فصلنامه ژنتیک نوین
سال چهاردهم شماره 3 (پیاپی 58، پاییز 1398)

The roots and stolons of Glycyrrhiza species constitute one of the most important crude drugs in the world and contain a large amount of glycyrrhizin, an oleanane-type triterpenoid saponin. In this study, relative expression of the genes involved in the biosynthesis pathway of this secondary metabolite, including bAS, SQS, CYP88D6 and CYP72A154, was investigated in the presence of methyl jasmonate in a culture of cell suspension of licorice. Methyl jasmonate was added at a concentration of 100 μM in a stationary growth phase to cell suspension.  The control experiment had no methyl jasmonate. The cells were harvested at intervals 24, 48 and 72 hours’. The experimental was done as split plot in time based on completely randomized design with two replications. The results showed that the two bAS and CYP88D6 genes had the highest expression levels in 48 h after the treatment. SQS and CYP72A154 genes, had the highest expression levels in 72 hours after treatment, but the expression of CYP72A154 didn’t show significant difference comparing to control. The bAS and SQS genes showed higher expression than CYP88D6 and CYP72A154 genes, Because the bAS and SQS genes are involved in the initial pathway of biosynthesis and also are involved in the biosynthesis of other saponins, such as soyasaponin, while The CYP88D6 and CYP72A154 genes are involved in the final pathway biosynthesis for glycyrrhizin. The previous studies had been confirmed presence of glycyrrhizin in the root. In this study, the increased expression of the genes involved in the biosynthesis pathway of glycyrrhizin in cell suspension culture confirmed the presence of glycyrrhizin in cell cultures of licorice.

Folliculogenesis is the maturation of the ovarian follicle, a densely packed shell of somatic cells that contains an immature oocyte. On the other hand, TEK signaling plays a very important role in folliculogenesis. It activates Ras/ERK/MYC, PI3K/AKT/mTORC1 and ovarian steroidogenesis activation pathways. These are the main pathways for cell growth, differentiation, migration, adhesion, proliferation, survival and protein synthesis. So, we have developed mathematical models relate to the different TEK signaling in dominant (> 10 mm) and subordinate follicles (< 5 mm) using systems biology markup language in Matlab environment. Simulation denotes the effect of different expression levels of ANGPT1, TEK, MYC, MAPK1, PIK3R1, MCL1 and EIF4EBP1 and increased expression of certain factors in folliculogenesis TEK signaling on each of the two important pathways where levels of pERK, pMYC, pAkt , pMCL1 and pEIF4EBP1 are increased in dominant follicles and pMYC is decreased in dominant follicles. Over activation of ERK and MYC which are the main cell growth and proliferation and over activation of Akt, MCl1, mTORC1 and EIF4EBP1 which are the main cell survival and protein synthesis factors act as promoting factors for folliculogenesis. Finally, the simulation of signaling pathways may give new insights into biological procedures.

New identified alleles of self-incompatibility locus in Pyrus communis and renumbering of these alleles caused some confusion in their evaluation and categorization. Therefore, this research was performed for a comprehensive bioinformatics review on all 27 S-alleles of this species, and identification of S-alleles in some native Iranian pear cultivars. The results demonstrated that 21 S-alleles, out of 27 identified S-alleles are completely sequenced, that subsequently all 13 parts of them, including promoter, signal region, conserved C1 to C5, intron region and 6 intermediate region were aligned. Bioinformatic analysis revealed considerable conservation also in promoter, signal and terminal parts of locus and their appropriateness for primer design, also showed presence of two CAAT and two coherent TAAT boxes in most promoter regions of S-alleles. Cluster analysis of various parts of alleles showed the complete similarity of polypeptide sequence of S119 and S121 alleles, but in intron region they were moderately different, this high similarity level causes some mistakes in S-alleling of some cultivars such as Dargazi. Comparison of the results also demonstrated that due to sequence similarities and gene flow of other species, the specific primer method is not a trustable method for S-alleling in pear. Finally, S-alleling of some cultivars confirmed S104 in two control cultivars, Coscia and Beurre Diel, also in central Asian cultivar, Krus Siehan, also S108 in Krusalun and S107 in Shekkari and Shahmiveh cultivars, while S119 in Esfarayeni and Dargazi. Interestingly, S35 from P. ussuriensis in Shirin Torkan, Konjuni and Kaftarbache cultivars and S19 from P.×bretschneideri as the second allele of Shekkari cultivar were detected that reconfirming the previously reported gene flow of other Pyrus species in the native Iranian pear cultivars.

The aim of this research was to sequence mitochondrial hypervariable region 1 (HVR1) in three Iranian sheep breeds (Lori, Lori Bakhtiari and Arabic) in order to explore the genetic and phylogenetic diversity of these breeds and also to investigate the maternal association of these breeds with other global breeds. Phylogenetic analysis was carried out using hypervariable region 1 (623 base pair) obtained from 33 animals (12 Arabic, 11 Lori and 10 Lori Bakhtiari breeds) from different parts of the Khuzestan province. After DNA extraction, the HVR1 region was amplified using PCR method. Then, PCR products were sequenced and analyzed using bioinformatics softwares. The phylogenetic tree pattern showed that the Lori and Arabic breeds belong to the haplotype group B and the Lori Bakhtiari breeds belonging to the haplotype group A, which could be due to the different biological origin of these breeds. Analysis of molecular variance (AMOVA) revealed 94% of the genetic variation existing among populations and 4% within populations. Also, the highest and lowest average haplotype diversity for Arabic and Lori breeds were calculated to be 0.95 and 0.61 respectively. Moreover, the calculated FST value indicated that there was a maximum genetic variation between the Lori and Lori-Bakhtiari sheep (0.146) and minimal genetic variation between the Arabic and Lari breeds (0.009). These results indicate high-divergence status of the three Iranian sheep breeds (Lori, Lori Bakhtiari and Arabic) and will influence breeding and conservation strategies adopted for these breeds.

The main objectives of this study were to evaluate the genetic diversity in 53 Aegilops accessions belonging to Aegilops tauschii, Ae. cylindrica and Ae. crassa as well as comparison of the efficiency of start codon targeted (SCoT) and targeted region amplification polymorphism (TRAP) markers. A total of 15 SCoT and six TRAP primer pairs amplified 165 and 201 polymorphic bands, respectively. TRAP primers indicated higher values for the number of polymorphic bands, marker index and resolving power compared to SCoT primers. However, SCoT primers showed the higher PIC value than TRAPs. The results of molecular variance analysis (AMOVA) were different for each of the marker systems, so that SCoT markers showed the highest portion of genetic variance referred to intra-species, while based on TRAP markers the highest variance was observed inter-species. SCoT markers indicated the highest values for all genetic parameters compared to TRAPs and Ae. cylindrica and Ae. tauschii exhibited the highest genetic variation based on SCoTs and TRAPs, respectively. Cluster analysis based on each marker systems as well as the pooled data classified all accessions into three main groups. The clustering pattern based on SCoT primers was clearer than TRAPs. Hence, the use of SCoT markers is recommended for population genetic structure analysis and grouping of them, while the use of TRAPs can be recommended for fine mapping studies.

Phytases are enzymes that hydrolysis phytic acid and makes mineral phosphorus available to animals. Natural and recombinant production of this enzyme is one of the important issues in protein engineering field. In this study, the synthetic Phytase gene Bac phy-wild was cloned into PUC57 vector and transformed to susceptible Escherichia coli-DH5α. In order to replace the polar amino acid with non-polar amino acid within the target mutagenic protein; we used a primer designed to tryptophan (S392W) for targeted mutagenicity in the amino acid serine 392 position. After confirming the mutation at the target site, the mutated gene was registered at the NCBI gene bank. To investigate the expression and temperature stability of the mutant enzyme and compare it to WT phytase, the WT and mutated Bac Phy-Mut genes were transferred to pET26b (+) expression vector. Recombinant vector construct was transferred to a to Escherichia coli-BL21. After screening of recombinant clones, SDS-PAGE analysis and protein concentration were used to evaluate the expression of recombinant protein. The results showed the presence of protein with a molecular weight of 42 kDa. Also the study of physicochemical properties of WT and mutated phytase showed that the optimum temperature was unchanged at 55 ° C and the pH was optimized to be 5. Comparison of thermal stability of the mutated phytase in compare with WT at 40, 50, 60, 70 and 80 ° C was improved by 18, 24, 19, 9 and 6%, respectively. Hence, the results of the current research showed that by using targeted mutagenesis method we succeed to improve the mutated phytase with higher thermal stability which could be obtained and mutant phytase could be used in the agricultural and environmental industries as food and feed additive of livestock, poultry and Fish as well as in medical application.

Improving the quality of water and soil through environmental methods and using the potential of plants to eliminate environmental contamination is very important. The Azolla has the ability to accumulate, degrade or remove metals from the environment, therefore, it is a suitable candidate plant for phytoremediation. In this regard, the present study investigates the accumulation of cadmium in three samples including two species of Azolla pinnata and A. filiculoides and a sample collected from Anzali wetland. For this purpose, cadmium adsorption was measured in three samples after 72 hours of treatment with three concentrations of 10, 50 and 500 μM Cd, by atomic absorption spectrophotometer. Mean comparison showed that the highest absorbance of cadmiun at 500 μm was related to A. pinnata (4673.8 mg Kg-1) followed by A. filiculoides (2747.1 mg Kg-1) and Azolla collected from Anzali wetland (2309.4 mg Kg-1). The cadmium absorption rate in all treated samples was significantly different from their control samples. The growth rate of Azolla samples was also decreased in response to elevated concentrations of cadmium. Increasing the expression of metallothionein gene in the samples in response to different levels of cadmium at 24, 48 and 72 hours after treatment showed a significant effect of cadmium on the expression of this gene in the Azolla samples. In general, application of cadmium treatment in the three samples showed that this plant has a high potential for absorption of heavy metals and uses for phytoremediation purposes.

Wild relatives of wheat are one of the important genetic resources for using in breeding programs. In this study, a set of diploid wild wheat possessing D genome (Aegilops tauschii Coss.) were evaluated for some photosynthetic-related traits under two control and water deficit stress conditions. Under stress condition, Analysis of variance showed significant differences among accessions for chlorophyll a, chlorophyll b, carotenoid content, photosynthesis capacity, maximum quantum yield of PSII and seedling fresh and dry biomasses. Water deficit stress reduced all traits and the highest reduction was recorded for seedling fresh and dry biomasses as well as chlorophyll b, respectively. Furthermore, genetic parameters were affected by water deficit stress so that stress condition increased all parameters compared with the control condition. Under stress condition, the genetic gain showed an increasing pattern and it ranged from 3.46 to 54.04. The highest genetic gain value was obtained for the seedling fresh weight (54.04%), chlorophyll b (29.93%) and photosynthesis capacity (26.44%), respectively. In both control and stress conditions, the highest environmental, phenotypic and genotypic coefficient of variances (ECV, PCV and GCV, respectively) were recorded for the last mentioned traits. In general, regarding the high level of genetic diversity in Ae. tauschii species, as well as a high rate of heritability for measured traits, future comprehensive evaluation on this germplasm, is recommended.

This research was performed with the purpose of screening of wheat genotypes and identification of resistance sources to yellow (stripe) rust disease. A total of 284 accessions from bread wheat collection of National Plant Gene Bank of Iran, which were previously received from 19 countries, were evaluated at adult plat stage under natural incidence of the disease in field condition of Sari, Iran. Based on components of resistance, 116 genotypes were selected for study in the second year at the same condition, among them 52 accessions were then chosen for evaluation at seedling stage in greenhouse. This evaluation was performed by four races 38E158A+, Yr27, 174E10A+, Yr27, 6E2A+, Yr27 and 238E190A+, Yr27. The results showed that 9 genotypes including 8252 and 8320 (Iran), 8395 and 8396 (Korea), 8103 (Afghanistan), 8150 (Portugal), 8348 (Algeria), 8426 (India) and 8472 (Turkey) were resistant to all the studied pathotypes. The presence of Yr1, Yr4, Yr10, and YrSP genes were postulated for these genotypes. One genotype from each country of Portugal, Iran, Japan, Korea, Italy and five genotype with unknown origin were resistant only against pathotype 6E2A+, Yr27, which are likely to carry YrSD or YrND. The comparison of resistance reactions in genotypes 8257, 8237, 8259 and 8332 (Iran), 8105 and 8108 (Afghanistan), 8169 (Portugal), 8458 (India), 8152 (Portugal) and 8362 (Australia) and virulence factors in the studied pathotypes suggested the presence of some unknown resistance genes in regarding genotypes. In the present study some genotypes with adult plant resistance were also identified. The total results of this research indicated the potential of bread wheat collection of National Plant Gene Bank of Iran to identify new sources and genes for resistance to yellow rust.

Tomato is one of vegetables of Solanaceae family that consumes in different ways. Tomato breeding for heterosis and resistance/tolerance to drought is an important program started since 2014 at Ilam University, for which identification of more variable parent lines was the main objective breeders. In the current research, in order to evaluate the relationship between and genetic distance of lines and isolation of the best parents among 36 tomato lines (collected from surround the globe) for hybridization, ISSR molecular markers were used. PCR reactions were performed using 11 primers which produced an appropriate and distinguishable model for the 36 studied genotypes. Totally 89 alleles were identified on agarose gel. Total number of alleles per primer ranged from 7 to 10. The highest and lowest polymorphism recorded for marker ISSR17 with 90 percent and marker LBMB B with 71 percent, respectively. Maximum and minimum of PIC were 0.45 (LBMB C, LBMB D, HB12) and 0.38 (ISSR17). The highest marker index (MI) was 3.8 (LBMB A) and 3.51 (primer 809), indicated a higher resolution of the primers compared to the others. Average of marker index was 3. Dice genetic similarity coefficient ranged from 0.10 to 0.96. Results of the research revealed that genotypes SolanumPimpinellifolium, S. Chilense LA1959 and S. ChilenseLA1972 are the best choices for heterosis breeding and crossing with indigenous Iranian varieties.