MODERN GENETICS JOURNAL (MGJ)
Year:2015 | Volume:10 | Issue:1|Papers Count:17

Laccase (EC 1.10.3.2) can use to broad range oxidation of phenolic and non-phenolic substrate. Yeast host cell is highly acceptable for the production because of advantages in promoter strength, secretion efficiency, or ease of growth to high cell density and non-toxicity than other host cell resources. The laccase gene from bacillus sp HR30 was synthesized with codon usage of pichia pastoris. Then, it was sub-cloned in expression vector (Ppink-a) and transferred in Pichia pastoris yeast host cell via electroporation method. For certification of cloning, the colony PCR and restriction enzyme digestion was done and the 1542 bp fragment was observed. Then, the laccase gene was expressed under different condition of temperature and copper sulphate concentration. The results showed that at these conditions, the enzyme activation on the syringaldazine substrate was variable. Also, reducing of temperature up to 20oC and 2 mM copper sulphate was caused improving of refolding and activity of enzyme.

Prunus necrotic ringspot virus (PNRSV), Apple mosaic virus (ApMV) and Arabis mosaic virus (ArMV) are important rose viruses in the world. The aims of this study were to detect the viruses and evaluate their distribution in damask rose floricultures of the Isfahan, Kerman and Markazi Provinces. To do this, total 749 leaf samples were collected randomly from floricultures of the mentioned provinces in 2012-13. Presence of the viruses in the samples was checked by DAS-ELISA using specific antibodies against each of the viruses. The result of ELISA test on collected samples in 2012 showed that 4.41%, 2.20% and 6.3% of the samples were infected with ArMV, ApMV and PNRSV, respectively. These percentages for samples collected in 2013, were 2.31%, 5.32% and 4.16%. Because of wider distribution of PNRSV in comparison to ArMV and ApMV, coat protein gene in four isolates of PNRSV was cloned and sequenced. Phylogenetic tree was drawn and recombination analyses were performed. Based on the phylogenetic tree, Iranian isolates were placed in PV96 phylogroup. This is the first report of genetic diversity of PNRSV in Iran and also the first incidence report of ArMV, ApMV and PNRSV in Isfahan and Markazi provinces.

RNA-seq is revolutionizing our ability to characterize abiotic stresses at the transcriptomic levels. In present study, using RNA-seq analysis, we tried to demonstrate the major processes related to cold stress of almond ovary. Results showed that the number of 358 differential expressed genes were involved in macromolecule metabolic process. Besides, to functional analysis of these genes, we determined the domain type of above mentioned genes. Results suggested that these genes have different combination of domains including pkinase, LRR, AP2, phytocorome and many other domains. Many of the detected domains and protein kinases were previously reported in different cold inducible and cold tolerant proteins. In addition, the highly over expression of 40S ribosomal proteins and NAC, sugessts that at least some partion of these genes are associated with cold stress resistance, and might have great potential to be applied in breeding programs for abiotic stress tolerance in almond and other Rosaceae species.

Bread Wheat is one of the most important crops which has main position in nutrition of the world’s population. Powdery mildew, caused by the biotrophic pathogen Blumeria graminis f.sp tritici (Bgt), is one of the most destructive diseases of wheat worldwide. Plants in response to biological agents produce numerous compounds such as reactive oxygen species, phytoalexins and a group of proteins called pathogenesis related (PR) proteins. In this research the expression rate of PAL and some PR genes including PR1, PR2, and PR3 were evaluated using quantitative PCR (qPCR) technique in two resistance and susceptible genotypes in response to Bgt. Primarily, 40 commercial Iranian wheat genotypes were screened with Bgt in seedling stage. On the basis of colonies number formed on samples, Tajan and Flat were selected as resistance and susceptible lines, respectively. To show the role of mentioned genes in powdery mildew disease resistance, these selected lines were inoculated to Bgt and sampling was carried out at 5 time courses. The experiment was performed in 3 independent replicate and the genes expression rate were normalized, in comparing with their relative internal reference gene, actin. Comparisons of genes expression patterns showed that the expression levels of all target genes was increased in both resistant and susceptible cultivars after infection with powdery mildew agent. Investigation of gene expression process showed that the expression level of all target genes in Tajan cultivar increased faster than Falat cv. In both cultivars, the maximum expression level of genes were observed at 24 h after infection to prevent the establishment of hastorium in host cells by Bgt. Maximum expression level of all genes in Tajan resistance cultivar were much higher than Falat susceptible cv. Results of this study direct that these evaluated genes are involved in resistance strategy beside the other major genes. In resistance and susceptible cultivars their transcript levels attenuated at 48 h, but this level in Falat cv. was lower than Tajan cv, consistently. As well, the PR1 transcript levels was major than the other genes. In peak time point the PR1 transcript levels in Tajan resistance cv was 2.2 fold more than Falat susceptible cultivar. This finding demonstrate that PR1 genes has crucial role in wheat powdery mildew resistance.

pomegranate is a rich source of valuable anthocyanin pigments which constitute the colors ranging from pink to dark purple in fruit skin and arils. The UDP-glucose: flavonoid 3- Oglycosyltransferase enzyme is responsible for the formation of anthocyanins in pigmented cells and catalyze the transfer of a sugar moiety into anthocyanidin. In this study, the cDNA of UDP glucose: flavonoid 3-O-glucosyltransferase gene (UFGT), the last enzyme in anthocyanin biosynthesis was isolated from the pomegranate fruit skin and sequenced. Moreover, the expression of this gene in three distinct pomegranate accessions (Poost Siyah Yazd, Malas Isfahn and Shirin Shabad Shiraz) differing in skin color and pattern of color accumulation was monitored. The deduced amino acid sequence of the cDNAs showed high homology to the sequences of other plants in the GenBank database such as grape and peach.Our results demonstrated that the expression of PgUFGT was relatively high in flowers in the three accessions, but increasing as the fruit continued to develop and peaking in Poost Siyah Yazd over-ripe fruit.Unlike this accession, in Shirin Shaabad Shiraz, PgUFGT expression was detected at low level in all fruit development stages except flower stage. Moreover, the expression of PgUFGT declined in young developing fruits, gradually increased again as the fruit matured and reached its peak when the fruit was over-ripened in Malas Isfahan. Overall, the expression of PgUFGT indicated that the expression of this gene increased dramatically during pomegranate fruit development. Furthermore, the protein structure prediction of PgUFGT showed the similarity of 52% to three dimensional structure of UDP-glucose: flavonoid glycosyltransferase in red grape. The obtained data suggested that both of proteins likely act in similar way to produce anthocyanin. Also, the results obtained from PgUFGT expression in three pomegranate accessions confirmed it.

In this study the expression of genes encoding some antioxidant enzymes in leaf senescence process and hypersensitive response in two wheat genotypes (Tajan and 10# line) was evaluated. Leaf samples were collected at 4 senescence stages: s1: starting yellowish of bottom leaves, s2: 20% of average leaf yellowish, s3: 40% of average leaf yellowish and s4: 60% of average leaf yellowish and green leaf as well as leaf sampling in hypersensitivity (HR) followed by inoculation of septoria tritici blotch pathogen (Mycosharella graminicola). Gene expression analysis (CAT: catalase, MT: metallothioneine, GC: glothamine cyntatase) carried out by using real time PCR technique. The results showed TBARM amount (cellular oxidative level) was at the highest level at HR and s4 senescence stage.In contrast, chlorophyll content declined at HR and during senescence stages from s1 to s4 in compare with green leaf. The gene expression ratio of CAT and MT increased steadily during senescence and it was at the highest level at s4 for both genes. Whether, the expression levels for these genes were different at HR.Indeed CAT and MT showed same activity as s2 and s4 respectively. GC transcript levels increased up to s3 and then dropped sharply. At HR, GC activity was same as s2 stage.

freezing tolerance is a major component of winter hardiness and the ability of plant to survive subfreezing temperatures. Recent advances of cloned genes and molecular markers in barley provide molecular breeders with the means to develop new and simple PCR-based molecular markers which can be used to select frost-tolerant genotypes quickly without stress simulation. In this paper, five molecular markers associated with two frost tolerance and vernalization QTLs (Vrn-H1/Fr-H1, Fr-H2 and Vrn-H2) were tested in 24 barley genotypes consisting of winter, facultative and spring habit types. Genotypes were characterized in terms of frost tolerance under artificial freezing test at -12oC. The marker HvBM5A related to Vrn-H1/Fr-H1, resulted to be the best predictor for assisted selection within this germplasm, because there is a highly significant difference in maximum quantum yield of the PSII photochemistry (Fv/Fm) and electrolyte leakage of organized groups in this marker. Also HvCBF3 marker was more associated than HvCBF14 at Fr-H2 with the frost tolerance. The evaluation of Fv/Fm showed that frost stress the photochemical efficiency of photosystem II to decline significantly that generally, this reduction in resistant genotypes (winter and intermediate growth types) was less than susceptible (spring habit). Furthermore frost stress causes serious damages on plants by injuring the cell membrane that investigation on this collection showed significant difference between mean of this trait in susceptible genotypes (spring type) 0.381 and mean of cold tolerant genotypes (winter and facultative growth habit types) 0.174.

Wheat is the oldest and most important agricultural product, with widest farming area in the world. Narrow Genetic base in cultivated wheat lines, prevents yield improvement according to increasing demand. So, introduction and identification of genetic resources from landraces and wild relatives are important in yield and adaptability improvement programs of commercial wheat cultivars. In this study, 45 genotypes of Aegilops umbellulata collection, from 8 different provinces, were used to investigation and identification of genetic diversity, genetic structure and genetic distance between populations. Ten RAPD markers were used to evaluation of genetic diversity. The results showed 97 bands, which 97.94 % were polymorphic. Observed number of alleles was 1.97 and effective number of alleles was 1.53. Data comparison based on Nei’s index, revealed a genetic diversity from 0.14 to 0.28 between different populations. Overall genetic diversity index was 0.32. Gene differentiation indices suggest high gene flow between population and low subpopulation deviation. Azarbayejan accessions showed highest diversity and minimum genetic distances with other populations, thus, Azarbayejan was suggested as center of diversity of Ae. umbellulata in Iran.

Drought stress, as the most important limiting factor, affected the growth and production of crops.In this study, cDNA-AFLP technique was used to identify transcripts involved in drought stress Polyethylene Glycol (PEG) at three levels: 0 (control), -0.9- and -1.4 MPa in Aeluropus littoralis which is the closest family to cereal. Among 21 isolated ESTs, 17 ESTs were obtained with the average length of 270 bp and 80% sequencing efficiency. The nucleotide sequences were compared with those in the GenBank database. Approximately 72% of the ESTs show homology to nucleotide or amino acid sequences in the GenBank database and 5 ESTs show no significant similarity to other protein and nucleic acid sequences identified in other organisms in the GenBank database which considered as novel and potential genes. Totally, 17 ESTs were recorded in NCBI database which are included zinc finger CCCH domain-containing protein, glyoxalase I and aspartic proteinase and the role of them in drought stress were discussed. These results can be effective to understand the molecular basis of drought resistance and genetic engineering in the improvement of crops resistance against stress and the production of resistant plants.

The high molecular weight glutenin subunit (HMW-GS) compositions of 154 Iranian bread wheat landraces were determined by sodium dodecyl sulphate polyacrylamide gel electrophoresis (SDSPAGE).Seeds were obtained from the Field Crops Research and Genetic Resources Unit of the Faculty of Agriculture, University of Tehran. The Iranian bread wheat landraces divided into 14 groups based on allelic compositions and 12 alleles were detected. The genetic diversity of Glu-B1 locus (0.274) was the highest compared to others loci. The Glu-1 loci score ranged from 4-10. The Most frequent HMW-GS compositions were N, 7+8 and 2+12. By studying the allelic diversity of high molecular weight glutenin subunits (HMW-GS), characterization of genetic resources and quality of wheat varieties can be done for improving the bread-making quality of hexaploid wheat.

GmOSBP and GmBZIP genes play a key role in signaling of response to certain physiological repercussion to stress and CAT gene plays a role through inhibition of ROS in plants This research was conducted to evaluate the expression profile of GmOSBP, CAT and GmBZIP genes in two soybean cultivars, Williams (tolerant) and L17 (susceptible) at salinity condition. Two soybean cultivars were cultured based on completely randomized design with three replicate in the greenhouse conditions, and salinity stress including zero (as non stress) and 300 mM NaCl treatments was performed at five-leaf stage plantlets for 7 days. The sampling from both from leaves and roots for two cultivars was performed in the same time at five-leaf stage. Total RNA was extracted from leaves and roots of both control and stressed plants. Then cDNA was synthesized and used for Real time PCR in two replications. To normalize data the housekeeping gene 18SrRNA was used. Data analysis based on Ct observations showed that the expression of three genes was increased under salinity stress in both leaf and root organs. The expression of GmBZIP gene in Williams was two-fold greater than L17 cultivar and its expression was higher in root than leaf. Also GmOSBP gene expression in L17 was two-fold greater than Williams’s cultivar and its expression was tree-fold greater in leaf than root. The expression of CAT gene in L17 was tree-fold greater than Williams and its expression was higher in root than leaf. According to increasing of salinity tolerance through the expression of these genes, it can be concluded that transferring of these genes may enhance salinity tolerance in other crops.

Quinoa (Chenopodium quinoa willd.) with excellent medicinal and alimentary properties is a plant native to the Andes mountains in South America. Quinoa recently has been subjected to investigation in Iran and a good adaptability was observed in some regions in the country.Availability of genetic variation within a germplasm is essential for plant breeding practices and releasing of new cultivars suitable for each particular agronomic condition. In the current study, genetic diversity of 45 quinoa morphotypes was evaluated using 11 microsatellite markers. Among all used microsatellites; QGAA001, QAAT074, QAAT084 and QAAT097 markers revealed the greatest number of allelic variation. Totally 27 alleles were distinguished using SSR markers and the average of observed polymorphism was 2.4 alleles for each locus. The average PIC of the samples was 0.62, the highest was 0.88 and the lowest was 0.18 in which were pertained to QAAT074 and QCA26 respectively. Cluster analysis grouped the morphotypes into 3 main clusters. Cluster A consisted of morphotypes from both Santamaria and Sajama accessions whereas the majority of morphotypes fallen into cluster C belonged to Sajama accession and only one morphotype belonged to Santamaria accession. The grouping results using PCoA analysis was similar to the cluster analysis indicating that both approaches can successfully be used in categorization studies. The results revealed the high genetic diversity presented among the quinoa morphotypes used in this study and, thus, provides substantial step forward for introducing quinoa in Iran as a new crop. In addition, the results of this research could be beneficiary for quinoa breeding and further genetic investigation and adaptability of quinoa.

OmpF is one of outer membrane proteins of gram negative bacteria that facilitates uptake of fluoroquinolones, including ciprofloxacin inside cell. Thus, regulating of ompF expression is an effective mechanism of antimicrobial resistance. The aim of this research was assessment of ompF expression in gyrA and gyrA marR double mutants. Expression of ompF gene in above mutants with low to intermediate MICs for ciprofloxacin and tetracycline in comparison with wild type, MG1655 was assessed by real time PCR. Statistical analysis of results obtained from real time PCR reaction did not show significant difference between expression of ompF in mutants and wild type. It was concluded that low and intermediate resistance to ciprofloxacin and tetracycline in investigated mutants did not cause decreased ompF expression.

effective myostatin gene or factor 8 on growth and differentiation operates as an inhibition moderator in growth and development of skeletal muscles. In the resent research, the blood sampling was randomly carried out of 58 Kordi sheep of Northern Khorasan and then the DNA extraction was done by optimized salting-out procedure. A 337 base-pair fragment from exon 3 of Myostatin gene was replicated using a couple of specific primer by polymerase chain reaction (PCR). It was used of restriction enzymes HaeIII, in order to tracing of different allelic forms of the Myostatin position by PCR-RFLP method. Of three possible genotypes in exon 3 of GDF8, it was observed only two genotypes of Mm and mm that their abundance was 0.15 and 0.85, respectively. Allelic frequency for M and m alleles was estimated 0.08 and 0.92, respectively. The analysis of linear models with fixed effects of genotype shows that the weight of lamb’s average in 3 and 6 months of heterozygote genotype is higher than the homozygote.