acid sequences of homologous DVR proteins were conducted, and leaf photosynthetic pigments, plant phenotypes and main agronomic traits of the mutants were investigated. 【Result】590ys and 525ys, two novel mutants accumulating divinyl chlorophyll, were obtained by screening 53 yellow-green leaf mutants isolated from ethyl methanesulfonate mutagenesis. Their mutant phenotypes were all controlled by a pair of recessive nuclear gene and moreover, the mutant genes were all mapped on the chromosomal region harbouring the 824ys mutant gene reported. Furthermore, allelism test confirmed that the mutant genes of 590ys and 525ys were allelic to that of 824ys. So 590ys, 525ys and 824ys mutants were all OsDVR mutants. However, mutational sites of OsDVR mutant gene and their encoded protein product in the three mutants were different, which resulted in extremely significant differences of chlorophyll contents and compositions, plant phenotypes, main agronomic traits and yields per plant among the three mutants. Among them, 525ys mutant had slight yellow leaves, and its growth and development, main agronomic traits and yield per plant were affected relatively less, indicating that 525ys mutant gene can basically meet the requirements as a leaf color marker gene introduced into rice male sterile lines. 【Conclusion】Different mutations in base sequence of OsDVR gene could result in extremely significant differences of mutant phenotype and yield per plant. The OsDVR mutant gene could be applicable in hybrid rice breeding as a leaf color marker gene.     

水稻叶色突变体基因定位研究进展

水稻叶色突变体是一类较易观察和获得的突变体,研究水稻叶色突变体基因进行定位有助于深入阐明叶色基因的调控机理,对水稻叶片光合遗传改良和产量提高具有重要的指导意义。因此,对于近年来有关水稻叶色突变体的分类来源、遗传机理以及基因定位等研究进展进行综述,并介绍了叶色突变体的分子机理及其在水稻中的应用,旨在为水稻叶色突变体的研究和利用奠定基础。     

GraS is critical for chloroplast development and affects yield in rice

Leaf color has been considered an important agronomic trait in rice (Oryza sativa L.) for a long time. The changes in leaf color affect the yield of rice. In this study, a green-revertible albino (graS) mutant was isolated from a ⁶⁰Co-gamma-irradiated mutant pool of indica cultivar Guangzhan 63-4S. The fine mapping indicated that graS mutant was mapped to chromosome 1, and was located in a confined region between markers ab134 and InDel 8 with genetic distances of 0.11 and 0.06 cM, respectively. Based on the annotation results, four open reading frames (ORFs) were predicted in this region. Sequence analysis revealed that LOC_Os01g55974 had a 2-bp nucleotide insertion (AA) in the coding region that led to premature termination at the 324th base. Sequence analysis and expression analysis of related genes indicated that LOC_Os01g55974 is the candidate gene of GraS. We studied the genome and protein sequences of LOC_Os01g55974, and the data showed that GraS contains a deoxycytidine deaminase domain, which was expressed ubiquitously in all tissues. Further investigation indicated that GraS plays an essential role in the regulation of chloroplast biosynthesis, photosynthetic capacity and yield. Moreover, leaf color mutant can be used as an effective marker for the purity of breeding and hybridization.     

水稻507ys黄绿叶突变体的遗传鉴定与候选基因分析

【目的】对水稻507ys黄绿叶突变体进行遗传鉴定与候选基因分析。【方法】用化学诱变剂甲基磺酸乙酯（EMS）处理粳稻品种日本晴（Nipponbare）,从突变体库中获得一份黄绿叶突变体507ys。对该突变体进行表型观察以及主要农艺性状调查分析。将507ys与正常绿色品种进行杂交,调查F1代的叶色表型和F2群体的叶色分离情况,分析该突变表型的遗传行为。利用（507ys/明恢63）的F2作为定位群体,对507ys突变基因进行精细定位且遴选候选基因,对候选基因进行DNA测序验证及编码蛋白序列同源性分析。同时,测定507ys突变体和野生型亲本的光合色素含量,并利用高效液相色谱（HPLC）精确分析它们的叶绿素组成成分,以进一步揭示507ys黄绿叶突变基因的候选基因。【结果】507ys黄绿叶突变体整个生育期呈黄绿色。与野生型亲本日本晴相比,507ys突变体在分蘖期叶片的叶绿素和类胡萝卜素含量分别下降52.1%和58.1%,成熟期株高、每株有效穗数、每穗着粒数和结实率分别减少8.3%、51.0%、7.4%和11.6%。507ys与正常绿色品种日本晴和明恢63杂交的F1表现正常的绿色,F2群体绿色正常植株与黄绿叶突变植株分离比符合3﹕1,表明507ys的黄绿叶突变性状由1对隐性核基因控制。该突变基因定位在第10染色体长臂近端部SSR标记RM333和InDel标记L3之间,遗传距离分别为0.56 cM和0.14 cM,两标记之间的物理距离约为60.2 kb,此区间内包含13个有注释的预测基因。基因组序列分析发现,507ys突变体中编码叶绿素酸酯a加氧酶的OsCAO1（LOC_Os10g41780）在编码区第2 198位碱基（CDS第1 057位碱基）处,碱基G突变为碱基A,造成编码蛋白的氨基酸序列第353位的谷氨酸（E）突变成赖氨酸（K）。对叶绿素组成成分分析表明,507ys突变体叶片中只有叶绿素a,没有叶绿素b。【结论】507ys突变体基因是已报道的叶绿素酸酯a加氧酶基因OsCAO1的等位基因。507ys突变体在OsCAO1外显子上发生的一个点突变使得其体内叶绿素酸酯a加氧酶失活,造成叶绿素b合成受阻。     

一个水稻卷叶突变体zw209的遗传分析与精细定位

叶片是植物光合作用的主要场所,优良的叶片形态有利于塑造理想株型,提高光合效率。为了研究叶片形态建成的分子机制,从水稻T-DNA突变体库中筛选到1个叶片极度卷曲的突变体zw209,突变体有2个显著特征:1突变体泡状细胞数量和面积均变小;2突变体的叶绿素含量较高,具有较高的光合效率。遗传分析表明,zw209的突变性状由一对隐性核基因控制。通过图位克隆,将基因(ZW209)精细定位到9号染色体长臂上,位于In Del136和In Del140之间的92.3 kb区域内。在这个区域内,尚未见报告已克隆的卷叶基因,很可能是一个新的基因位点。上述结果明确了突变体的表型特征及遗传规律,为克隆ZW209基因和揭示其作用机理奠定基础。     

水稻脆性突变体nbc(t)的主要特性和脆性基因的初步定位

对脆性突变体的表型、主要农艺性状和稻米品质进行了考查,并对脆性基因进行了初步定位。结果表明:脆茎水稻nbc(t)与野生亲本9311在株高等形态特征上无明显区别,仅在机械强度上有区别;nbc(t)的根、茎、叶、穗及种子都很脆,易折断,且表现全生育期脆性。脆茎突变体nbc(t)的主要农艺性状与9311相似,产量略低于9311,除了整精米率偏低以外,其他稻米品质性状与9311相似。脆茎突变体nbc(t)与广占63-4S所配组合的产量和稻米品质与扬两优6号相当。与9311相比,nbc(t)茎秆的抗折力相当,但抗张力明显降低,纤维素含量约降低17%。遗传分析表明,nbc(t)的脆茎特性可能由2个基因位点控制,初步定位在9号染色体上的2个区段,一个位于9号染色体上段SSR标记RM3700和RM24371之间,遗传距离分别为1.3cM和3.1cM;另一个位于9号染色体下段INDEL标记CL062和CL045外侧,遗传距离分别为1.6cM和6.0cM。     

水稻yl1黄叶突变体的基因克隆与功能分析

叶片是植物进行光合作用的重要器官,是作物产量形成的基础。作物叶色突变体不仅是研究叶绿体发育机制、培育高光效新种质的前提,而且可以作为性状标记,应用于良种繁育及杂交育种。在经化学诱变剂甲基磺酸乙酯(EMS)处理的粳稻品种Kitaake突变体库中,筛选得到黄叶突变体yl1,yl1突变体的株高、穗粒数、千粒重与野生型相比均降低。遗传分析表明,yl1黄叶表型由单隐型基因控制,图位克隆表明yl1编码叶绿素酸酯a氧化酶(chlorophyllide a oxygenase,OsCAO1,LOC_ Os10g41780)基因发生点突变;yl1突变体OsCAO1突变位点位于催化功能域,没有改变其在叶绿体中的定位,且在不同物种之间高度保守,这表明该位点可能是OsCAO1的酶活关键氨基酸位点。     

水稻类病斑突变体LM1基因的图位克隆

类病斑坏死突变体（lesion mimic mutant,LMM）是一类表型类似于植物过敏反应的突变体,对植物抵御外界病原菌浸染的抗病机理研究具有重有意义。以水稻突变体库中类病斑突变体lm1为材料,经图位克隆获得突变基因LM1,同时测定了相关生理指标。分析结果表明：与野生型相比,突变体叶片细胞中的丙二醛（MDA）积累水平明显升高,而叶绿素含量、Fv/Fm却显著低于野生型。遗传分析及精细定位显示,lm1突变体表型受一对隐性核基因LOC_Os12g16720控制,并将该基因命名为LM1,LM1位于水稻第12号染色体短臂的着丝粒附近,是SPL1的等位基因, 编码细胞色素P450单加氧酶;LM1突变导致其功能缺失,使植物体内活性氧得到累积,引发细胞程序性死亡并诱导病斑形成。     

水稻窄叶突变体Narrow leaf 11(nal11)的基因定位

[目的]定位和分析水稻窄叶突变体基因,为水稻叶片发育调控及株型育种提供参考依据.[方法]用甲基磺酸乙酯(EMS)诱导泸恢17,获得稳定的窄叶突变体(Narrow leaf 11,nal11),调查其与野生型泸恢17抽穗期功能叶的长和宽、分蘖数及成熟期株高.对nal11和绵恢727正反交获得的F2代进行遗传分析及基因定位.[结果]nal11抽穗期剑叶、倒2叶和倒3叶的宽度与野生型泸恢17存在显著(P<0.05,下同)或极显著(P<0.01,下同)差异,分别为野生型泸恢17的60.7%、57.9%和75.8%,但长度无显著差异(P>0.05);nal11株高为野生型泸恢17的90.3%,存在显著差异;nal11分蘖数极显著增加,为野生型的150.0%.nal11和绵恢727正反交后,F1代均表现正常叶宽,F2代叶宽发生性状分离,正常叶宽与窄叶植株数比例经χ2检验均符合3:1,表明nal11是受核单基因控制的隐性突变.利用SSR标记将nal11定位在水稻第4号染色体RM7290和RM16720标记之间约322 kb范围内,其与2个标记的遗传距离均为0.8 cM,覆盖了32.236 kb的物理区域,在定位区域内有5个注释基因,即Os04g26834、Os04g26850、Os04g26870、Os04g26880和Os04g26841,其序列与前人克隆的窄叶基因无重复.[结论]获得一个新的水稻窄叶突变体(nal11),其窄叶性状由1个隐性核基因控制,定位于水稻第4号染色体RM7290和RM16720标记之间,对功能叶、株高和分蘖数的表型有明显影响.     

Fine mapping of a novel wax crystal-sparse leaf3 gene in rice

Cuticular wax plays an important role in protecting plants against water loss and pathogen infection and in the adaptations to environmental stresses. The genetic mechanism of the biosynthesis and accumulation of epicuticular wax in rice remains largely unknown. Here,we show a spontaneous mutant displaying wax crystal-sparse leaves and decreased content of epicuticular wax that was derived from the cytoplasmic male sterility(CMS) restorer line Zhenhui 714. Compared with the wild type Zhenhui 714,the mutant exhibited hydrophilic features on leaf surface and more sensitivity to drought stress. The mutation also caused lower grain number per panicle and thousand grain weight,leading to the decline of yield. Genetic analysis indicates that the mutation is controlled by a single recessive gene,named wax crystal-sparse leaf3(wsl3). Using segregation populations derived from crosses of mutant/Zhendao 88 and mutant/Wuyujing 3,respectively,the wsl3 gene was fine-mapped to a 110-kb region between markers c3-16 and c3-22 on chromosome 3. According to the rice reference genome and gene analysis,we conclude that a novel gene/mechanism involved in regulation of rice cuticular wax formation.     

水稻温敏黄转绿突变体v5的鉴定和基因定位

通过对粳稻品种‘日本晴’进行60Coγ射线诱变,从M2中筛选到一个温度敏感的黄转绿突变体v5(virescent5)。突变体植株在较低温度(20～24℃)下叶色表现为黄色,而植株在较高温度(26～30℃)下叶色几乎表现为绿色。对叶绿素含量的测定显示出低温条件下突变体叶绿素含量显著低于野生型植株;对相关农艺性状考察显示出突变体单株产量显著低于野生型。遗传分析表明,该突变表型受一对隐性核基因控制,利用F2群体(v5/Kasalath)将OsV5基因定位于第9染色体长臂,在BAC AP005838上标记STS1与BAC AP005702上标记STS5312之间166.5 kb的区段上。     

两个新的水稻叶色突变体形态结构与遗传定位研究

【目的】对2个新的水稻叶色突变体进行形态结构与遗传分析,并且初步定位这2个突变基因。【方法】在水稻育种材料中分别发现了一株白色条纹叶突变体和一株黄叶突变体,经多代自交已形成稳定的突变系。对突变体的主要形态特征与叶绿素组分等进行分析,观察叶绿体的超微结构,并以这2个突变系杂交产生的F2群体作为定位群体,应用SSR标记对突变基因进行初定位。【结果】与其野生型相比,白色条纹叶突变体的单株穗数减少12.86%,生育期延长11.27%,黄色叶突变体的株高降低31.08%,千粒重减少14.55%,生育期延长17.86%,并且2种突变体的叶绿素含量都显著低于其野生型。电镜观察结果表明：2种突变体的类囊体结构异常,与野生型水稻相比,黄色叶突变体的类囊体片层数变少,白色条纹叶中条纹部分的类囊体片层结构几乎消失,正常绿色部分的类囊体结构没有变化。遗传分析表明：这2种突变性状均受1对隐性核基因控制,位于不同染色体上,将突变基因暂时命名为st9(t)(stripe)、chl12(t) (Chlorophyll-deficit)。将st9(t)定位到第一染色体短臂最末端,与分子标记RM1331相距9.6 cM,且与标记RM3252等共分离;将chl12(t)定位到第三染色体短臂,与分子标记RM411、RM8208之间的遗传距离分别是1.2、5.1 cM。【结论】发现了2个叶色突变新基因,为下一步的基因克隆与功能分析奠定了基础。     

陆地棉亚红株突变体的研究进展及育种应用前景

介绍了一个新的陆地棉色素突变体,即亚红株突变体,对该突变体的发现、遗传分析及分子标记研究进展进行了综述。分析了其与已在陆地棉中鉴定出与红色素沉着有关的3个基因位点R1、R2和Rd之间的关系,并提出了此突变体在育种上的应用前景,尤其是其性状在提高光合效能上的优势,为棉花实现高光效育种提供了一条途径,对棉花高产育种具有重要意义。     

一个新的水稻黄化突变体的光合作用及叶绿素荧光特性研究

对水稻黄化自然突变体(安农标810S)及其野生型(安农810S)的叶绿素含量、光合速率、叶绿素荧光动力学参数及单株产量进行了比较分析,结果显示:突变体在整个生长发育阶段总叶绿素含量均显著低于野生型,但突变体与野生型各自所配的杂交F1代叶绿素含量没有差异;从叶绿素a/b的比值来看,突变体的比值明显大于野生型;从净光合速率来看,无论是在1 d中的不同时间段,还是在不同发育时期,突变体均高于野生型;在不同发育时期和1 d中的不同时间段,突变体的FV/Fm、FV′/Fm′、φPS2、ETR2及QP均大于野生型,突变体的QN低于野生型;突变体自交繁殖的单株产量高于野生型,且从各自所配的杂交后代F1来看,突变体有较强的配合力。     

水稻白化转绿突变体研究进展

白化是水稻叶色突变中常见的叶绿素缺失突变,白化转绿突变体在光合作用机理、基因调控和遗传育种等理论研究和实际应用方面均有较大的利用价值。该文综述了近年来国内外有关水稻白化转绿突变体的研究进展,包括水稻白化转绿突变体的来源、白化转绿突变体与叶绿素合成的关系、影响白化的内外因素、相关基因的克隆以及作为标记性状在育种中的应用,为水稻白化转绿突变体的研究提供参考。     

一个水稻散生突变体tag1的遗传分析及基因定位

株型对水稻产量有着重要影响。作为影响株型的重要因素之一,分蘖角度是水稻高产育种上考虑的重要农艺性状。散生突变体tag1(tiller angle 1)在水稻品种台北309转基因后代中被发现。与野生型相比,主要突变表型为分蘖角度和叶夹角显著增大,并伴随株高、穗长、结实率和枝梗数等性状上的变化。遗传分析表明,tag1的突变性状由一对隐性基因控制。利用F2(tag1/明恢63)群体作为定位群体,突变基因被定位在第11染色体长臂In Del标记AC35795和M7之间,遗传距离分别为0.38 c M和0.95 c M。在该区间内存在一个已被克隆的控制水稻分蘖角度的基因LAZY1。测序比对分析发现,突变体的TAG1存在一个119 bp的缺失,包括第4个外显子50 bp,第4个内含子69 bp,及二者间的剪切位点。因此TAG1是LAZY1的一个新的等位基因,重新命名为LAZY1-2。lazy1和tag1在突变表型上存在一定差异,这可能是LAZY1基因序列的突变位点不同造成的。     

水稻卷叶突变体rlm1的基因克隆

叶片是植物光合作用的主要场所,优良的叶片形态有利于塑造理想的株型,提高光合效率。为了研究叶片形态建成的分子机制,自水稻T-DNA插入突变体库中筛选获得1个叶片半卷曲的卷叶突变体(roll leaf mutant,命名为rlm1),突变体rlm1主要特征为成熟叶片沿中脉向内卷,叶片最终卷成直立半圆筒状,叶片卷曲度达0.64,叶片直立参数达95,且光合效率显著优于野生型。通过图位克隆技术,确定突变体rlm1突变位点位于LOC_Os03g06654基因的第1个内含子,LOC_Os03g06654基因编码黄素单氧化酶(flavin-containing monooxygenase),RT-PCR表达分析表明LOC_Os03g06654基因因T-DNA插入而导致完全失活。该基因与已报道的水稻卷叶基因Os COW1(Constitutively Wilted 1)为等位基因,而且突变体rlm1所表现的农艺性状均佳,可期待利用该突变体进行高光合的育种实践。     

一个光敏色素B调控的水稻NBS-LRR基因的表达模式分析

利用水稻基因芯片比较野生型、phyA和phyB突变体受连续红光照射后的基因表达图谱,从中筛选到一个受光敏色素B(phyB)介导的红光信号特异诱导的、编码NBS-LRR类蛋白的基因,命名为PB-LRR(phyB-regulated NBS-LRR).为了揭示PB-LRR基因在水稻生长发育中的作用,对该基因表达的器官特异性...     

具有黄绿叶标记的香型籼稻不育系选育研究

【目的】转育成携带有标记性状的实用香型籼稻三系不育系及保持系。【方法】利用常规杂交自然分离突变体黄绿色叶片、有香味植株,转育成带有标记性状和香味的实用型不育系。【结果】选育的黄绿叶片突变体不育系其叶色自苗期至成熟期均表现为黄绿色,与正常叶色极易区别,并且未发现黄色退化现象。采用热水法鉴别具有香味的株系转育成带黄绿叶标记的香型籼稻不育系,其株高在41.5—56.5cm,植株叶片及谷粒颖壳呈黄绿色,其中叶鞘、叶耳及叶片边缘呈紫红色,分蘖力中等;其中黄标3A的穗长24.9cm,穗子着粒数145.0粒,谷粒呈长粒梭状形;千粒重28g,播始期85d,其保持系基部结实好,充实饱满。柱头呈紫黑色,柱头较大、而外露好。与恢复系测交、常规杂交的正、反交,其F1代均表现为正常绿色;测交、常规正、反交F2代群体平均分离比例分别为:2.92﹕1、2.76﹕1和2.96﹕1,经卡方检测,F2代群体分离符合3﹕1比例,表明黄绿色叶片呈隐性遗传,受1对隐性基因控制。【结论】黄绿叶标记不育系带有香味,且叶色与正常叶色有显著差异,受1对隐性基因控制。黄绿色叶片这一标记性状遗传稳定。与不同恢复系配组,F1代表现一般配合力强,可恢复性好。     

Quantitative analysis of the effect of the PAY1 gene on rice canopy structure during different reproductive stages

The leaf and stem types are core structural characteristics of the rice phenotype that determine the light interception ability of the canopy and directly affect crop yield. The PLANT ARCHITECTURE AND YIELD 1 (PAY1) gene has been shown to alter the prostrate growth habit of wild rice and to inhibit the wild rice prostrate growth gene PROSTRATE GROWTH 1 (PROG1). In this paper, the wild rice introgression line YIL55, which contains the PROG1 gene; its mutant, PAY1; and its parent, TQ, were used as test varieties to construct three-dimensional (3D) canopy structure models based on 3D digital assay technology. On this basis, quantitative analyses of the PAY1 gene and the plant leaf and stem types at the jointing, heading and filling stages were performed. Under the influence of the PAY1 gene, the plant stem and leaf angles from vertical decreased significantly; the plants were upright, with larger leaves; the culm angle changed from loose to compact; and the average tiller angle during the three key reproductive stages decreased from 44.9, 28.5 and 21.3° to 17.6, 8.4 and 10.5°, respectively. Moreover, the PAY1 mutant retained the PROG1 gene characteristic of exhibiting dynamic changes in the tiller angle throughout the growth period, and its culm angle changed from loose during the jointing stage to compact during the heading stage. The measurements of photosynthetically active radiation (PAR) in the canopy also showed that the mutant PAY1 allowed more PAR to reach the bottom of the canopy than the other varieties. The light-extinction coefficients for PAY1 at the jointing, heading and filling stages were 0.535, 0.312 and 0.586, respectively, which were lower than those of the other two varieties. In this study, the influence of the PAY1 gene on rice canopy structural characteristics was quantitatively analyzed to provide effective canopy structure parameters for breeding the ideal plant type.