نشریه بیوتکنولوژی و بیوشیمی غلات
سال دوم شماره 2 (پیاپی 6، تابستان 1402)

Barley (Hordeum vulgare L.) is known as the fourth most important grain crop in the world. Drought stress is also one of the main factors that reduce the growth and yield of barley, and grain yield is directly related to the morphological characteristics of the flag leaf.. In order to identify the main and epistatic genomic regions and investigate their interaction with the environment for morphological traits related to flag leaf, including length, width, area, and flag leaf sheath length, an experiment was conducted with 136 doubled haploid lines of barley along with their parents 'Nure' and 'Tremois' using an alpha lattice design with two replications under normal and drought stress conditions at the Zabol Agricultural Research Station, Iran, in 2015-16 growing season. The genetic map of the target population comprised 543 markers (including DArT, SSR, SNP, and AFLP markers) for the QTL analysis. In this study, single-locus and two-locus QTL analyses were performed using the composite interval mapping (CIM) and mixed-model based composite interval mapping (MCIM) methods, respectively. Based on a single-locus analysis, five QTLs were identified, including three putative QTLs, Q1FLW-1H.SZ, Q1SHFLL-1H.NZ and Q2SHFLL-1H.NZ at bPb-6343-bPb-8081, bPb-1419-bPb-9180, and bPb-9108-bPb-6343 marker intervals, and two suggestive QTLs, on chromosomes 1H and 5H for the traits of flag leaf width (FLW, cm) and flag leaf sheath length (SHFLL, cm). Based on a two-locus analysis, eight additive (M-QTL) QTLs and 15 pairs of epistatic effects (E-QTL) were detected for different traits, among which two pairs of epistatic QTLs for the FLW trait showed significant QTL × environment interactions (AAEI). All detected M-QTLs for flag-leaf-related traits across two environments, without additive × environment interactions (AEI), were identified as stable QTLs, and the highest number of stable QTLs was associated with chromosome 1H. In this study, drought stress-tolerant QTLs were identified in all barley chromosomes except chromosome 2H. Most of the QTLs associated with the investigated traits in the Nure/Tremois population in the present study have not been reported previously and, therefore, are novel. The stable and putative QTLs controlling flag-leaf-related traits in this study, along with the markers linked to these QTLs, can be utilized in marker-assisted selection (MAS) and gene pyramiding breeding programs after confirmation and validation in different environmental conditions and genetic backgrounds to create high-yielding and drought-tolerant barley cultivars. Additionally, these findings will accelerate the understanding of the genetic relationships among spike-related traits at the molecular level.

Wheat ranks first among crops regarding cultivated area and production worldwide and in Iran. Drought stress is the most common abiotic stress that decreases the grain yield of crops. Physiologic characteristics with some traits related to grain yield and its components are used as indices to evaluate wheat cultivars under drought stress conditions. This experiment aimed to study the grain yield components and some physiologic characteristics of the flag leaf and their relationship with grain yield in commercial wheat varieties under post-anthesis drought stress conditions.
This experiment was conducted at Razi University. The experiment was arranged as a split plot basis on a randomized complete block design with three replications. Two moisture treatments (no stress and post-anthesis drought stress) were placed in the main plots and 14 wheat cultivars were considered in the sub-plots. Traits related to grain yield (including biologic, grain and straw yields and harvest index), grain yield components (including number of spikes per square meter, number of grains per spike and 1000-grains weight) and some physiologic characteristics of flag leaf (including flag leaf area, chlorophyll a, chlorophyll b, total chlorophyll, chlorophyll a/b ratio and proline content) were measured.
The effect of drought stress on biological yield, grain yield, harvest index, number of grains per spike, 1000-grain weight, flag leaf area, chlorophyll a/b ratio and flag leaf proline content was significant, but on straw yield, number of spikes per square meter, chlorophyll a, chlorophyll b, and total chlorophyll was not significant. Wheat cultivars significantly differed in the mentioned characteristics (except for chlorophyll a, chlorophyll b and total chlorophyll). The damage of post-anthesis drought stress on the grain yield reduction was more than straw yield (41 vs. 9% reduction). Among the grain yield components, 1000-grain weight had greater damage than the number of grains per spike (35 vs. 11% reduction) in response to drought stress. Drought stress decreased the amount of chlorophyll but increased the chlorophyll a/b ratio and proline content of the flag leaf. Correlation analysis showed that under normal moisture conditions, harvest index and chlorophyll content in flag leaf had the highest positive correlation with grain yield. While, under post-anthesis drought stress conditions, the highest positive correlation belonged to the proline content of flag leaf and 1000-grain weight.
Some physiologic characteristics of flag leaf (such as proline) with grain yield components (such as 1000-grain weight) could be used to evaluate wheat cultivars under post-anthesis drought stress conditions.Keywords: 1000-grain weight, flag leaf area, harvest index, proline.

Wheat is a strategic product in the world's food security, as it occupies one-sixth of the world's cultivated area. It is the main food crop in Iran, and at the same time, most of the cultivated areas of this crop face environmental stresses such as drought, which leads to a decrease in its yield. Drought stress causes a wide range of plant reactions, including growth and yield changes. Therefore, to achieve maximum adaptability and yield in drought conditions, the present experiment was conducted in Karaj's climatic conditions on eight important wheat genotypes (obtained from the National Plant Gene Bank of Iran).
The experiment was conducted as a split-plot design based on randomized complete block design (RCBD) with three replicates during the 2016-2017 cropping season in Karaj. The main factor was the irrigation regime at two levels, including control (irrigation based on the water requirement of the plant during the growth period until physiological maturity) and water deficit stress (interruption of irrigation from the beginning of spike appearance until physiological maturity) in the main plots and eight different bread wheat genotypes (Star, Sirvan, Roshan, Mehrgan, 2019, 3737, 3729, and 4228) in the sub-plots. Seed yield and physiological traits, including leaf relative water content, leaf photosynthetic pigment content, leaf proline content, and the activity of antioxidant enzymes (catalase and superoxide dismutase) were measured at the end of grain filling stage; then, data analyzed using principal component analysis and cluster analysis.
According to the results, a significant difference was observed between different genotypes of wheat in terms of reduced biological yield and grain yield under water deficit stress compared to the control treatment. In these conditions, the highest and lowest decrease in grain yield was observed in genotypes 3737 (47%) and 4228 (24%), respectively. Also, the content of photosynthetic pigments and the relative water content of the studied genotypes decreased under water deficit condition. Despite the decrease in "chlorophyll a" content in genotypes 4228 and 3729, there was no significant difference with the control treatment under water deficit conditions. The most changes in these conditions were related to Genotype 3737, with a 25% decrease compared to the control treatment. Contrary to plant performance, leaf proline content increased as an important factor in water deficit conditions. The highest accumulation of proline in leaves was observed in Genotype 4228 at 9.1 times under water deficit conditions and the lowest amount in Genotype 3729 at 2.5 times compared to normal irrigation conditions. The results of principal component analysis (PCA) showed that Genotypes 4228 and 3729 had a negative and significant relationship with Mehregan and Star. The results of agglomerative hierarchical clustering (AHC) demonstrated that Genotypes 3729 and 4228 were placed in Cluster 1, genotypes 1399, 3737 and Sirvan were grouped together in Cluster 2, and Genotypes Star, Mehrgan and Roshan were located in Cluster 3.
Physiological responses of bread wheat genotypes to water deficit stress at the end of the growing season were different, which caused different alternations in the growth and yield of plants under water deficit conditions.

Using plant tissue culture, including embryo culture, is one of the ways to increase genetic diversity to withstand environmental stresses, including salinity tolerance in plants. In this study, the reaction of 20 different durum wheat (Triticum turgidum var. durum Desf.) genotypes, including improved lines, native accessions, and two control cultivars, Zardak and Saji, were evaluated to the induction of callus formation and plant regeneration through mature embryo culture and comparison of callus response to salt stress condition. Murashige and Skoog (MS) medium was used for the mature embryo culture of durum wheat. For assessment of genotypes to salt stress, growing morphogenic calli were exposed to different concentrations of NaCl (0, 4, 8, 12, 16 and 20 dSm-1) added to the culture medium. The response of genotypes to in vitro NaCl stress was analyzed as a 20 × 6 factorial experiment based on a completely randomized design with three replications. Comparison of genotypes for callus induction from mature embryos was based on callus induction frequency, relative fresh weight growth of callus (RFWG), relative growth rate (RGR) and callus growth rate. The relative fresh weight growth of callus (RFWG), relative growth rate (RGR), callus growth rate and necrosis percent of callus were used for salt stress. The analysis of the correlation coefficients of the investigated traits in the conditions of callus induction showed that the callus induction percentage has a positive and significant correlation (P<0.01) with the callus growth rate. A positive and significant correlation (P<0.05) was also observed between the relative callus growth and the relative callus growth rate. Under salinity stress, relative callus growth showed a positive and significant correlation (P<0.01) with callus relative growth rate and a negative and significant correlation with callus chlorosis percentage. The results cluster analysis showed that genotypes 65-12-3-3, 75-5-3-5 and Zardak were classified in the same group of tolerant salt stress genotypes. Based on the results, Zardak variety and accessions 65-12-3-3 and 75-5-3-5 due to high relative growth rate and relative growth of callus and low chlorosis percentage in the saline environment were identified as tolerant genotypes to salinity under in vitro conditions. Genotype 25-25-1-5 had the highest response to adult embryo culture. The results showed a significant variation in the ability to callus induction in the genetic material under salinity stress conditions, which can be used in the durum wheat breeding program. It could be suggested to evaluate genotypes in field conditions and study the relationships between traits in vitro and in vivo. Selected cells and plants provide a tool to determine the mechanisms involved in salt stress tolerance. The laboratory screening method for tolerance to salinity stress of plant genotypes can provide a suitable path for developing salinity-tolerant lines in durum wheat.

Durum wheat is known as the eighth agricultural product in the world and is used for spaghetti and pasta. In many studies, in vitro cultivation methods have been used a lot to shorten breeding periods. The use of different explants in the production of wheat embryo callus is known as the best explant in the production of callus. Plant regeneration from callus is also a complex morphogenetic phenomenon that can be affected by various internal and external factors.

An experiment was conducted to study callus induction and regeneration of immature embryos with 11 genotypes of durum wheat. In the callus induction stage, two characteristics, callus induction, and callus growth rate based on the completely randomized design with five replications were measured. Callus derived from an immature embryo was conducted in MS medium containing BAP (0.5, 1 and 1.5 g/l) using an 11*3 factorial experiment based on the completely randomized design with three replications.

Statistical analysis results in the callus induction stage showed a significant difference in the percentage of callus induction and callus growth rate among genotypes. Mean comparison results indicated that in the callus growth rate of genotypes, respectively Stj3//Bcr/lks4 and Quadalete//Erp/mol/3/unk/4/Mrb3/Mnal, had the fastest growth rate and at percentage callus induction genotype 25-25-1-5 showed the highest induction. In the regeneration phase with different concentrations of BAP growth regulator, genotypes 409 and Stj3//Bcr/lks4 showed the highest regeneration. Therefore, based on the results obtained in this study, Genotype Stj3//Bcr/lks4 was introduced as the best genotype among others due to giving the highest callus induction and regeneration.

Therefore, according to the availability of wheat embryos, it can be considered a suitable source in tissue culture through independent factors, which are located in the nucleus or cytoplasm. According to the results, it could be stated that the success in tissue culture is influenced by various factors such as genotype, type of micro sample, and concentration of growth regulators, which differs from one cultivar to another.

If there is enough nitrogen, potassium and phosphorus in the soil, the crops will have vegetative growth, more leaf area and proper performance. Due to the arid and semi-arid nature of Iran, the amount of organic matter in the soils is low and as a result, they have low nitrogen levels. Most plants suffer from nitrogen deficiency in the conditions, so it is necessary to provide nitrogen through chemical and organic fertilizers. Nowadays, due to the problems caused by the excessive use of chemical fertilizers, biological fertilizers have attracted more attention. Biological fertilizers are defined as a set of preservatives with a large number of one or more beneficial soil micro-organisms or their metabolic products, which are mostly consumed to provide the nutrients required by the plant and create suitable physical and chemical conditions for the growth of the plant. Bio-fertilizers are produced in the form of live inoculum for use in the soil or use with seeds.

The experiment was carried out as a factorial design based on a randomized complete block design with four replications. The factors used included: corn cultivars in two levels (Simon and BC), the second factor was Azotobacter chrocooccum bacteria in two levels (use of bacteria and no bacteria) and the third factor nitrogen fertilizer in two levels (use of nitrogen and no nitrogen). The attributes of plant height, number of ears per plant and per unit area, number of seeds per ear, ear diameter, weight of seeds per ear, fresh weight, dry weight, hundred seed weight, total harvest index, ear harvest index, seed yield per hectare, leaf area index, chlorophyll a, and chlorophyll b were measured.

Based on the results of mean comparisons, the highest plant height, the number of ears per plant and per unit area, the number of seeds per ear, ear diameter, weight of seeds per ear, fresh weight, dry weight, weight of 100 seeds, total harvest index, seed yield per hectare, leaf area index, chlorophyll a, chlorophyll b were obtained in the treatment of nitrogen and azotobacter in the Simon cultivar, and the lowest amounts were recorded in the control treatment (no nitrogen and no azotobacter) in the B.C. cultivar.

The use of nitrogen fertilizer along with seed inoculation with azotobacter in comparison with no seed inoculation leads to an increase in plant height, number of seeds in a row, ear diameter, number of leaves, harvest index, leaf area index, fresh and dry weight of leaves and finally grain yield. Combining biological and chemical fertilizers seems to be a suitable method for improving the performance and consumption of nitrogen fertilizers. Therefore, to increase grain yield, it could be suggested that azotobacter be used in seed inoculation and optimal use of nitrogen to reduce the consumption of chemical fertilizers.

Knowledge of genetic diversity and understanding the genetic architecture of complex traits in crop species can play a key role in exploiting genetic diversity and lead to crop cultivation's adaptation, development and expansion in different geographical environments. In this regard, identifying and utilizing informative markers related to quantitative traits is a prerequisite for application in breeding programs and marker-assisted selection (MAS). One of the major uses of these markers is the construction of genome-wide molecular maps and genetic analysis. In genome-wide association study (GWAS) and genomic selection (GS), determining of extent and level of linkage disequilibrium (LD) is critical in sample size and marker density. Moreover, selection for increasing frequency in new mutations advantageous only in a subset of populations leaves some signatures in the genome. Locations of selection signatures are often correlated with genes and QTLs affecting economically important traits. Access to next-generation sequencing technologies, high phenotypic data and a variety of sophisticated statistical tools have enabled LD-based association mapping studies in plants to identify gene loci controlling quantitative traits successfully. Alternatively, the LD-based association mapping (AM) approach can enhance the genetic map resolution to a greater extent due to the representation of a more comprehensive gene pool and more recombination events in history. Association mapping has emerged as a tool to resolve complex trait variation down to the sequence level by exploiting evolutionary recombination events at the population level. Genetic diversity, LD extent in genome and intra-population relationship, determine quality of mapping, marker diversity, statistical methods and power of mapping. LD in population is the foundation of GWAS, whereas it is always affected by genetic drift, population stratification and natural selection. Of the above threats, population stratification is recognized as a major one to the validity of GWAS results. Therefore, knowledge concerning the pattern of LD is essential for performing GWAS and GS. As one of the most important measures in population genetics, LD is the basis of effective population size estimation, genomic selection, GWAS study and QTL mapping. Due to the importance of LD method in mapping studies of the quantitative traits, in this review, we will present LD, its application in population, current status, limitations, and its use in plant breeding, especially in cereals. Finally, this study provides some information about commonly used statistical software and packages in the calculation of LD, and the challenges and perspectives of this approach have been discussed in this study as well.