مجله پژوهش های ژنتیک گیاهی
سال ششم شماره 1 (بهار و تابستان 1398)

Plant Breeding has utilized a wide range of techniques and methods to improve the quality and quantity of plants. The molecular markers are the tools that have provided a new perspective for plant breeding advancements. This article has reviewed the various advantages and uses of molecular markers and the utilization of the high potential of natural polymorphisms within communities, combined with the abilities of conventional plant breeding methods. The marker attributes are not subject to environmental influence and their high frequency of these markers in their number, high structural diversity are as part of their benefits in identifying identities, determining the genetic diversity of species and studying relationship between populations. They may aid in discovering more information about protecting and maintaining genetic stock collections, identifying varieties, determining genes with chromosomal location and the number of genes controlling traits. Genome sequencing, the preparation of physical and genetic maps, genomic fingerprinting of plants are some of the other applications of this tool in plant breeding. The high efficiency of selection with the help of markers in selection of genotypes has been emphasized as the parent of crosses and selection with the help of a marker in breeding programs and genomic selection. New technologies offers new opportunities to shape genetic variation in the improvement of specific plant breeding programs. Nowadays development of next-generation sequencing technology, genome sequencing and high throughput approaches for markers have facilitated EST-derived simple sequence repeat (EST-SSR) marker development as well as single nucleotide polymorphism (SNP) marker. These markers can be successfully employed in accelerating research and plant breeding programs.

The medicinal plant, Trachyspermum ammi is a rich source of active pharmaceutical ingredients with pharmaceutics effects. Microsatellite markers play a key role in the genome and gene expression, especially in secondary metabolite biosynthesis in medicinal plants. For the first time, transcriptome sequencing of this herb medicine was carried out to identify the microsatellite markers of this species. After pair-end sequencing with the Illumina HiSeq 2000 platform, the quality of the reads was evaluated by FastQC software, trimming was performed by Trimmomatic software and De novo assembly was done with Trinity software. In this study, 11,468 unitranscripts (7913 unigenes) were found to contain 13593 potential microsatellites. The most abundant microsatellite types were di-nucleotide (67%) and tri-nucleotide (24%). Also, six repeated SSRs were the most abundant repeats. The predominant sequence was AG / CT (31%). Sixty-five percent of SSRs were belonged to class II (10-20 nucleotides) and 35% to class I (more than 20 nucleotides). The frequency of SSRs found to be approximately one per 10.1 kB of assembled sequence. More than 57 percent of unigenes containing SSRs were blasted with carrot genome. This showed that T. ammi was an Apiaceae family member and had a high similarity to the carrot genome. A total of 3437 unigenes (43%) were categorized functionally, which among them 2,219 unigenes (64.6%) belonged to the "metabolic process" category and 71 unigenes (1.2%) were assigned to the "secondary metabolic process". In this study, 12 genes were detected in the terpenoid backbone biosynthesis pathway, that their transcripts were containing a microsatellite. These SSRs probably contribute to the genes expression and the biosynthesis of metabolites, especially secondary metabolites. The development of these markers can be used for future studies of marker-assisted selection, genetic diversity and construct genetic maps in this medicinal plant.

The transition from the vegetative phase to reproductive phase is the most important event in production and genetic innovation. This phenomenon is influenced by many genetic and environmental factors in plants. According to studies carried out in this field, one of the environmental factors affects the reproductive and flowering process is Trichoderma species, which is abundant in soil. This study was carried out to evaluate the ability of two recombinant Trichoderma harzianum strains containing chimeric chit 42 (with CHBD domain) and wild-type strain to promote common bean flowering and yield increase in vivo condition. To do this, flowering parameters such as a number of flowers, flowering time and effective parameters in yield were evaluated. Also, expression level of some flowering-related genes such as FT and SOC1 were measured using real-time PCR. The results showed that the bean plants treated with recombinant strains had a significantly increased number of flowers and earlier flowering compared to the control and wild type Trichoderma. Also, plants treated with recombinant strains showed a significant difference in the number and weight of the pod compared to the plant treated with wild type strain and non-treated plants. In addition, the plants treated with T13 strain showed more expression levels of the FT and SOC1 genes (with ratio of 3.42 and 3.41 fold respectively) compared to other treatments and control plant. Finally, T13 recombinant strain exhibited a better performance compared to the other strains through a positive effect on flowering and then increased the crop yield.

Glycyrrhiza is one of the important medicinal plants that is in danger of extinction. Search for finding accessions that have a higher glycyrrhizic acid is very important in breeding programs. Functional genomics methods such as EST sequencing prepare the ability to identify consensus gene families among studied species and interpretation of the genome. In this research, 55960 EST sequences of two different species of this plant along with the protein sequences were analyzed in order to identify the molecular aspects and functional analysis of the genome and the gene network involved in the biosynthesis of glycyrrhizin. Also, in order to validation of results, relative expression of four important genes in the pathway of glycyrrhizin biosynthesis including squalene synthase (SQS), β-amyrin synthase (BAS), β –amyrin 11-oxidase (CYP88D6) and UDP-glucuronyl transferase (UGT) were evaluated. After trimming and qualitative evaluation the sequences, 6427 contig sequences and 30895 singleton (37322 unigene) were generated, which covered a total of 26884666 bp (7.06%) of the licorice genome. Genome functional activity showed that most genes play a role in the catalytic activity and cellular and metabolic processes in which these genes interact within cells and intracellular organels. Locating this group of genes showed that the genes involved in glycyrrhizin biosynthesis pathway were localized in endoplasmic reticulum. Results of validation using qRT-PCR showed that in the autumn and in the rhizome tissue, the genes of BAS, CYP88D6, UGT and SQS were up-regulated. The results of this study can be valuable for genomic sequencing, functional groups, genetic diversity and functional genomics of this plant.

Salinity is an important limiting factor in the production of more plants, including rice. Due to the limited amount of cultivated area, identification of tolerant genotypes to environmental stresses and especially salinity is very important. The aim of this study was to investigate the genetic diversity among 114 recombinant lines derived from the intersection of local Tarom × Khazar cultivars under non stress conditions and salinity levels of 8 dS/m in reproductive stage in a completely randomized design. Combined analysis of variance showed that the differences between lines was significant for all traits. Genotypic variation coefficients also showed that the highest genetic variation among the evaluated recombinant lines was related to the number of panicles per plant. In contrast, days to 50% flowering showed the least genetic variation among these lines. In non stress and stress conditions, the highest genotypic and phenotypic correlation coefficient was observed between grain yield and number of fill grain in seedlings. Based on the cluster analysis of grain yield, the lines were classified into four groups under normal conditions and were classified into three groups under salinity conditions. The third-party lines in both cases had a higher average than the overall average. In general, the results of this study showed that there is a significant genetic variation between the studied lines in terms of salt tolerance and this variety can be used in subsequent corrective programs. Accordingly, lines 83, 81, 56, 39, 37 and 89 were the most sensitive lines and lines 107, 101, 16, 100, 84, 98, 47, 32, 14, 29, 95, 63, 5, 49, 92 and 10 were the most tolerant lines to salinity stresses of 8 dS/m and they also had higher yields and yield components. Strained lines are proposed directly for cultivating saline or for transferring salt tolerance to commercial cultivars through future breeding programs.

German chamomile (Matricaria chamomilla L.) is an s an annual plant of the composite family Asteraceae. This plant is native to the Mediterranean region, and some researchers have reported its origins in Asia. The aim of this study was to investigate the genetic diversity of nine chamomile populations using the SCoT marker. Ten SCoT primers were used. A total of 141 bands were produced, of which 140 bands (96.5%) showed polymorphism. Cluster analysis was performed using UPGMA algorithm based on Jaccard's coefficient of similarity. The results of cluster analysis and principal components analysis divided the chamomile population into four groups. The results of the analysis of molecular variance (AMOVA) showed that the inter-group variation was greater than the intra-group variation, so that 55% of variation was related to the diversity among the groups. The results of this study showed that SCoT markers have high efficiency in determining the genetic diversity and relationships of the chamomile populations.

In the present study, in order to evaluate the drought tolerance indices and their relationship with ISSR markers, 12 rapeseed genotypes were studied using a factorial experiment based on completely randomized block design under the three irrigation treatments (control and irrigation after drainage of 60 and 85% moisture content) in the greenhouse of Mohaghegh Ardabili University, Iran. Drought tolerance genotypes were evaluated by quantitative indices including MP, GMP, SSI, STI and TOL. Cultivars in all five of indices at two levels of stress exhibited significant differences. Regarding the results of the mean comparison at both levels of stress, SLMO46 was identified as the most resistant cultivar with the highest amount of MP and STI, and Karun was the most sensitive one with the highest amount of SSI index. According to the results of factor analysis, in the first level of stress, Sarigol32 and Karun were sensitive, and in the second level of stress, Talaye and Sarigol32 were sensitive as well. SLMO46 was known to be resistant to stress in both levels of stress. Phenotypic correlation of grain yield under stress and non-stress conditions was investigated in two levels of stress with 5 drought indices. In first level of stress condition, grain yield had a positive and significant correlation with mean productivity, geometric mean of productivity and stress tolerance index. In the second level of stress condition, the same correlation was observed with the difference that there was no significant correlation between drought tolerance and tolerance indices. Canonical correlation analysis was performed between drought indices and molecular markers. Five ISSR primers (5, 9, 11, 14 and 19) with the highest polymorphic percentages were used for calculation using the first factor coefficients. ISSR-PCR was used to identify some of the molecular markers associated with drought tolerance indices. A total of 106 clear and score-able loci were amplified by 18 ISSR primers, of which 60 bands (56.6%) were polymorphic. Finally, according to the results, these markers can be used in rapeseed breeding programs for drought tolerance.

MicroRNAs (miRNAs), as a group of non-coding small RNAs, play key roles in regulating the growth, development and response of plants to various stresses. In this study, the expression patterns of three drought responsive miRNAs (miR159c, miR160a,b and miR169v) were compared in both drought tolerant (Yaghuti) and drought sensitive (Bidanesefid) grapevine cultivars using qRT-PCR under drought stress conditions. For identification the potential regulatory elements in the promoter regions of investigated miRNAs, the upstream sequences of their pre-miRNAs were analyzed using PlantCARE database. Drought related motifs such as ABREs and ABSs were identified in the regulatory regions of investigated miRNAs. Three transcription factors related to Auxin and ABA signaling were identified as the most important target genes for investigated miRNAs. The expression patterns of studied miRNAs were different affected from miRNA kind and grapevine cultivar. So, the expression of miR159 remained unchanged in Bidanesefid and increased in Yaghuti cultivar under drought stress condition, but the expression of miR160 and miR169 changed reversely in both cultivars. Cultivar dependent expression showed that miRNA responses to drought stress are also different among very relative genotypes with different drought susceptibility. Generally, considering the role of the potential target genes of investigated miRNAs, it seems that the change in the expression of evaluated miRNAs ultimately leads to the better tolerate of drought stress in Yaghuti cultivar.

For the purpose of graphical analysis and estimation of genetic parameters related to yield and its components, six tobacco genotypes was crossed in half -diallel mating design. In this study, the F2 progenies and their parents were evaluated in a randomized complete block design with three replications. The results of analysis of variance indicated a significant difference between genotypes for plant height, leaf number, leaf dry weight and leaf fresh weight. Hayman graphical method was utilized for analysis of data. Diallel analysis indicated existence of an additive and dominant actions in inheritance of all studied traits. Traits including plant height, leaf number, leaf length and width, stem diameter, internode distance and leaf dry weight possessed greater additive effects meanwhile in the leaf weight the dominance effect was greater. The additive gene action detected for leaf dry weight (yield) manifested the influence of selection methods in breeding of this trait. Due to the fact that the fresh weight of the leaf was controlled by dominance effects, so hybrid-based methods are effective in modifying this trait. Also, additive and dominance gene action were contributed in heritability of yield, plant height and number of leaves. Regarding the mean values of dominance degree and results of graphical analysis, the gene action for leaf fresh weight was over-domnance and so, the heterosis phenomenon could be used to increase and improve this trait. For the traits including plant height, leaf number and yield, the gene action type was relative dominance.

Recently, new molecular breeding and genetic engineering approaches have emerged to overcome the limitations of conventional breeding methods in generating disease-resistance transgenic plants. The use of antimicrobial peptides (AMPs) to produce transgenic plants resistant to a wide range of plant pathogens has achieved great success. Among huge number of AMPs, Dermaseptin B1 (DrsB1), an antimicrobial cationic 31 amino acids peptide, exhibits significant antimicrobial activities towards a wide range of pathogens. In order to increase the antimicrobial efficacy of DrsB1, the DrsB1 encoding DNA sequence was either fused to the N- or C-terminus of the sequence encoding chitin-binding domain (CBD) of Avr4 gene from Cladosporium fulvum and constructs (CBD-DrsB1 and DrsB1-CBD) were used for tobacco leaf disk Agrobacterium-mediated transformation. Polymerase chain reaction (PCR), semi-quantitative RT-PCR and SDS-PAGE analysis indicated the integration of transgenes in tobacco genome and expression of the recombinant genes in transgenic plants, respectively. The antimicrobial activity of extracted recombinant peptides were assessed against a number of plant and human pathogens. Both recombinant peptides had statistically significant (P<0.01) inhibitory effects on the growth and development of fungi pathogens. Also, CFU test result showed that extracted recombinant peptides from transgenic plants, had a relatively high inhibitory effect on plant pathogens. The CBD-DrsB1 recombinant peptide demonstrated a higher antibacterial activity, whereas the DrsB1-CBD recombinant peptide performed a greater antifungal activity. In addition, the expression of DrsB1-CBD recombinant peptide significantly inhibited R.solani fungal infection in comparison with Pythium sp. interestingly, fungi with a higher amount of cell wall chitin were more vulnerable to recombinant peptides, suggesting recombinant peptides present a higher affinity for cell wall chitin. Owing to the high antimicrobial activity and novelty of recombinant peptides, this strategy for the first time, could be used to generate transgenic crop plants resistant to devastating plant pathogens.