مجله بیوتکنولوژی کشاورزی
سال هفتم شماره 2 (پیاپی 18، تابستان 1394)

Body weight is one of the most important traits which has quantitative traits inheritance. QTL mapping not only provides useful information regarding the status and number of genes controlling quantitative traits. It can also help the breeders in marker assisted selection. This study aimed to identify QTL affecting body weight on chromosomes 1 and 5 in Markhoz goats using microsatellite markers. The studied population consisted of 248 animals from 8 half sib families genotyped for eleven microsatellites on chromosomes 1 and 5. Phenotypic data including body weight at birth, 4 months, 6 months, 9 months and 1 months of age that were corrected for fixed effects including year of birth, sex and type of birth. QTL analysis was performed based on regression analysis. In this research significant QTL was not found on chromosome 1 while one chromosomal regions associated with weight at 4 and 6 months of age was found on chromosome 5. The location of the detected QTL was 2 to 8 cM near the OarFCB005 marker. Maybe this region of chromosome 5 contain genes that affect growth trait. Considering that the locations of this QTL was near the location of OarFCB005 marker, if further investigation were performed, this marker could be used in marker-assisted selection.

The ecological balance of microbial community (MC) is very important as an index of soil ecosystem health due to the pivotal and vital roles of MC. Microbial culture methods are suitable for this purpose because the majority of soil microorganisms unable to grow on synthetic media. DNA- based molecular analyses just provide microbial diversity data. Then a rapid method for the assessment of soil microbial community structure is using phospholipid fatty acid (PLFA) patterns (because they are degraded rapidly after cells death, by the action of phosphatases). Moreover, PLFA provide broad information dealing with microbial diversity, biomass and their nutritional - physiological status. Certain and specific PLFAs, viz. trans/cis ratio (monounsaturated fatty acids (16:1w7t) to 16:1w7c), cy/pre ratio (cyclopropyl (cy17:0 + cy19:0) fatty acids to precursor (16:1ω7+18:1ω7) fatty acids, S/M ratio (saturated to monosaturated fatty acids), G-/G+ ratio (Gram negative bacteria to Gram positive bacteria fatty acids), F/B ratio (saprophytic fungi to bacterial fatty acids) were able as indicators of physiological or nutritional status in different environmental conditions, such as heavy metals polluted soils, changes in pH, depth changes drought, salinity, different agricultural managements and flooding are used for evaluation of soil microbial communities. The method of PLFA analysis in soil and determining microbial diversity, biomass and their physiological status are addressed here.

Different enzymes in oyster mushroom (pleurotus oatreatus), can degradate the lignin compounds from the beginning of mycelium growth up to the end of fruiting period in the environment of compost and straw. Wood, plant residue and most of plant wastes in the nature are called lignocelluloses compounds. Lignocelluloses is mainly composed of cellulose, hemicelluloses and lignin. Magnesium peroxides enzyme (EC:1:11:1:13) is one the most common peroxides destroying lignin which produced by most wood decomposing mushrooms as well as many compost decomposing mushrooms. In order to cloning of mnp gene, RNA was extracted from oyster edible mushroom (P.ostreatus var.florida) and cDNA was synthesized and then the primers was designed based on the sequence of the mnp gene using Primer Premier (V.5.0) software and was amplified by PCR. First, the gene was inserted to pTG-19T plasmid and was confirmed using sequencing. In continue, mnp gene was cloned in to the p13H88 plasmid. Then it was transferred to Ecoli (DH5a) using ice-melt method and its presence was confirmed by enzymatic digestion. The results showed that mnp gene is cloned in p13H88 plasmid and the recombinant plasmid was named as p13H88-FM.

Thioredoxins (Trxs) are low-molecular-mass proteins with two cysteins in their active site WC(G/P)PC which are involved in reversible reduction of disulfide bonds. In contrast to animals and prokaryotes that typically possess one or a few genes encoding Trxs, higher plants contain eight different Trx types: f, m, x, y, z, o, s, and h. The cytoplasmic type Trxs (htype) constitute a particularly large subgroup in higher plans. For instance, in the genome of rice (Oryza sativa), nine genes encoding putative Trx h were identified. Among these nine isoforms OsTrx20, OsTrx1, OsTrx18 and OsTrx23 have additional cys residue in N-terminal in comparison to the other OsTrxh isoforms. In order to study the critical role of this cysteine in OsTrx23, we replaced it with serine using site-directed mutagenesis. Both wild type and mutant proteins were heterologously expressed in Rosetta (DE3)- a strain of Escherichia coli-. The purification of these proteins enabled us to compare their activity in reaction with insulin as substrate. In addition, the dimerization of proteins were analysed under non- reducingand reducing condition with DTT. The results shows that whereas wild type OsTrx23 is present in monomer and dimer forms, the mutant OsTrx23(C11S) is dominantly in monomer form. The activity of mutant was almost similar to wild type. These results suggest that Cys11 at the Nterminal of OsTrx23 has a key role in dimerization of this protein by creating disulfide bonds.

In this study, 10438 ESTs related to the interaction of wheat and Mycosphaerella graminicola (Anamorph: Zymoseptoria tritici) were searched in the NCBI Genebank. Non redundant related proteins of these ESTs in NCBI genebank were analyzed by Seqtools software. Among 10438 studied ESTs, 3868 ESTs had related proteins. These proteins were classified in 279 groups based on the type of them. Related pathways of 112 proteins were found in the KEGG site. Subsequently, the relationship between 55 searched pathways and defense responses in plants interact with pathogen was studied. Finally, these pathways were classified based on function in eight groups. Functional groups were included: Defense, Energy, Metabolism, Protein Process, RNA process, Cell cycle, Protein degradation and Signaling. The maximum number (745) of ESTs was related to proteins involved in cellular energy group, following by functional pathway related to defenses with 251 ESTs.

Anthocyanins have attracted a great deal of attention due to their capability in protecting humans from chronic diseases. Production of natural anthocyanins from fresh plant material has many limitations. Therefore, the use of plant biotechnology for the production of anthocyanin has been put in focus without such restrictions. From the other hand, some crab apples can produce anthocyanins in their various organs including in vitro-grown calli. In this study, in order to investigate the effect of different concentrations of magnesium and nitrogen source for in vitro production of anthocyanin from callus cultures of a red-fleshed crab apple genotype, leaf explants from in vitro grown seedling was exploited to produce callus. After obtaining sufficient amount of callus, treatments were exerted in a completely randomized design with four replications for each treatment and four explants in each replication. With addition of magnesium to the medium, anthocyanin production increased up to 3 fold and the highest concentration of anthocyanins was observed in the highest magnesium concentration (40 mM). However, at this concentration, the growth rate of calli reduced. Changing the nitrogen source also affected the production of anthocyanins. The best nitrogen source for the production of anthocyanin was the combination of 10 mM ammonium and 18.8 mM nitrate as well as 5 mM ammonium and 25 mM nitrate.

Due to complex identification of young pistachio cultivars (Pistacia vera L.) using morphological traits, advance molecular tools have provided a new prospect for DNA fingerprinting. In this study, specific molecular keys were identified for 10 Iranian pistachio cultivars (Pistacia vera L.) using 25 SSR markers and 12 specific AFLP primer combinations. Out of 25 SSR markers, 4 were polymorphic. However, specific molecular keys were not identified using SSR markers. Twelve specific AFLP primer combinations produced specific molecular keys for 10 Iranian pistachio cultivars. Out of 12 specific AFLP primer combinations, two produced specific molecular keys for 8 Iranian pistachio cultivars. Specific primer combination “M_TTT-E_GTC” in Abbasali, Ahmad Aghaei, Badami Sefid, Fandoghi 48, Momtaz, Ohadi va Shahpasand, and “M_GAG-E_CAG” in Akbari produced specific keys. The results showed that there was a similar genetic background within Iranian pistachio mother's trees. The specific molecular keys were verified on 36 pistachio mother's trees and the results were confirmed at two independent laboratories. The reported specific molecular keys can be used for identification of 10 Iranian pistachio cultivars.

The citrus bacterial canker is among the most important diseases in Mexican lime gardens in southern area of Iran. The disease is caused by Xanthomonas citri subsp. citri (Xcc). The PthA bacterial protein, as an effector protein, is crucial in pathogenesis pathway. Inhibition of its functions through antibody could lead to suppression of disease in plant. The present study was performed for isolation and expression of pthA gene and purification of PthA recombinant protein. For this aim the specific primers were designed for isolation of 606 bp of hydroxyl terminal of this gene followed by PCR amplification and cloning into pTZ57R/T vector. The coding sequence of truncated pthA was inserted to pET28a bacterial expression vector as a C-terminal fusion to 6XHis tag. Production of recombinant protein was performed in BL21(DE3) strain of E. coli. The purification was done under native condition through affinity chromatography on nickel column. Total yield was assessed by SDS-PAGE and western blotting analysis. The results confirmed production of PthA recombinant protein with molecular weight around 26 kDa. The purified recombinant protein could be used as an antigen for production of recombinant monoclonal antibodies.

In order to evaluate the effects of five concentrations of sucrose and PEG (0.0, 0.05, 0.11,0.17 and 0.23 mol.l-1) on microtubers initiation, formation and growth, a factorial experiment based on completely randomized design with five replications was carried out. Nodes fromin vitro potato shoots were cultured for microtuberization in MS medium and incubated in constant darkness in growth room at 20 ±1oC. Analysis of variance revealed significant differences among treatments for initiation and formation of microtubers, as well as microtuber length, diameter and fresh weight. Sucrose was more effective than PEG on microtuberization process (all traits).Thus; it was show that application of high levels of sucrose was very useful in microtuberization. Increasing of sucrose concentrations efficiently improved in vitro microtubers number without negative side effects on fresh weight and size. PEG led to a decrease in microtuberization (all traits). High concentrations of PEG restricted microtuberization due to reduce water and nutrients uptake by node explants from medium. On the other hand the use of high concentrations of osmotic material was significantly effective on microtubers dormancy which is useful in germplasm exchange. Results of study like the one presented here, could be very useful in better understanding of physiological mechanisms involved in microtuberization process and useful in approaching breeding objectives as well.

Phosphoenolpyruvate carboxykinase (PEPCK) catalyzes the first step of the gluconeogenesis cycle. There are two isozymes forms of this enzyme, cytosolic phosphoenolpyruvate carboxykinase (PEPCK-C) and mitochondrial phosphoenolpyruvate carboxykinase (PEPCK-M) which present in mitochondria and cytoplasm Current research has been conducted to identify the allelic polymorphism in the PEPCK-M gene in breeder hens of native fowls and commercial broiler and layer chickens. Blood samples were collected randomly from 150 birds of three strains and DNA was extracted using modified salting out method. A fragment of 401 bp in length was amplified from exon 9 of PEPCK M gene. For genotyping of each sample the PCR products were digested by AccI restriction enzyme. The frequency of AccI- and AccI+ allele was estimated at 0.73 and 0.27 in native fowls population, 0.6 and 0.4 in broiler and 0.85 and 0.15 in layer lines, respectively. Two genotypes of AccI -/- and AccI -/+ were observed with the frequency of 0.46 and 0.54 in native fowls, 0.20 and 0.80 in broiler line and 0.70 and 0.30 in commercial layer line, respectively. The comparison of allelic frequency showed no statistical differences between native fowls population with broiler and layer lines, but this results indicated significantly differences between broiler and layer lines (P<0.05).The comparison of genotypic frequency showed that there were significant differences between three populations (P<0.05). No any AccI +/+genotype was detected in the genotyped samples. The difference in genotypic distribution may be as a consequence of different selection strategies used in these populations.