A low-temperature method for maintaining plant regeneration activity in embryogenic callus of rice (Oryza sativa L.)

 A method was developed to maintain plant regeneration activity of rice cells (Oryza sativa L.) using embryogenic callus. Calluses were cultured in suspension, then on solid medium, to form compact globular callus resistant to low-temperature stress and with high plant regeneration activity. Callus preserved at 5  °C for 5 months regenerated plants from protoplasts at a frequency higher than from non-preserved callus from cv. Nipponbare, and cv. Koshihikari, but at lower rates from cv. Akitakomachi. Similar results were obtained from protoplasts of the three cultivars. Callus preserved at 5  °C for 8 months incurred cell damage, yet some surviving cells divided in suspension culture and eventually regenerated whole plants. Preserved and non-preserved regenerated plants showed similar levels of somaclonal variation. Received: 7 January 1999 / Revision received: 28 April 1999 / Accepted: 26 May 1999     

Reactions of six cultivars of rice (Oryza sativa L.) to three fungal diseases

Abstract

Fungal diseases are the most important threat for rice production in the world. Rice fields were visited and sampled from diseased bushes to identify fungal diseases in Kohgiluyeh and Boyer-Ahmad province, in the south-west of Iran. Fungi were isolated from diseased tissues by Agar-plate method, and purified by single-spore or hyphal-tip methods. They were identified by studying morphological characteristics. The virulence of isolated fungi was evaluated on six dominant rice cultivars, Champa of temperate region, Champa of subtropical region, Gerdeh, Shamim, Fajr and Tarom, under greenhouse conditions. The experiment was set up in a factorial experiment in completely randomized design with four replicates. The incidence of three fungal diseases: brown spot caused by Curvularia australiensis and Bipolaris cookei, foot rot caused by Fusarium globosum and Fusarium sambucinum, and black root rot caused by Exserohilum pedicellatum, was proven in this study. Although, all of the tested cultivars significantly showed different reaction to these fungi, but the cultivars Champa of subtropical region and Tarom were resistant to foot rot as well as the black root rot and the cultivar Gerdeh was resistant to brown spot disease.     

Using minimum DNA marker loci for accurate population classification in rice (Oryza sativa L.)

Because of the rich diversity among rice accessions grown around the world in distinct environments, traditional methods using morphology, cross compatibility and geography for classifying rice accessions according to different sub-populations have given way to use of molecular markers. Having a few robust markers that can quickly assign population structure to germplasm will facilitate making more informed choices about genetic diversity within seedbanks and breeding genepools. WHICHLOCI is a computer program that selects the best combination of loci for population assignment through empirical analysis of molecular marker data. This program has been used in surveys of plant species, for fish population assignment, and in human ancestry analysis. Using WHICHLOCI, we ranked the discriminatory power of 72 DNA markers used to genotype 1,604 accessions of the USDA rice core collection, and developed panels with a minimum number of markers for population assignment with 99% or higher accuracy. A total of 14 markers with high discriminatory power, genetic diversity, allelic frequency, and polymorphic information content were identified. A panel of just four markers, RM551, RM11, RM224 and RM44, was effective in assigning germplasm accessions to any of five sub-populations with 99.4% accuracy. Panels using only three markers were effective for assignment of rice germplasm to specific sub-populations, tropical japonica, temperate japonica, indica, aus, and aromatic. Assignment to tropical japonica, temperate japonica, or indica sub-populations was highly reliable using 3–4 markers, demonstrated by the high correlation with assignment using 72 markers. However, population assignment to aus and aromatic groups was less reliable, possibly due to the smaller representation of this material in the USDA core collection. More reference cultivars may be needed to improve population assignment to these two groups. This study demonstrated that a small number of DNA markers is effective for classification of germplasm into five sub-populations in rice. This will facilitate rapid screening of large rice germplasm banks for population assignment at a modest cost. The resulting information will be valuable to researchers to verify population classification of germplasm prior to initiating genetic studies, maximizing genetic diversity between sub-populations, or minimizing cross incompatibility while maximizing allelic diversity within specific sub-populations.     

Integration of crop rotation and arbuscular mycorrhiza (AM) inoculum application for enhancing AM activity to improve phosphorus nutrition and yield of upland rice (Oryza sativa L.)

Upland rice (Oryza sativa L.) is a major crop of Eastern India grown during the wet season (June/July to September/October). Aerobic soils of the upland rice system, which are acidic and inherently phosphorus (P) limiting, support native arbuscular mycorrhizal (AM) activity. Attempts were made to improve P nutrition of upland rice by exploiting this natural situation through different crop rotations and application of AM fungal (AMF) inoculum. The effect of a 2-year crop rotation of maize (Zea mays L.) followed by horse gram (Dolichos biflorus L.) in the first year and upland rice in the second year on native AM activity was compared to three existing systems, with and without application of a soil–root-based inoculum. Integration of AM fungal inoculation with the maize–horse gram rotation had synergistic/additive effects in terms of AMF colonization (+22.7 to +42.7%), plant P acquisition (+11.2 to +23.7%), and grain yield of rice variety Vandana (+25.7 to +34.3%).     

Effect of sorbitol concentration on regeneration of embryogenic calli in upland rice varieties (Oryza sativa L.)

This study describes the impact of sorbitol on plantlets regeneration frequency (PRF) of four rice cultivars (japonica, Oryza sativa L.) both of which mature and immature embryo-derived calli were investigated. The variance analysis results showed that PRF of the three elite upland rice cultivars, Handao297 (HD297), Handao502 (HD502), Handao65 (HD65) and one lowland rice cultivar Zhongzuo93 (ZZ93) were significantly increased with addition of appropriate amount of sorbitol in culture media. Supplementing appropriate sorbitol in the media of a continous culture from induction and maintenance to regeneration for mature embryo-derived calli could improve PRF dramatically, originally from 27.6% up to a maximum of 71.8%. Especially to low regenerative capacity (LRC) cultivar HD65, the PRF was increased over 7-fold (from 9.7% to 74.0%). The optimum concentrations of sorbitol for calli induction, subculture and differentiation were 5, 20 and 40 g/l, respectively. Adding sorbitol, only in maintenance media at concentration of 20 g/l, also enhanced the PRF greatly in all the cultivars from 27.6% to 43.3%. Similar results were observed when incorporating with maltose in regenerating media both in immature and mature embryo-derived calli. The optimal concentration was 25 g/l sorbitol + 20 g/l maltose and 20 g/l sorbitol + 25 g/l maltose, respectively. HD297 appeared to be the most responsive genotype compared to other cultivars in PRF, 99.2% in immature embryo-derived calli and 76.8% in mature embryo-derived calli. The results and relevant conclusions might be valuable to establish an efficient plant regeneration system from somatic embryogenesis culture in upland rice.     

Genetic variation detected with RAPD markers among upland and lowland rice cultivars (Oryza sativa L.)

Genetic variation of nine upland and four lowland rice cultivars (Oryza sativa L.) was investigated at the DNA level using the randomly amplified polymorphic DNA (RAPD) method via the polymerase chain reaction (PCR). Forty-two random primers were used to amplify DNA segments and 260 PCR products were obtained. The results of agarosegel electrophoretic analysis of these PCR products indicated that 208 (80%) were polymorphic. All 42 primers used in this experiment were amplified and typically generated one-to-four major bands. Only two primers showed no polymorphisms. In general, a higher level of polymorphism was found between japonica and indica subspecies while fewer polymorphisms were found between upland and lowland cultivars within the indica subspecies. A dendrogram that shows the genetic distances of 13 rice cultivars was constructed based on their DNA polymorphisms. Classification of rice cultivars based on the results from the RAPD analysis was identical to the previous classification based on isozyme analysis. This study demonstrated that RAPD analysis is a useful tool in determining the genetic relationships among rice cultivars.     

Zinc-copper antagonism in the nutrition of rice (Oryza sativa L.)

Summary The effect of various doses of copper and zinc on their uptake and on the yield of rice were studied. Copper applications increased copper contents in the plants without effecting the zinc contents. However, zinc applications though increased zinc contents but markedly decreased the copper contents in the plants. This antagonistic effect of zinc on copper suggests that zinc applications can reduce rice yield if available copper is marginal in the soils. re]19720628     

Low irradiance stress tolerance in rice (Oryza sativa L.)

The effect of low irradiance on three rice cultivars (shade tolerant cvs. Swarnaprabha and CO 43 and shade susceptible cv. IR 20) was studied. The large subunit (LSU) of ribulose-1,5-bisphosphate carboxylase/oxygenase with molecular mass of 55 kDa was reduced in cv. IR 20 grown under low irradiance (LI). Native protein profile studied showed, under LI, reduction in the contents of proteins with RF values 0.03, 0.11 and 0.37. Analysis of chloroplast polypeptides revealed an induction of light-harvesting chlorphyll-protein 2 (LHCP2) under shade. The induction was more expressed in cv. CO 43 than in cv. IR 20. Under LI, in vivo labelled protein bands in the molecular range of 26 - 27 kDa were induced. These proteins were highly turned over in the LI-grown plants of cv. CO 43 than in cv. IR 20. A signal for rbcL gene sequences in EcoRI digested lanes was also found. Isozyme analysis of peroxidase showed an induction of a new band with RF 0.43 in cv. IR 20 subjected to LI. This revised version was published online in July 2006 with corrections to the Cover Date.     

Arsenic toxicity to rice (Oryza sativa L.) in Bangladesh

Natural contamination of groundwater with arsenic (As) occurs around the world but is most widespread in the river basin deltas of South and Southeast Asia. Shallow groundwater is extensively used in the Bengal basin for irrigation of rice in the dry winter season, leading to the possibility of As accumulation in soils, toxicity to rice and increased levels of As in rice grain and straw. The impact of As contaminated irrigation water on soil-As content and rice productivity was studied over two winter-season rice crops in the command area of a single tubewell in Faridpur district, Bangladesh. After 16–17 years of use of the tubewell, a spatially variable build up of As and other chemical constituents of the water (Fe, Mn and P) was observed over the command area, with soil-As levels ranging from about 10 to 70 mg kg^?1. A simple mass balance calculation using the current water As level of 0.13 mg As L^?1 suggested that 96% of the added arsenic was retained in the soil. When BRRI dhan 29 rice was grown in two successive years across this soil-As gradient, yield declined progressively from 7–9 to 2–3 t ha^?1 with increasing soil-As concentration. The average yield loss over the 8 ha command area was estimated to be 16%. Rice-straw As content increased with increasing soil-As concentration; however, the toxicity of As to rice resulted in reduced grain-As concentrations in one of the 2 years. The likelihood of As-induced yield reductions and As accumulation in straw and grain has implications to agricultural sustainability, food quality and food security in As-affected regions throughout South and Southeast Asia.     

Mapping of QTLs for phosphorus-deficiency tolerance in rice (Oryza sativa L.)

 Phosphorus (P) deficiency of soils is a major yield-limiting factor in rice production. Increasing the P-deficiency tolerance of rice cultivars may represent a more cost-effective solution than relying on fertilizer application. The objective of this study was to identify putative QTLs for P-deficiency tolerance in rice, using 98 backcross inbred lines derived from a japonica×indica cross and genotyped at 245 RFLP marker loci. Lines were grown on P-deficient soil and P uptake, internal P-use efficiency, dry weight, and tiller number were determined. Three QTLs were identified for dry weight and four QTLs for P uptake, together explaining 45.4% and 54.5% of the variation for the respective traits. Peaks for both traits were in good agreement which was to be expected considering the tight correlation of r=0.96 between dry weight and P uptake. For both traits the QTL linked to marker C443 on chromosome 12 had a major effect. Two of the three QTLs detected for internal P-use efficiency, including the major one on chromosome 12, coincided with QTLs for P uptake; however, whereas indica alleles increased P uptake they reduced P-use efficiency. We concluded that this was not due to the tight linkage of two genes in repulsion but rather due to an indirect effect of P uptake on P-use efficiency. Most lines with high use efficiency were characterized by very low P uptake and dry weight and apparently experienced extreme P-deficiency stress. Their higher P-use efficiency was thus the result of highly sub-optimal tissue-P concentrations and did not represent a positive adaptation to low P availability. The number of tillers produced under P deficiency is viewed as an indirect indicator of P-deficiency tolerance in rice. In addition to the major QTL on chromosome 12 already identified for all other traits, two QTLs on chromosome 4 and 12 were identified for tiller number. Their position, however, coincided with QTLs for tiller number reported elsewhere under P-sufficient conditions and therefore appear to be not related to P-deficiency tolerance. In this study P-deficiency tolerance was mainly caused by differences in P uptake and not in P-use efficiency. Using a trait indirectly related to P-deficiency tolerance such as tiller number, we detected a major QTL but none of the minor QTLs detected for P uptake or dry weight. Received: 9 February 1998 / Accepted: 29 April 1998     

Mapping QTLs for phosphorus deficiency tolerance in rice (Oryza sativa L.)

 The amplified fragment length polymorphism (AFLP) technique combined with selective genotyping was used to map quantitative trait loci (QTLs) associated with tolerance for phosphorus (P) deficiency in rice. P deficiency tolerant cultivar IR20 was crossed to IR55178-3B-9-3 (sensitive to P-deficiency) and 285 recombinant inbred lines (RILs) were produced by single-seed descent. The RILs were phenotyped for the trait by growing them in P-sufficient (10.0 mg/l) and P-deficient (0.5 mg/l) nutrient solution and determining their relative tillering ability at 28 days after seeding, and relative shoot dry weight and relative root dry weight at 42 days after seeding. Forty two of each of the extreme RILs (sensitive and tolerant) and the parents were subjected to AFLP analysis. A map consisting of 217 AFLP markers was constructed. Its length was 1371.8 cM with an average interval size of 7.62 cM. To assign linkage groups to chromosomes, 30 AFLP and 26 RFLP markers distributed over the 12 chromosomes were employed as anchor markers. Based on the constructed map, a major QTL for P-deficiency tolerance, designated PHO, was located on chromosome 12 and confirmed by RFLP markers RG9 and RG241 on the same chromosome. Several minor QTLs were mapped on chromosomes 1, 6, and 9. Received: 21 April 1998 / Accepted: 9 June 1998     

RFLP analysis of rice (Oryza sativa L.) introgression lines

Summary Fifty-two introgression lines (BC₂F₈) from crosses between two Oryza sativa parents and five accessions of O. officinalis were analyzed for the introgression of O. officinalis chromosome segments. DNA from the parents and introgression lines was analyzed with 177 RFLP markers located at approximately 10-cM intervals over the rice chromosomes. Most probe/enzyme combinations detected RFLPs between the parents. Of the 174 informative markers, 28 identified putative O. officinalis introgressed chromosome segments in 1 or more of the introgression lines. Introgressed segments were found on 11 of the 12 rice chromosomes. In most cases of introgression, O. sativa RFLP alleles were replaced by O. officinalis alleles. Introgressed segments were very small in size and similar in plants derived from early and later generations. Some nonconventional recombination mechanism may be involved in the transfer of such small chromosomal segments from O. officinalis chromosomes to those of O. sativa. Some of the introgressed segments show association with genes for brown planthopper (BPH) resistance in some introgressed lines, but not in others. Thus, none of the RFLP markers could be unambiguously associated with BPH resistance.     

Chronopathological aspects of disease incidence in rice (Oryza sativa L.)

Rice seedlings maintained under uncontrolled glasshouse conditions were inoculated with conidial suspensions of a fungal pathogen, Helminthosporium oryzae, at various times during the 24 h. Significant increase in the percent germination and germ tube length of conidia were observed in the rice samples inoculated at 02:00 and 06:00h. The 24 h temporal variation in leaf temperature was positively correlated with variation in stomatal movements. The results indicate a 24 h rhythm in the behavior of the fungal pathogen on the host in relation to the conditions of the growing environment. In all the inoculated seedlings, the appearance of a large number of brown leaf spots was confined to the light span. Among the plants inoculated, earlier initiation of brown leaf spot appearance, maximum number of leaf spots, and highest disease severity were observed when plants were inoculated at 02:00h. There was a positive correlation between disease severity of the host and in vivo values of percent germination of conidia and germ tube length of the pathogen in plants inoculated between 02:00 and 06:00h. The findings of this study implicate that light intensity and temperature could play a predominant role in controlling disease susceptibility rhythms in plants.     

QTL mapping for the paste viscosity characteristics in rice (Oryza sativa L.)

In order to understand the genetic basis of the paste viscosity characteristics (RVA profile, which is tested on the Rapid Visco Analyser) of the rice grain, we mapped QTLs for RVA profile parameters using a DH population derived from a cross between an indica variety, Zai-Ye-Qing 8 (ZYQ8), and a japonica variety, Jing-Xi 17 (JX17). Evidence of genotype-by-environment interaction was found by comparing the mapped QTLs between two locations, Hainan (HN) and Hangzhou (HZ). A total of 20 QTLs for six parameters of the RVA profiles were identified at least one location. Only the waxy locus (wx) located on chromosome 6 was detected significantly at both environments for five traits, i.e. hot paste viscosity (HPV), cool paste viscosity (CPV), breakdown viscosity (BDV), consistency viscosity (CSV) and setback viscosity (SBV). This locus explained 19.5%–63.7% of the total variations at both environments, suggesting that the RVA profiles were mainly controlled by the wx gene. HPV, CPV, BDV, CSV and SBV were also controlled by other QTLs whose effects on the respective parameter were detected only in one environment, while for the peak viscosity (PKV), only 2 QTLs, 1 at HN,the other at HZ, were identified. These results indicate that RVA profiles are obviously affected by environment. Received: 18 July 1999 / Accepted: 27 August 1999     

High photosynthetic efficiency of a rice (Oryza sativa L.) xantha mutant

Comparative analysis revealed that a xantha rice mutant (cv. Huangyu B) had higher ratios of chlorophyll (Chl) a/b and carotenoids/Chl, and higher photosynthetic efficiency than its wild type parent (cv. II32 B). Unexpectedly, the mutant had higher net photosynthetic rate (P _N) than II32 B. This might have resulted from its lower non-photochemical quenching (q_N) but higher maximal photochemical efficiency (F_V/F_M), higher excitation energy capture efficiency of photosystem 2 (PS2) reaction centres (F_V′/F_M′), higher photochemical quenching (q_P), higher effective PS2 quantum yield (Φ_PS2), and higher non-cyclic electron transport rate (ETR). This is the first report of a chlorophyll mutant that has higher photosynthetic efficiency and main Chl fluorescence parameters than its wild type. This mutant could become a unique material both for the basic research on photosynthesis and for the development of high yielding rice cultivars.     

Genetic analysis of plant regeneration in rice (Oryza sativa L.)

Summary A diallel study involving reciprocal crosses of four genotypes (IR8, 36, 54, and 64) was carried out to understand the genetic mechanism of plant regeneration from immature embryo-calli in rice. Regeneration frequency (% of calli that produced plants) varied from a high of 86% for IR54 to a low of 0% for IR36, while regenerated plants per embryo numbered from 0 to 7 when these same IR lines and the F₁ hybrids were compared. Combining ability analysis revealed that both nuclear (with both additive and dominant effects) and cytoplasmic genes are important in controlling regeneration in rice. Parental lines and F1 hybrids with high ability to regenerate were identified.     

Genetic analysis of salinity tolerance in rice (Oryza sativa L.)

Summary The genetics of salinity tolerance in rice was investigated by a nine-parent complete diallel including reciprocals. Test materials involved susceptible (IR28, IR29, and MI-48), moderately tolerant (IR4595-4-1-13, IR9884-54-3-1E-P1, and IR10206-29-2-1), and tolerant (Nona Bokra, Pokkali, and SR26B) parents. Twoweek-old seedlings were grown in a salinized (EC = 12 dS/m) culture solution for 19 days under controlled conditions in the IRRI phytotron. Typical characteristics of salinity tolerance in rice were found to be Na⁺ exclusion and an increased absorption of K⁺ to maintain a good Na-K balance in the shoot. Genetic component analysis (GCA) revealed that a low Na-K ratio is governed by both additive and dominance gene effects. The trait exhibited overdominance, and two groups of genes were detected. Environmental effects were large, and the heritability of the trait was low. Our findings suggest that when breeding for salt tolerance, selection must be done in a later generation and under controlled conditions in order to minimize environmental effects. Modified bulk and single-seed descent would be the suitable breeding methods. Combining ability analysis revealed that both GCA and specific combining ability (SCA) effects were important in the genetics of salt tolerance. Moderately tolerant parents — e.g., IR4595-4-1-13 and IR9884-54-3-1E-P1 — were the best general combiners. Most of the best combinations had susceptible parents crossed either to moderate or tolerant parents. The presence of reciprocal effects among crosses necessitates the use of susceptible parents as males in hybridization programs. Large heterotic effects suggest the potential of hybrid rice for salt-affected lands.