水稻耐盐生理及分子机制研究进展

随着全球气候变化和土地盐碱化问题的加剧,提高水稻(Oryza sativa L.)在盐碱地环境下的生长能力成为了农业生产的一个关键挑战。“以种适地”策略的实现需要深入理解水稻的耐盐机制,并在此基础上进行育种改良。本研究总结了近年来关于水稻耐盐调控基因的研究成果,并依据其参与的生物学过程进行了功能性分类。详细分析了水稻对盐胁迫的感知、以及随后激活的多种生理调节机制,包括渗透调节、离子稳态、抗氧化防御系统和养分平衡等。重点讨论了水稻中几个关键的盐胁迫信号途径,包括SOS(Salt Overly Sensitive)途径、MAPK(Mitogen-Activated Protein Kinase)级联途径以及激素调节途径,这些途径在水稻适应盐胁迫环境中起着至关重要的作用。通过综述现有文献资料,本文旨在提供一个全面的水稻耐盐性调控基因及其功能的概览,为水稻耐盐育种工作提供科学依据,同时为提高水稻在盐碱地环境下的产量和质量提供参考。

盐碱地的生物修复方法

中国盐碱地面积占全球盐碱地总面积的1/10,严重制约了农业生产的发展。因此,寻找有效的盐碱地修复方法对于提高农业产量和保障粮食安全具有重要意义。目前,盐碱地修复的方法有化学修复、工程修复和生物修复等,而生物修复以其经济高效和绿色环保的特点受到了广泛关注。生物修复通过植物和微生物等生物资源来改善盐碱地的土壤和环境。具体而言,盐生植物能够在盐碱地中生长繁殖,通过其特殊的生理机制,能够有效修复高浓度盐碱地,为后续作物种植创造良好条件。此外,普通作物也可以通过基因挖掘和品种培育的方式,增强其耐盐性,从而在盐碱地上实现正常生长,达到修复盐碱地的目的。除了植物,微生物也在盐碱地修复中发挥着重要作用。一些特定的微生物能够提高作物的耐盐性,或者通过降解盐碱土壤中的盐分和碱性物质,从而降低土壤的盐碱度,改善土壤环境。总的来说,生物修复是一种绿色环保,且符合中国可持续农业和循环经济发展战略的盐碱地修复方法。

水稻斑马叶突变体zl7的鉴定与基因的精细定位

【目的】斑马叶突变体作为水稻叶色突变体的重要种质资源,是研究植物光合作用机制和高光效育种的理想材料,对于解析光合作用机理和提高水稻产量具有重要意义。【方法】用甲基磺酸乙酯(EMS)诱变粳稻品种春江06建立突变体库,从突变体库中筛选到1份苗期为斑马叶的突变体,该突变体被命名为zl7(zebra leaf7)。在常规大田种植条件下分别比较突变体与野生型在苗期、抽穗期和成熟期叶色表型和产量性状差异,通过透射电镜实验分析叶片叶绿体发育情况,利用图位克隆方法克隆候选基因,利用荧光定量PCR方法分析参与叶绿素合成和叶绿体发育相关基因的表达水平。【结果】从苗期开始,突变体zl7表现出典型的斑马叶,叶绿素含量降低,直到抽穗期,斑马叶表型消失,叶片逐渐复绿,叶绿素含量无明显差异。光合速率测定和电镜观察结果显示,突变体zl7的光合速率、气孔导度下降,叶绿体发育异常。与野生型相比,突变体的株高、分蘖、穗长、一次枝梗、二次枝梗和每穗粒数均显著降低,而粒长、粒宽和千粒重均略有增加。荧光定量PCR结果表明突变体中参与叶绿素降解相关基因的表达量显著升高,而参与叶绿素合成和叶绿体发育相关基因的表达量显著降低。遗传分析表明,该突变体受一对隐性核基因调控。通过图位克隆将该基因定位在第7染色体,测序发现突变体的目标基因ZL7编码区发生单碱基替换,导致一个氨基酸由丝氨酸变为天冬酰胺。【结论】ZL7(Zebra Leaf 7)突变导致叶绿体发育异常,水稻叶片出现斑马叶表型,该基因在叶绿素合成及叶绿体发育中起重要作用。

Chlorophyllide-a Oxygenase 1(OsCAO1) Over-Expression Affects Rice Photosynthetic Rate and Grain Yield

Leaf color and photosynthesis are important factors for rice growth and development.Hence,improving the photosynthetic rate is an effective approach for increasing rice yield.We isolated a gene,chlorophyllide-a oxygenase 1(OsCAO1),which characterized a rice near-isogenic line named fgl(faded green leaf).

水稻窄叶突变体nal12的鉴定与基因精细定位

【目的】水稻叶片是理想株型的主要组成部分。筛选和鉴定新的叶形突变材料,可为研究叶发育调控机制和塑造叶片理想形态打下基础。【方法】由粳稻品种武运粳31经EMS(ethyl methane sulphonate)诱变获得窄叶突变体nal12(narrow leaf 12);通过表型测量分析剑叶至倒4叶的叶片长度、宽度、大脉数和小脉数,并进行组织切片和显微观察;将突变体nal12与籼稻品种台中本地1号(TN1)杂交,在F_2分离群体中选取了1709个具窄叶突变表型的单株,通过已有的SSR、STS和新开发的分子标记进行精细定位。【结果】nal12在幼苗期就表现出了窄叶性状,在成熟期植株较野生型矮小,分蘖增多,茎秆变细。经组织切片观察分析,nal12叶片变窄是由于大脉和小脉数量减少造成。遗传分析表明该窄叶表型受单一隐性核基因调控;最终将该基因定位在第10染色体上LC2-RF37与LC4R-RF39标记之间约64.7 kb的范围内。该区间内共有10个开放阅读框,其中并无已报道的窄叶相关基因。qRT-PCR结果表明,NAL12基因的突变会影响生长素合成与运输相关基因的表达。【结论】NAL12为一个新的叶形调控基因。这为进一步研究水稻叶片发育,丰富其分子调控网络打下了基础。

水稻淡绿叶基因PGL11的鉴定与精细定位

【目的】叶片是水稻进行光合作用的主要场所,叶片颜色的变化与水稻的生长发育直接相关。发掘水稻叶色突变体,是水稻功能基因组学研究的重要遗传基础。【方法】利用EMS诱变日本晴获得一个能稳定遗传的淡绿叶突变体,暂命名为pgl11(pale green leaf 11)。在不同生育期测定野生型与突变体的叶绿素含量。在苗期,取野生型与突变体叶片进行叶绿体结构的透射电镜观察。在分蘖期,测定野生型与突变体的光合参数并观察气孔结构。在成熟期,测定野生型和pgl11的主要农艺性状。以pgl11为母本,南京6号为父本构建相应的F2群体,采用图位克隆的方法,对该基因进行定位。【结果】从苗期开始,突变体pgl11的每一片新叶均表现为淡绿色,叶绿素含量显著降低,叶绿体发育异常。随着叶片的生长,叶色由淡绿逐渐转绿,至抽穗期时叶绿素含量亦无明显差异。pgl11还表现光合速率、气孔导度明显下降,胞间CO_2浓度上升。扫描电镜观察发现,突变体pgl11的气孔发育异常。与野生型相比,突变体的农艺性状如株高、剑叶宽、二次枝梗数、每穗粒数、粒长、粒宽、千粒重以及结实率等均显著降低。对叶绿素合成、光合作用以及质体发育相关基因的表达量测定表明,突变体pgl11中参与叶绿体转录和翻译相关基因的表达量显著升高,而叶绿素合成和光合作用相关基因的表达量显著下降。遗传分析表明,该突变表型受一对隐性核基因控制。通过图位克隆的方法将该基因定位于第1染色体上的C6和C8标记之间,物理距离约为110 kb。【结论】该定位区间内未见有叶色相关基因报道,推测PGL11基因可能是一个新的水稻叶色基因。

籼稻C84和粳稻春江16B重组自交系遗传图谱构建及籽粒性状QTL定位与验证

【目的】本研究旨在挖掘水稻粒型新基因、探索其分子机理,解析籽粒发育调控遗传网络奠定基础,并为通过分子标记聚合有利基因开展超级稻分子设计育种提供理论依据。【方法】以植株和籽粒形态差异较大的晚粳稻品种春江16B(CJ16B)和广亲和中籼稻背景恢复系C84为亲本构建含有188个家系的重组自交系为作图群体,利用158对在双亲中存在多态性差异的分子标记,构建了遗传连锁图谱,总遗传距离为1428.40 cM,平均标记间距为9.04 cM。在构建遗传图谱的基础上,完成RIL188个株系籽粒的粒长、粒宽、粒厚、长宽比和千粒重等5个性状考查并进行QTL定位。【结果】在海南陵水和浙江杭州两地共检测到籽粒相关主效QTL 30个,包括籽粒QTL新座位18个,解释遗传变异3.51%~17.25%。其中粒长、粒宽、粒厚和长宽比QTL位点分别为9个、5个、5个和6个,千粒重QTL位点5个。经基因座位比对,发现有5个QTL区间与已克隆的调控籽粒形态相关基因座位相近,我们通过对双亲目标基因的测序并根据差异位点设计d CAPs分子标记进行验证。【结论】该RIL群体及其遗传图谱可用于水稻重要农艺性状主效QTL基因的定位和克隆,新定位的18个粒型QTL可以为水稻籽粒发育调控网络提供补充和资料积累。

Progress on Molecular Mechanism of Aluminum Resistance in Rice

Aluminum(Al)toxicity in acid soils is a significant limitation to crop production worldwide,as 13%of the world's rice is produced in acid soil with high Al content.Rice is likely the most Al-resistant cereal and also the cereal,where Al resistance is the most genetically complex with external detoxification and internal tolerance.Many Al-resistance genes in rice have been cloned,including Al resistance transcription factor 1(ART1)and other transcription factors,organic acid transporter genes,and metal ion transporter gene.This review summarized the recent characterized genes affecting Al tolerance in rice and the interrelationships between Al and other plant nutrients.

Rapid Creation of New Photoperiod-/Thermo-Sensitive Genic Male-Sterile Rice Materials by CRISPR/Cas9 System

Rice is an important food crop in China, and the development of hybrid rice is a crucial way to increase grain yield. The creation of dual-purpose nuclear-sterile lines for two-line hybrid breeding has become vital for commercial rice breeding. We constructed the pC1300-2 x35 S::Cas9-sgRNA^(PTGMS2-1) expression vector for editing the male fertility gene PTGMS2-1 in two widely compatible rice varieties, 93-11 and Huazhan, by using the CRISPR/Cas9 system. We obtained the marker-free photoperiod-/thermo-sensitive genic male-sterile(P/TGMS) lines in T_1 generation. According to the experiments in phytotron with four temperature and photoperiod treatments, we found the temperature is the main factor for restoring the pollen fertility of ptgms2-1 mutants in 93-11 and Huazhan, and the photoperiod also has some effects on pollen fertility in two different rice backgrounds. The application of cultivating new male-sterile lines by genome editing system will significantly accelerate the rice breeding process.

Coleoptile Purple Line Regulated by A-P Gene System Is a Valuable Marker Trait for Seed Purity Identification in Hybrid Rice

In plants,a large number of anthocyanin biosynthetic genes encoding enzymes and regulatory genes encoding transcription factors are required for anthocyanin synthesis.Coleoptile purple lines are two purple lines on both sides of coleoptiles after seed germination.However,the molecular mechanism of coleoptile purple line is not clear in rice so far.In this study,two major dominant genes,coleoptile purple line 1(OsCPL1,also known as OsC1)and coleoptile purple line 2(OsCPL2),were isolated via map-based cloning,and both of them were required for anthocyanin biosynthesis of coleoptile purple line in rice.The knockout and complementation experiments confirmed that OsC1 was required for purple color in most organs,such as coleoptile line,sheath,auricle,stigma and apiculus,whereas OsCPL2 was just required for coleoptile purple line.OsC1 was predominantly expressed in coleoptiles,flag leaves,and green panicles,and highly expressed in young leaves,whereas OsCPL2 was predominantly expressed in coleoptiles,and extremely lowly expressed in the other tested organs.Loss-of-function of either OsC1 or OsCPL2 resulted in significant reduction of transcript levels of multiple anthocyanin biosynthesis genes in coleoptiles.Coleoptile purple line was further used as a marker trait in hybrid rice.Purity identification in hybrid rice seeds via coleoptile purple line just needed a little water,soil and a small plate and could be completed within 5 d.Molecular marker and field identification analyses indicated that coleoptile purple line was reliable for the hybrid seed purity identification.Our findings disclosed that coleoptile purple line in rice was regulated by two major dominant genes,OsC1 and OsCPL2,and can be used as a simple,rapid,accurate and economic marker trait for seed purity identification in hybrid rice.

水稻粒宽突变体gw4的鉴定与基因定位

【目的】水稻粒形是影响水稻产量和决定稻米外观品质的主要性状之一。筛选和鉴定新的粒形突变材料,可为研究水稻籽粒发育的调控机制奠定基础。【方法】粳稻品种中花11经1%的EMS处理,在诱变群体中获得一份窄粒突变体gw4(grain width on chromosome 4);分析粒形和其他主要农艺性状,在扫描电镜下观察颖壳细胞变化;利用突变体与籼稻品种台中本地1号配组的F2分离群体,选择隐性个体完成基因的精细定位;开展生物信息和测序分析,确定定位区间的候选基因;采用RT-PCR分析该基因在根、茎、叶、鞘、穗等组织中的表达模式及其他粒形相关基因的表达水平。【结果】与野生型相比,除了表现窄粒外,gw4的粒长、千粒重、每穗粒数、一次枝梗数和二次枝梗数等显著下降;扫描电镜发现gw4的颖壳内外表皮细胞均小于野生型;遗传分析表明该窄粒表型受一对单隐性核基因控制;通过开发的新标记最终将该基因定位在第4染色体BS6与EX49两个标记之间约31.74 kb的范围内;测序结果发现在LOC_Os04g01590基因编码区发生了一个由G至A的单碱基突变,导致原来编码的甘氨酸变成了天冬氨酸;qRT-PCR结果表明,LOC_Os04g01590主要在幼穗中表达,且在突变体中表达显著下降。【结论】GW4主要调控水稻粒宽的发育,预测LOC_Os04g01590为其候选基因。这为进一步丰富粒形的遗传调控网络打下了基础。

Genome-Wide Association Analysis and Allelic Mining of Grain Shape-Related Traits in Rice

Excavating single nucleotide polymorphisms (SNPs) significantly associated with rice grain shape and predicting candidate genes through genome-wide association study (GWAS) can provide a theoretical basis for discovery and utilization of excellent genetic resources in rice. Based on 16 352 SNPs, 161 natural indica rice varieties with various grain sizes in southern China were used for GWAS of grain shape-related traits, referring to grain length (GL), grain width (GW), 1000-grain weight (TGW), and grain length/width (GLW). Phenotypic statistics showed that coefficient of variation values for these four traits GL, GW, TGW and GLW were 9.92%, 9.09%, 20.20% and 16.38%, respectively. Each trait showed a normal distribution, and there was a certain correlation between these traits. Through general linear model correlation analysis, a total of 38 significant loci were identified, and a range of 100 kb upstream and downstream of the significant loci was identified as the candidate interval. On chromosome 3, GS3 and qGL3 were found to regulate GL. On chromosome 6, TGW6 and GW6a were found to regulate TGW. Also, some QTLs related to grain shape were found on chromosomes 5 and 9. Besides that, using sequenced 3K-germplasm resources, we found that there are 22 overlapped varieties between these two natural populations. Twenty-six SNPs and fourteen haplotypes were identified in five regions of GS3 genes. The detection of multiple candidate genes/QTLs within the candidate interval is beneficial for further excavation of superior rice genetic resources.

Functional Analysis of Three Rice Chloroplast Transit Peptides

Chloroplast transit peptides(CTPs) can be used to transport non-chloroplastic proteins into the chloroplasts. Here, we studied the CTPs of three rice(Oryza sativa L.) chloroplast-localized proteins and found that their CTPs could be used to transport non-chloroplast-localized proteins into the chloroplasts. Fusion proteins lacking the CTP remained located in the cytoplasm. Furthermore, we constructed green fluorescent protein fusion vectors with the three CTPs and three non-chloroplast-localized proteins, Ghd10, MULTI-FLORET SPIKELET1(MFS1), and SHORTENED UPPERMOST INTERNODE 1(SUI1). After transforming these constructs into rice protoplasts, the fusion proteins all localized in the chloroplasts. Collectively, our results showed that these CTPs can transport non-chloroplast-localized proteins into the chloroplasts, and more importantly, these CTPs can be applied to engineer chloroplast metabolism.

Identification of QTLs for Cadmium Tolerance During Seedling Stage and Validation of qCDSL1 in Rice

Cadmium(Cd)is a non-essential toxic metal that is harmful to plants.To investigate the genetic mechanism of Cd tolerance in rice,quantitative trait loci(QTLs)associated with Cd tolerance at the seedling stage were analyzed using a recombinant inbred line(RIL)population derived from a cross between PA64s and 93-11.A total of 36 QTLs associated with shoot length,root length,shoot dry weight,root dry weight and total dry weight were detected in Hangzhou and Lingshui of China.Among them,15 QTLs were identified under the control condition and 15 QTLs were identified under the Cd stress condition,and 6 QTLs for Cd tolerant coefficient were detected on chromosomes 1,3,7 and 9.The qCDSL1.1 and qCDSL1.2 were identified in Hangzhou and Lingshui,respectively,and had overlapping intervals on chromosome 1.To further confirm the effects of qCDSL1.1 and qCDSL1.2,we developed a chromosome segment substitution line(CSSL),CSSLqCDSL1,in 93-11 background harboring qCDSL1.1/qCDSL1.2 from PA64s.Compared to 93-11,CSSLqCDSL1 had increased shoot length under the Cd stress condition.These results pave the way for further isolation of those genes controlling Cd tolerance in rice and marker-assistant selection of rice elite varieties with Cd tolerance.

A Recessive Mutant of argonaute 1b/gsnl4 Leads to Narrow Leaf, Small Grain Size and Low Seed Setting in Rice

A gsnl4 mutant characterized by small grain size,narrow leaf and low seed-setting rate was obtained by ethyl methane sulfonate(EMS)mutagenesis of a japonica rice variety Wuyunjing 21.Genetic analysis showed that gsnl4 is a loss-of-function mutant.A single-base mutation in gsnl4 resulted in the substitution of Ser to Asn in the Piwi domain of OsAGOlb protein.CRISPR/Cas9-mediated editing of OsAGOlb yielded a mutant phenotypically resembling gsnl4.Furthermore,miRNA-Seq analysis showed that the transcript expression levels of miRNAs in the signal transduction pathways related to pollen development,leaf morphology and honnone activation were significantly different between the gsnl4 mutant and the wild type(WT)plants.Several miRNAs were downregulated,and their target genes were upregulated in gsnl4 mutants.The auxin content in the root tips of the gsnl4 mutant decreased,and the expression of most auxin-related genes was altered.In summary,GSNL4 not only regulates organ development by controlling cell division and expansion,but also plays an important role in regulating auxin transport in rice.

Identification of a novel tillering dwarf mutant and fine mapping of the TDDL(T) gene in rice (Oryza sativa L.)

Rice plant architecture is an important agronomic trait that affects the grain yield. To understand the molecular mechanism that controls plant architecture, a tillering dwarf mutant with darker-green leaves derived from an indica cultivar IR64 treated with EMS is characterized. The mutant, designated as tddl(t), is nonallelic to the known tillering dwarf mutants. It is controlled by one recessive nuclear gene, TDDL(T), and grouped into the dn-type dwarfism according to Takeda's definition. The dwarfism of the mutant is independent of gibberellic acid based on the analyses of two GA-mediated processes. The independence of brassinosteroid (BR) and naphthal-3-acetic acid (NAA) of the tddl(t) mutant, together with the decreased size of parenchyma cells in the vascular bundle, indicates that the TDDL(T) gene might participate in another hormone pathway. TDDL(T) is fine mapped within an 85.51 kb region on the long arm of rice chromosome 4, where 20 ORFs are predicted by RiceGAAS (http://ricegaas.dna.affrc. go.jp/rgadb/). Further cloning of TDDL(T) will benefit both marker assisted selection (MAS) of plant architecture and dissection of the molecular mechanism underlying tillering dwarf in rice.