Ahmad Arzani

Improving salinity tolerance in crop plants: a biotechnological view

Salinity limits the production capabilities of agricultural soils in large areas of the world. Both breeding and screening germplasm for salt tolerance encounter the following limitations: (a) different phenotypic responses of plants at different growth stages, (b) different physiological mechanisms, (c) complicated genotype × environment interactions, and (d) variability of the salt-affected field in its chemical and physical soil composition. Plant molecular and physiological traits provide the bases for efficient germplasm screening procedures through traditional breeding, molecular breeding, and transgenic approaches. However, the quantitative nature of salinity stress tolerance and the problems associated with developing appropriate and replicable testing environments make it difficult to distinguish salt-tolerant lines from sensitive lines. In order to develop more efficient screening procedures for germplasm evaluation and improvement of salt tolerance, implementation of a rapid and reliable screening procedure is essential. Field selection for salinity tolerance is a laborious task; therefore, plant breeders are seeking reliable ways to assess the salt tolerance of plant germplasm. Salt tolerance in several plant species may operate at the cellular level, and glycophytes are believed to have special cellular mechanisms for salt tolerance. Ion exclusion, ion sequestration, osmotic adjustment, macromolecule protection, and membrane transport system adaptation to saline environments are important strategies that may confer salt tolerance to plants. Cell and tissue culture techniques have been used to obtain salt tolerant plants employing two in vitro culture approaches. The first approach is selection of mutant cell lines from cultured cells and plant regeneration from such cells (somaclones). In vitro screening of plant germplasm for salt tolerance is the second approach, and a successful employment of this method in durum wheat is presented here. Doubled haploid lines derived from pollen culture of F1 hybrids of salt-tolerant parents are promising tools to further improve salt tolerance of plant cultivars. Enhancement of resistance against both hyper-osmotic stress and ion toxicity may also be achieved via molecular breeding of salt-tolerant plants using either molecular markers or genetic engineering.

Response of durum wheat cultivars to immature embryo culture, callus induction and in vitro salt stress

Response of twenty eight cultivars of durum wheat (Triticum turgidum var. durum) to immature embryo culture, callus production and in vitro salt tolerance was evaluated. For assessment of cultivars to salt tolerance, growing morphogenic calli were exposed to different concentrations of NaCl (0, 0.3, 0.6, 0.9, 1.2, 1.5, 1.8 and 2.1% w/v) added to the culture medium during two subsequent subcultures (4 weeks each). Comparison of cultivars for callus induction from immature embryo was based on callus induction frequency and fresh weight growth of callus (FWG). While, for salt tolerance, the relative fresh weight growth (RFWG) and necrosis percent of callus were used. There were significant differences among cultivars for potential of regeneration from immature embryo, and ‘Shahivandi’ a native durum wheat cultivar originating from western Iran was superior among the cultivars tested. The FWG distinguished cultivars more than callus induction frequency did for callus induction evaluation. Hence, a range of FWG from 1.23 to 14.65 g was observed in ‘Mexical-75’ and ‘Omrabi-5’ cultivars, respectively. Growing calli derived from cultivars ‘PI 40100’ and ‘Dipper-6’ showed superiority for tolerating salinity under in vitro conditions.

Essential oil composition, total phenolic, flavonoid contents, and antioxidant activity of Thymus species collected from different regions of Iran

In this study, the essential oil (EO) composition, flavonoid and phenolic contents, and antioxidant activities of fourteen Thymus accessions belonging to ten species were evaluated. Gas chromatography-mass spectrometry (GC-MS) analysis revealed the presence of 38 compounds with the major constituents including thymol (12.4-79.74%), carvacrol (4.37-42.14%), geraniol (0.3-22.44%), and p-cymene (0.8-12.86%). Cluster analysis identified three groups of high thymol, geraniol/linalool, and high carvacrol. The highest phenolic and flavonoid contents were detected in T. daenensis-1 (70.6mg tannic acid equivalents (TAE) g-1 DW) and T. vulgaris (8.55mg quercetin equivalents (QE) g-1 DW), respectively. The antioxidant activities of the samples were determined using 1,1-diphenyl-2-picrylhydrazyl (DPPH) and reducing power assay. The results demonstrated that T. daenensis-3 (IC50=273.36), T. vulgaris (IC50=289.3), and T. fedtschenkoi-3 (IC50=339.22) possessed higher antioxidant activities than the others. Finally, the Thymus species with high bioactive compounds may be recommended for further food applications.

Identification of microsatellite markers linked with yield components under drought stress at terminal growth stages in durum wheat

Grain yield and yield components are the main important traits involved in durum wheat (Triticum turgidum L.) improvement programs. The purpose of this research was to identify quantitative trait loci (QTL) associated with yield components such as 1000 grain weight (TGW), grain weight per spike (GWS), number of grains per spike (GNS), spike number per m2 (SN), spike weight (SW), spike harvest index (SHI) and harvest index (HI) using microsatellite markers. Populations of F3 and F4 lines derived from 151 F2 individuals developed from a cross between Oste-Gata, a drought tolerant, and Massara-1, a drought susceptible durum wheat genotypes, were used. The populations were evaluated under four environmental conditions including two irrigation regimes of drought stress at terminal growth stages and normal field conditions in two growing seasons. Two hundred microsatellite markers reported for A and B genomes of bread wheat were used for parental polymorphism analysis and 30 polymorphic markers were applied to genotype 151 F2:3 families. QTL analysis was performed using genome-wide single marker regression analysis (SMA) and composite interval mapping (CIM). The results of SMA revealed that about 20% of the phenotypic variation of harvest index and TGW could be explained by Xcfd22-7B and Xcfa2114-6A markers in different environmental conditions. Similarly, Xgwm181-3B, Xwmc405-7B and Xgwm148-3B and marker Xwmc166-7B were found to be associated with SHI and GWS, respectively. A total of 20 minor and major QTL were detected; five for TGW, two for GWS, two for GNS, three for SN, five for HI, two for SHI and one for SW. The mapped QTL associated with ten markers. Moreover, some of these QTL were prominent and stable under drought stress and non drought stress environments and explained up to 49.5% of the phenotypic variation.

Study of genetic diversity in safflower genotypes using agro-morphological traits and RAPD markers

This research was conducted to study the genetic diversity in safflower (Carthamus tinctorius L.) using agro-morphological traits and RAPD markers. Sixteen selected lines derived from landraces growing in various agro-climatic regions of Iran along with four exotic genotypes were evaluated in a randomized complete block design with three replications under field conditions. Days to emergence, days to initial flowering, days to flowering, days to maturity, plant height, branches per plant, capitula per plant, seeds per capitulum, 1,000-seed weight, seed yield per plant, seed yield, and reaction to powdery mildew (Leveillula taurica Arnaud) were evaluated in this study. Genetic diversity of the genotypes was assessed by RAPD markers. The results indicated significant differences among genotypes for the agro-morphological traits and clustering based on these traits classified the genotypes into five groups. Analysis of the RAPD markers revealed 15 polymorphic primers out of 50 used primers. Based on RAPD data, the highest genetic similarity was observed between the cultivars of “AC Sunset,” “AC Sterling” from Canada and the lowest relatedness observed between a local breeding line “E2428” and genotype “GE62923” from Germany. Cluster analysis based on RAPD markers and 54% coefficient of similarity divided the genotypes into five distinct groups. Comparing the clusters based on agro-morphological traits with those from molecular markers showed slight similarities. The finding of high genetic variation for agro-morphological traits and polymorphism at DNA level reveal that agronomic traits can be improved by selection programs.

Genetic variation in safflower (Carthamus tinctorious L.) for seed quality-related traits and inter-simple sequence repeat (ISSR) markers.

Safflower (Carthamus tinctorious L.) is an oilseed crop that is valued as a source of high quality vegetable oil. The genetic diversity of 16 safflower genotypes originated from different geographical regions of Iran and some with exotic origin were evaluated. Eight different seed quality-related traits including fatty acid composition of seed oil (stearic acid, palmitic acid, oleic acid and linoleic acid), the contents of, oil, protein, fiber and ash in its seeds, as well as 20 inter-simple sequence repeat (ISSR) polymorphic primers were used in this study. Analysis of variance showed significant variation in genotypes for the seed quality-related traits. Based on ISSR markers, a total of 204 bands were amplified and 149 bands (about 70%) of these were polymorphic. Cluster analysis based on either biochemical or molecular markers classified the genotypes into four groups, showing some similarities between molecular and biochemical markers for evaluated genotypes. A logical similarity between the genotype clusters based on molecular data with their geographical origins was observed.

Evaluation of Oil Compositions of Iranian Mints (Mentha ssp.)

Abstract Dried aerial parts of 12 accessions of Iranian mints, three of which belonged to Mentha longifolia (L.) Huds. and the remaining of which belonged to Mentha spicata L., were used in this study. These were water distilled, and the essential oils analyzed by GC (FID) and GC/MS. Results indicated a significant variation in oil composition within the accessions. cis-Carveol (53.5–78.2%) was found as the main constituent in the oils of three M. longifolia accessions. The studied accessions of M. spicata indicated six different chemotypes, each of which was characterized by a high contribution of the following compound: 1 terpinen-4-yl acetate (51.7–55.3%); 2 carvone oxide (52.5%); 3 piperitone and menthol (49.3% and 15.7%, respectively); 4 carvacrol (49.6%); 5 isomenthone and piperitone oxide (39.1% and 23.3%, respectively); (6) p-cymene and trans-carvyl acetate (48.6% and 32.2%, respectively). The chemical variability of the tested accessions suggests the possibility of further exploiting and improving of the populations in a breeding program.

Smart Engineering of Genetic Resources for Enhanced Salinity Tolerance in Crop Plants

ABSTRACT Salinity is a consistent factor of crop productivity loss in the world and in particular arid and semi-arid areas where the soil salinity and saline water are major problems. Plants employ various mechanisms to cope with salinity stress and activate an array of stress-responsive genes to counteract the salinity-induced osmotic and ionic stresses. Genetic improvement for salinity tolerance is challenging, and thus progress attained over the several decades has been far less than anticipated. The generation of an explosion of knowledge and technology related to genetics and genomics over the last few decades is promising in providing powerful tools for future development of salinity-tolerant cultivars. Despite a major progress in defining the underlying mechanisms of salinity tolerance, there are still major challenges to be overcome in translating and integrating the resultant information at the molecular level into plant-breeding practices. Various approaches have been suggested to improve the efficiency of plant breeding for increasing plant productivity under saline environments. In this context, breeding for salinity tolerance in crops largely depends upon the availability of genetic resources of tolerance, reliable screening techniques, identification of genetic components of tolerance, and successful genetic manipulation of desired genetic backgrounds. The efficiency of selection and breeding in the stressful environments can be improved through marker-assisted selection. To date, this is almost exclusively applied to major genes, but this requires to be extended to quantitative trait loci (QTLs) controlling complex traits such as salinity tolerance to greatly enhance the impact. Moreover, methodologies for high-throughput genotyping, and the development of an array of “functional” markers can be much supportive. The introduction of novel genes or alteration in the expression patterns of the existing genes through the generation of transgenic plants can also be employed to overcome the limits in classical plant breeding. The introgression of wild halophytic attribute genes facilitated by genetic engineering is an alternative approach to bypass interspecific hybridization barriers, which will stimulate breakthrough in the future agriculture. The molecular dissection of salinity-tolerance trait, accompanying the classical quantitative genetics, is a substantial progress in updating tools and methods for the manipulation of plant genomes. Methods of gene discovery such as identification of candidate genes, QTL cloning, linkage and association mapping, and functional genomics such as identification of transcripts and proteins involved in salinity tolerance are necessary to manipulate the molecular mechanisms underlying the complex phenotype of salinity tolerance. Some of the challenges and opportunities have also been addressed in the present review with a particular emphasis on molecular breeding approaches to be employed in combination with other crop improvement strategies to develop salinity-tolerant cultivars.

Meiotic behaviour of tetraploid wheats (Triticum turgidum L.) and their synthetic hexaploid wheat derivates influenced by meiotic restitution and heat stress

Meiotic restitution is considered to be a common mechanism of polyploidization in plants and hence is one of the most important processes in plant speciation. Meiotic behaviour of plant chromosomes is influenced by both genetic and environmental factors. In this study, the meiotic behaviour of cereal crops was investigated, which includes tetraploid wheat genotypes (with and without the meiotic restitution trait) and their derivates (synthetic hexaploid wheats and a doubled haploid (DH) line), grown at two planting dates in the field. In addition, two local landraces of emmer wheat (Triticum turgidum ssp. dicoccum), one wheat cultivar (Chinese spring), one DH triticale cultivar (Eleanor) and one rye accession were included. Immature spikes of mid-autumn and end-winter sowing plants were collected in April and May 2008, respectively, fixed in Carnoy’s solution and stained with hematoxylin. Pollen mother cells (PMCs) from anthers at different stages of meiotic process were analysed for their chromosomal behaviour and irregularities. Meiotic aberrations such as laggards, chromosome bridges, micronuclei, abnormal cytokines, chromatin pulling and meiotic restitution were observed and the studied genotypes were accordingly ranked as follows: triticale > synthetic hexaploid wheats > tetraploid wheats possessing meiotic restitution > tetraploid wheats lacking meiotic restitution > rye. The results indicated that the samples that had been planted in the autumn, thus experiencing an optimum temperature level at the flowering stage, exhibited less meiotic irregularities than winter planting samples that encountered heat stress at the flowering period.

The effect of colchicine on triticale anther-derived plants: Microspore pre-treatment and haploid-plant treatment using a hydroponic recovery system

In cereals, chromosome doubling of microspore-derived haploid plants is a critical step in producing doubled haploid plants. This investigation was undertaken to study the effect of incorporation of colchicine in the induction medium for anther culture, and the effect of colchicine on anther culture-derived plants of triticale grown under controlled greenhouse conditions. In the latter case, chromosome doubling of adult sterile plants derived from anther culture of fourteen triticale populations was attempted, where androgenetic plants with non-dehiscent anthers were cloned and subjected to the colchicine treatment, and then grown with the aid of hydroponics. The hydroponic system provided optimal conditions for recovery of the affected haploids from the toxic effects of colchicine treatment and all colchicine-treated plants survived. A topcross-F1 (TC1F1) population with timopheevii cytoplasm produced the highest percentage of plants with seed-set either due to chromosome doubling by colchicine (98%) or spontaneous doubling of chromosome number (15%). Colchicine-treated anthers performed inferior than control in both induction and regeneration phases. One of the key observation of this study was the reversal from reproductive stage back to the vegetative stage which in turn enabled further cloning of haploid plants under hydroponic conditions once they were identified as sterile. The one hundred percent survival rate of in vitro-derived plants, 100% survival rate of colchicine treated haploid plants and the high chromosome doubling success rate (X = 82.3) observed in this study imply that a temperature-controlled greenhouse with an hydroponic system provides an efficient environment for inducing chromosome doubling of haploid plants in cereals.