Morphological and Molecular Characterization of Novel Salt-Tolerant Rice Germplasms from the Philippines and Bangladesh

To screen for new sources of salinity tolerance, 688 traditional rice varieties from the Philippines and Bangladesh were obtained, and their tolerance to hypersaline conditions at the seedling stage was examined. A total of 29 Philippine lines and 15 Bangladeshi lines were scored as salt-tolerant.Morphological assessment(plant height, biomass and Na-K ratio) revealed that among the 44 salt-tolerant accessions, Casibon, Kalagnon and Ikogan had significantly higher relative shoot length difference, relative shoot growth reduction and shoot Na-K ratio than the tolerant check FL478.Additionally, AC and Akundo exhibited significantly higher Na-K ratios than the other genotypes. The genetic diversity of the 44 genotypes was assessed using 34 simple sequence repeat markers. A total of 133 alleles were detected across all loci. Cluster analysis showed that AC, Akundo and Kuplod were clustered along with FL478, indicating a strong genetic relatedness between these genotypes. IR29(susceptible check) was singly separated. The haplotype analysis revealed that none of the 44 genotypes had a similar allele combination as FL478. These accessions are of interest since each genotype might be different from the classical salinity-tolerant Pokkali.     

Differential Sensitivity of Rice Cultivars to Salinity and Its Relation to Ion Accumulation and Root Tip Structure

Abstract

Effects of NaCl on the growth, ion content, root cap structure and Casparian band development were examined in four rice (Oryza sativa L.) cultivars with different salt resistance (salt-sensitive indica-type IR 24 and japonica-type Nipponbare and salt-resistant indica-type Nona Bokra and Pokkali). Experiments were conducted to find the differences in salinity resistance during early seedling and developed seedling stages among the cultivars. For salinity treatment, sodium chloride (NaCl) was added to nutrient solution at concentrations of 0, 25 and 50 mM for 7 days from germination to the 7th day (early seedling stage) or from the 7th day to 14th day (developed seedling stage). Growth inhibition by salinity was more prominent in the early seedling stage than in the developed seedling stage. Based on the growth, the order of the sensitivity was IR24 > Nipponbare > Nona Bokra > Pokkali. The growth of NaCl-treated rice cultivars relative to control was significantly and negatively correlated with the Na⁺ content and Na⁺/K⁺ ratio in roots and shoots in both stages. Scanning electron microscopic observation revealed that the root cap tissues proliferated and extended to the basal part of the root tip by salinity. The length of root cap was, however, reduced by 50 mM NaCl in sensitive cultivars due to peeling off. An endodermal Casparian band was formed in the basal region of the root tip. Development of the Casparian band was more prominent in sensitive cultivars than in tolerant cultivars. Root cap proliferation might be related to NaCl resistance in rice seedlings, but the Casparian band may not function efficiently in Na⁺ exclusion. Essentially the present results suggest that exclusion of Na⁺ from roots plays a critical role in expression of Na⁺ resistance in rice seedlings and the root cap is important for Na⁺ exclusion.     

Haplotyping of Rice Genotypes Using Simple Sequence Repeat Markers Associated with Salt Tolerance

Salt stress is a major problem in most of the rice growing areas in the world. A major QTL Saltol associated with salt tolerance at the seedling stage has been mapped on chromosome 1 in rice. This study aimed to characterize the haplotype diversity at Saltol and additional QTLs associated with salt tolerance. Salt tolerance at the seedling stage was assessed in 54 rice genotypes in the scale of 1 to 9 score at EC = 10 d Sm-1 under controlled environmental conditions. Seven new breeding lines including three KMR3/O. rufipogon introgression lines showed similar salt tolerant ability as FL478 and can be good sources of new genes/alleles for salt tolerance. Simple sequence repeat(SSR) marker RM289 showed only two alleles and RM8094 showed seven alleles. Polymorphic information content value varied from 0.55 for RM289 to 0.99 for RM8094 and RM493. Based on 14 SSR markers, the 54 lines were clearly separated into two major clusters. Fourteen haplotypes were identified based on Saltol linked markers with FL478 as the reference. Alleles of RM8094 and RM3412 can discriminate between the salt tolerant and susceptible genotypes clearly and hence can be useful in marker-assisted selection at the seedling stage. Other markers RM10720 on chromosome 1 and RM149 and RM264 on chromosome 8 can also distinguish tolerant and susceptible lines but with lesser stringency.     

Effects of Soil Salinity and Alkalinity on Grain Quality of Tolerant,Semi-Tolerant and Sensitive Rice Genotypes

Soil salinity and alkalinity adversely affects the productivity and grain quality of rice.The grain quality of 19 rice genotypes characterized as salt tolerant (T),semi-tolerant (ST) and sensitive (S) was assessed in lysimeters containing saline and highly alkaline soils.Head rice recovery was reduced by salinity stress whereas it was not affected by alkalinity stress.The ratio of length to width (grain dimensions) was significantly reduced in the T genotype even at low electrical conductivity (EC,4 mS/cm) and alkalinity (pH 9.5),whereas in the ST genotype,it was significantly reduced at high salinity (EC 8 mS/cm).There was no significant effect of any levels of salinity or alkalinity on grain dimensions in the S genotype.Amylose content was significantly reduced in T and ST groups even at low EC (4 mS/cm) and alkalinity (pH 9.5) and the effect in the S genotype was only at high salinity.Starch content showed significant reduction at high salinity and alkalinity (EC 8 mS/cm and pH 9.8) in the T and ST genotypes and no significant effect was observed in the S genotype.The effect of both levels of salinity (EC 4 and 8 mS/cm) and high alkalinity (pH 9.8) on gel consistency was observed only in the S genotype.The tolerant genotypes IR36 under high salinity,and CSR10 and CSR11 under alkali stress showed less reduction in amylose content.The T genotype BR4-10,and ST genotypes CSR30,CSR29 and CSR13 showed better gel consistency under saline and alkali stress.Amylose content was affected even at low salinity stress and thus important to be considered in breeding rice for salt tolerance.Overall,the grain quality of T and ST genotypes was less affected by saline and alkali stress compared to S ones.     

水稻耐盐性数量性状位点的初步检测

用RFLP分析技术和分蘖株系法对由耐盐性品种Pokkali和盐敏感品种Peta配制的BC1(Peta/Pokkali∥Peta)群体分别检测水稻苗期和成熟期耐盐性数量性状位点(QTLs)。表型鉴定在含(处理)或不含(对照)60 mol/m3 NaCl的营养液中进行,苗期观测盐害级别、苗Na+含量和鲜重/干重比值3项指标,成熟期测定10种农艺性状处理与对照的相对值。从水稻12条染色体上筛选出43个多态性标记,对上述指标分别作点分析,共检出15个连锁标记。连锁标记的分布特点显示,在研究所涉及的基因组范围内存在4个影响苗期耐盐性的QTL,其增效等位基因均来自耐盐品种Pokkali;影响成熟期耐盐性的QTL分布于7条染色体的1或2个连锁区间上,其有利基因来自双亲;RG678和RZ400B~RZ792附近的2个QTL在全生育期都能表达较强耐盐性。     

Responses of Lowland,Upland and Aerobic Rice Genotypes to Water Limitation During Different Phases

Rice yield reduction due to water limitation depends on its severity and duration and on the phenological stage of its occurrence. We exposed three contrasting rice genotypes, IR64, UPLRi7 and Apo (adapted to lowland, upland and aerobic conditions, respectively), to three water regimes (puddle, 100% and 60% field capacity) in pots during the vegetative (GSI), flowering (GSII) and grain filling (GSIII) stages. Stress at all the three stages significantly reduced yield especially in lowland genotype IR64. Effect of water limitation was more severe at GSII than at the other two stages. Stress at GSI stage reduced both source activity (leaf area and photosynthetic rate) and sink capacity (tiller number or panicle number per pot). When stress was imposed at GSII, spikelet fertility was most affected in all the three genotypes. In both GSII and GSIII, although leaf area was constant in all the three water regimes, estimated relative whole-plant photosynthesis was strongly associated with yield reduction. Reduced photosynthesis due to stress at any given stage was found to have direct impact on yield. Compared to the other genotypes, Apo had deeper roots and maintained a better water relation, thus, higher carbon gain and spikelet viability, and ultimately, higher biomass and productivity under water-limited conditions. Therefore, screening for these stage-dependent adaptive mechanisms is crucial in breeding for sustained rice production under water limitation.     

不同水稻基因型地上部钾素累积和转运规律的研究

 经水培苗期筛选后选用对低钾胁迫抗性不同的9种水稻基因型进行田间试验，结果表明, 低钾胁迫减少水稻体内钾的累积量，增加体内钾的运转速率。不同钾处理间水稻地上部钾素累积量和同一处理内不同水稻基因型间吸钾能力在生育前期差异均较大，后期则相对较小。水稻体内钾累积与运转能力的基因型差异非常显著。与钾低效基因型相比，钾高效基因型低钾胁迫下苗期地上部钾素累积量较大，孕穗期上位叶和下位叶中钾含量均相对较高，上位叶、下位叶间钾含量的差异也较大。     

不同水稻基因型地上部钾素累积和转运规律的研究（英文）

经水培苗期筛选后选用对低钾胁迫抗性不同的 9种水稻基因型进行田间试验 ,结果表明 ,低钾胁迫减少水稻体内钾的累积量 ,增加体内钾的运转速率。不同钾处理间水稻地上部钾素累积量和同一处理内不同水稻基因型间吸钾能力在生育前期差异均较大 ,后期则相对较小。水稻体内钾累积与运转能力的基因型差异非常显著。与钾低效基因型相比 ,钾高效基因型低钾胁迫下苗期地上部钾素累积量较大 ,孕穗期上位叶和下位叶中钾含量均相对较高 ,上位叶、下位叶间钾含量的差异也较大。     

低磷胁迫下不同基因型水稻阶段性磷营养特征

采用盆栽试验研究了低磷胁迫条件下不同磷效率基因型水稻在分蘖期、孕穗期和成熟期的磷营养特征。在分蘖期,耐低磷基因型水稻吸磷量无论在何种磷处理条件下均显著高于低磷敏感基因型,低磷胁迫时,除99112T外其他耐低磷基因型水稻的磷利用效率也显著高于低磷敏感基因型;到孕穗期,耐低磷基因型水稻在低磷胁迫时仍具有较高的吸磷量,但是磷利用效率与低磷敏感基因型相比却没有明显差异;至成熟期,耐低磷基因型水稻的磷效率的贡献因子依次为：磷利用效率、吸收效率和转运效率。较低的吸磷量是99012S（低磷敏感基因型）在不同生育时期磷效率低下的主因,在生育前期磷吸收和利用效率低下及后期磷利用效率低下是另一个低磷敏感基因型99056S磷效率低下的主要原因。低磷胁迫还滞后了低磷敏感基因型的磷吸收,并随着低磷胁迫的加剧而增强。     

Identification of Rice Accessions Associated with K~+/Na~+ Ratio and Salt Tolerance Based on Physiological and Molecular Responses

The key for rice plant survival under Na Cl salt stress is maintaining a high K~+/Na~+ ratio in its cells. Selection for salt tolerance rice genotypes based on phenotypic performance alone will delay in progress in breeding. Use of molecular markers in tandem with physiological studies will help in better identification of salt tolerant rice accessions. Eight rice accessions along with the check Dongjin were screened using 1/2 Yoshida solution with 50 mmol/L NaCl at the seedling stage. The accessions IT001158, IT246674, IT260533 and IT291341 were classified as salt tolerant based on their K~+/Na~+ ratios. Seventeen SSR markers reported to be associated with K~+/Na~+ ratio were used to screen the accessions. Five SSR markers(RM8053, RM345, RM318, RM253 and RM7075) could differentiate accessions classified based on their K~+/Na~+ ratios. Banding pattern of the accessions was scored compared to the banding pattern of Dongjin. The study differentiated accessions based on their association of K~+/Na~+ ratio with molecular markers which are very reliable. These markers can play a significant role in screening large set of rice germplasms for salt tolerance and also help in identification of high-yielding varieties with better salt tolerance. The salt tolerant accessions can be taken forward into developing better varieties by conventional breeding and exploring genes for salt tolerance.     

盐胁迫对杂交稻隆两优华占及其亲本种子萌发和幼苗生长的影响

以杂交稻组合隆两优华占及其2个亲本隆科638S和华占为试验材料,萌发期设置6个NaCl质量分数梯度(0.2%、0.4%、0.6%、0.8%、1.0%和1.2%)和苗期设置3个NaCl质量分数梯度(0.4%、0.8%和1.2%)来模拟盐胁迫试验,以清水为对照,分析盐胁迫对杂交稻及其亲本种子萌发和幼苗生长的影响。结果表明,在各质量分数的盐胁迫处理下,萌发期各项指标均不同程度地受到盐胁迫影响,多数指标值均随盐质量分数升高而下降,且质量分数0.6%是隆两优华占及其亲本的耐盐临界值。在0.2%~0.6%NaCl胁迫下3个品种发芽率、发芽指数、活力指数、胚芽长的盐胁迫抑制率在品种间存在显著差异,进一步通过隶属函数值综合评价,得出3个品种萌发期耐盐性由强到弱分别为隆两优华占、华占和隆科638S;苗期盐胁迫下3个品种中隆两优华占地上部分K^+/Na^+比值最大,表明其耐盐性最强。综合萌发期隶属函数值和苗期K^+/Na^+比值可以得出,杂交组合隆两优华占与其亲本比较,耐盐性更强,说明水稻的耐盐性具有一定的杂种优势。     

Characterizing the Saltol Quantitative Trait Locus for Salinity Tolerance in Rice

This study characterized Pokkali-derived quantitative trait loci (QTLs) for seedling stage salinity tolerance in preparation for use in marker-assisted breeding. An analysis of 100 SSR markers on 140 IR29/Pokkali recombinant inbred lines (RILs) confirmed the location of the Saltol QTL on chromosome 1 and identified additional QTLs associated with tolerance. Analysis of a series of backcross lines and near-isogenic lines (NILs) developed to better characterize the effect of the Saltol locus revealed that Saltol mainly acted to control shoot Na⁺/K⁺ homeostasis. Multiple QTLs were required to acquire a high level of tolerance. Unexpectedly, multiple Pokkali alleles at Saltol were detected within the RIL population and between backcross lines, and representative lines were compared with seven Pokkali accessions to better characterize this allelic variation. Thus, while the Saltol locus presents a complex scenario, it provides an opportunity for marker-assisted backcrossing to improve salt tolerance of popular varieties followed by targeting multiple loci through QTL pyramiding for areas with higher salt stress.     

Antioxidant Defense Mechanisms of Salinity Tolerance in Rice Genotypes

In order to elucidate the role of antioxidant responses in salinity tolerance in rice genotypes under salt stress, experiments were conducted using four rice varieties, including salt-sensitive BRRI dhan 28 and three salt-tolerant varieties BRRI dhan 47, BINA dhan 8 and BINA dhan 10. Thirty-day-old rice seedlings were transplanted into pots. At the active tillering stage(35 d after transplanting), plants were exposed to different salinity levels(0, 20, 40 and 60 mmol/L NaCl). Salt stress caused a significant reduction in growth for all the rice genotypes. Growth reduction was higher in the salt-sensitive genotype than in the salt-tolerant ones, and BINA dhan 10 showed higher salt tolerance in all measured physiological parameters. The reduction in shoot and root biomass was found to be minimal in BINA dhan 10. Chlorophyll content significantly decreased under salt stress except for BINA dhan 10. Proline content significantly increased in salt-tolerant rice genotypes with increased salt concentration, and the highest proline content was obtained from BINA dhan 10 under salt stress. Catalase and ascorbate peroxidase activities significantly decreased in salt-sensitive genotype whereas significantly increased in salt-tolerant ones with increasing salt concentration. However, salt stress significantly decreased guaiacol peroxidase activity in all the rice genotypes irrespective of salt tolerance. K~+/Na~+ ratio also significantly decreased in shoots and roots of all the rice genotypes. The salt-tolerant genotype BINA dhan 10 maintained higher levels of chlorophyll and proline contents as well as catalase and ascorbate peroxidase activities under salt stress, thus, this might be the underlying mechanism for salt tolerance.     

Cold tolerance during early reproductive growth of chickpea (Cicer arietinum L.): genetic variation in gamete development and function

Chilling temperatures at flowering cause floral abortion in most chickpea cultivars. Recent evaluations of germplasm showed distinct genotypic differences in pod and seed set at low temperature but the morpho-physiological basis for such variation is unclear. Observations in the field during cold spells of December and January, and at 15/5°C and 15/0°C (day/night) regimes in growth rooms showed distinct genetic variation in flower morphology, gamete development (viability and size of pollen and ovules) and function (pollen germination and tube growth, ovule viability and fertilization, etc.). The greater pod-setting ability of tolerant lines (ICCVs 88502 and 88503) than the sensitive cultivars (Annigeri, Pant G 114, etc.) was associated with a higher pollen vigour (germination and tube growth) and ovule viability at low temperature in the former. The number of ovules was not affected by cold stress in all cultivars/lines but pollen size and viability were reduced in Annigeri. The reduced ovule fertilization, associated with decline in pollen tube growth and ovule viability, was the major cause for poor seed set at low temperatures. The magnitude of effects on gamete function varied with cultivar/line and severity of stress. Function of both pollen and ovules was adversely affected in Annigeri in 15/5° and 15/0°C regimes. In contrast, the reduction in pollen vigour was more than in ovule viability in ICCV 88510 and Pant G 114 at 15/5°C. At 15/0°C, however, both pollen vigour and ovule viability were reduced. A small increase in pod set of sensitive genotypes from manual pollinations with pollen from plants in the warm regime further suggested that pollen function was more adversely affected than pistil function at moderately low temperatures.     

水稻冷胁迫响应基因OsSADMC功能标记的开发和利用

通过对95份水稻种质资源进行苗期耐冷性鉴定,筛选出10份苗期耐冷(存活率≥60%)和9份冷敏感的材料(存活率为零),并对水稻冷胁迫响应基因OsSADMC进行了表达分析,发现对照和冷胁迫处理条件下该基因在耐冷品种中的表达量都显著高于冷敏感品种。对耐冷材料丽江新团黑谷(LTH)和冷敏感材料IR29的基因编码区域进行了克隆和测序分析,在两个品种间鉴定了10个SNP位点,其中第749碱基处存在的一个碱基变异(C突变为T),从而使丝氨酸变为亮氨酸。根据其中一个SNP位点设计了OsSADMC的功能标记SADMC-CAPS1,该标记可以准确地鉴定出19份耐冷性不同水稻种质资源OsSADMC的基因型和耐冷性。     

Enclosed stigma contributes to higher spikelet fertility for rice(Oryza sativa L.) subjected to heat stress

With global warming, rice plants may be subjected to heat stress more regularly during the heatsensitive flowering stage, causing spikelet sterility and grain yield loss.Stigma exsertion is considered to increase pollen reception and promote female reproductive success.The aim of this study was to investigate the role of stigma exsertion on spikelet fertility at high temperatures.Five rice cultivars(Liangyoupeijiu, Shanyou 63, Huanghuazhan, Nagina 22, and IR64) with differing degrees of stigma exsertion were cultivated and exposed to high temperature at anthesis.Heat-tolerant cultivars did not always show a high percentage of spikelets with exserted stigmas, and vice versa.Irrespective of the presence of more pollen grains on exserted stigmas, spikelets with exserted stigmas did not show greater spikelet fertility than spikelets with fewer exserted stigmas or hidden stigmas under heat stress.GA3 application augmented the percentage of spikelets with exserted stigmas;however, it did not increase spikelet fertility under heat stress.Spikelet fertility of whole panicles was negatively correlated with the percentage of spikelets with exserted stigmas, but positively with that with hidden stigmas.Viability of the hidden stigmas was less reduced than that of exserted stigmas under heat stress, suggesting that hidden stigmas have an advantage in maintaining viability.Heat stress delayed anther dehiscence and reduced the viabilities of both exserted stigmas and pollens, thereby causing low spikelet fertility.Together, these results suggest that high spikelet fertility does not depend on stigma exsertion and that enclosed stigma generally contributes to higher spikelet fertility and heat tolerance under high-temperature conditions during flowering in rice.     

Differential Proteins Expressed in Rice Leaves and Grains in Response to Salinity and Exogenous Spermidine Treatments

Exogenous application of spermidine(Spd) has been reported to modulate physiological processes and alleviate salt-induced damage to growth and productivity of several plants including rice. Employing a proteomic approach, we aimed at identifying rice leaf and grain proteins differentially expressing under salt stress, and in response to Spd prior to Na Cl treatment. A total of 9 and 20 differentially expressed protein spots were identified in the leaves of salt-tolerant(Pokkali) and saltsensitive(KDML105) rice cultivars, respectively. Differential proteins common to both cultivars included a photosynthetic light reaction protein(oxygen-evolving complex protein 1), enzymes of Calvin cycle and glycolysis(fructose-bisphosphate aldolase and triose-phosphate isomerase), malate dehydrogenase, superoxide dismutase and a hypothetical protein(Os I_18213). Most proteins were present at higher intensities in Pokkali leaves. The photosynthetic oxygen-evolving enhancer protein 2 was detected only in Pokkali and was up-regulated by salt-stress and further enhanced by Spd treatment. All three spots identified as superoxide dismutase in KDML105 were up-regulated by Na Cl but down-regulated when treated with Spd prior to Na Cl, indicating that Spd acted directly as antioxidants. Important differential stress proteins detected in mature grains of both rice cultivars were late embryogenesis abundant proteins with protective roles and an antioxidant protein, 1-Cys-peroxiredoxin. Higher salt tolerance of Pokkali partly resulted from higher intensities and more responsiveness of the proteins relating to photosynthesis light reactions, energy metabolism, antioxidant enzymes in the leaves, and stress proteins with protective roles in the grains.     

磷对水稻高Fe2+胁迫的缓解作用

采用溶液培养法研究了高Fe2+胁迫下不同磷水平对粳稻Azucena（耐铁毒基因型）和籼稻IR64（铁毒敏感基因型）的生长和生理特性的影响。结果表明,与正常供Fe2+相比,高Fe2+胁迫抑制了水稻地上部和根系的生长,降低了干物质积累量和叶片叶绿素含量。外源供磷水平的提高,水稻地上部和根系生长受铁毒抑制程度有所减弱,叶片丙二醛含量和质膜透性下降,POD活性和叶绿素含量增加,而SOD活性则有所下降。表明在一定程度上,磷营养对提高水稻的耐Fe2+毒害具有重要作用,而磷对缓解Azucena的Fe2+毒害的效果较为明显。     

Selection of Rice Genotypes for Salinity Tolerance Through Morpho-Biochemical Assessment

The present study reported the morpho-biochemical evaluation of 15 selected rice genotypes for salt tolerance at the seedling stage. Growth parameters including shoot length, root length, plant biomass, plant turgid weight, plant dry weight along with relative water content were measured after exposure to saline solution （with electrical conductivity value of 12 dS/m）. Genotypes, showing significant differential responses towards salinity in the fields, were assessed through 14 salinity-linked morpho-biochemical attributes, measured at 14 d after exposure of seedling in saline nutrient solution. Relative water content, chlorophyll a/b, peroxidase activity and plant biomass were identified as potential indicators of salt tolerance. Principal component analysis and successive Hierarchical clustering using Euclidean distance revealed that Talmugur, Gheus, Ghunsi, Langalmura, Sabitapalui, and Sholerpona were promising genotypes for further breeding programmes in rice. The maximum Euclidean distance was plotted between Thavallakanan and Talmugur （7.49）, followed by Thavallakanan and Langalmura （6.82）, indicating these combinations may be exploited as parental lines in hybridization programmes to develop salinity tolerant variety.     

Alterations of Panicle Antioxidant Metabolism and Carbohydrate Content and Pistil Water Potential Involved in Spikelet Sterility in Rice under Water-Deficit Stress

Two rice genotypes with different drought tolerance,namely Jin 23B(drought tolerant)and Zhenshan 97B(drought sensitive),were used to study the antioxidant enzyme activities,soluble sugar and starch contents in spikelets,pistil water potential and pollen number on a stigma under water-deficit stress at the flowering stage,which were involved in the spikelet sterility.Compared with respective controls,drought stress induced more serious decreases of superoxide dismutase(SOD),peroxidase(POD)and catalase(CAT)ac...