Crop Biotechnology
Year:2013 | Volume:2 | Issue:3|Papers Count:8

Rhizomania is one of the most important Sugar beet diseases throughout the world. The disease is caused by beet necrotic yellow vein virus (BNYVV). The Polymyxa betae (Keskin) is the only natural transmitting agent of the disease between the plants. The fungus is an obligate parasite and could not be cultured in the media, then detection of fungus is done normally by microscopic observation. In order to facilitate detection process, present study is done to develop specific antibodies against this fungus by applying recombinant protein. The Glutathione-S-transferase (GST) as a specific immunogenic protein is a critical enzyme expressed in zoospores, sporangia and resting spores and could be a good candidate to develop a serological test for P. betae. For this aim, the DNA region encoding fungal GST gene was isolated and cloned into pET28a bacterial expression vector. Large scale expression of the recombinant protein was performed in Escherichia coli (Migula). Purification was carried out by applying immobilized metal affinity chromatography under native conditions. The purified recombinant GST protein was used for immunization of rabbit. Purification of immunoglobulin was performed by affinity chromatography using protein A column. The purified antibodies were applied for efficient detection of infected plant in serological assays.

Fusarium head blight (FHB) is a disease that causes major economic losses in wheat and the other cereal crops production worldwide. Moreover, contamination of food with the trichothecene mycotoxin deoxynivalenol (DON) produced by Fusarium graminearum is a major health concern for humans and animals because trichothecenes are potent cytotoxins of eukaryotic cells. Trichothecene mycotoxins inhibit translation by targeting ribosomal protein L3 at the peptidyl transferase center. In this study, we modified a Tomato (Lycopersicon esculentum) cDNA encoding the ribosomal protein RPL3 so that the amino acid residue 258 was changed from tryptophan to cysteine and the amino acid residue 259 was change from histidine to tyrosine. All version of the tomato RPL3 were introduced to DON-sensitive pdr5 and ayt1 mutant strains of Saccharomyces cerevisiae. When transgenic yeast were compared for growth in presence of DON, differences in growth rate and survival were observed among the yeasts expressing the modified versions of the tomato RPL3 genes, compared to those expressing the wild-type yeast RPL3 gene. These results could create a new field in developing FHB resistance varieties of wheat through genetic manipulation.

Maize dwarf mosaic virus is one of the most important and most extensive viruses of maize in the world. In this paper investigated molecular comparison and nucleotide sequence analysis of coding regions of proteins of genomic 5' region of Maize dwarf mosaic with other isolate and other cereal potyviruses. In order to this, maize plants with mosaic symptoms and dwarfing were collected from Golestan province. After verification of plants by Elisa test against MDMV antiserum, an infected sample was selected for isolation of total RNA. Amplification of genomic 5' region has been conducted using five specific primer pair that was obtained 5 overlap segments including P1, HC-Pro ،p3, 6k1genes, as well as 5'-UTR region of the genome and was obtained 3460bp. The comparative analysis of nucleotide sequences of MDMV isolates showed that the Iranian (Golestan) isolate have 95.7 and 92.3 percent similarity at the amino acid level and 90.5 and 85.1 percent at the nucleotide level with Bulgarian and Spanish isolates respectively, and with other homogeneous viruses, MDMV had similarity maximum with sorghum mosaic virus from Texas and sugarcane mosaic virus from china 72.9 and 72.3 percent respectively.

In this study, an F2:3 populations derived from the cross between Hiberna and Pfyner was used to analyze the inheritance of yield and its components in barley by generation mean analysis and to map the corresponding QTLs (quantitative trait loci) by microsatellite markers. Generation mean analysis suggested that both additive and dominance effects were important for most of the traits evaluated, but dominance and non-allelic interaction had a more pronounced effect for days to maturity, number of grains per spike, spike length and plant height. The highest heritability was obtained for number of tillers, indicating that this trait is controlled by additive effects. The additive effects played major role in the inheritance of grain yield per plant, since heritability of this trait was low. The linkage map constructed by 159 microsatellite markers covered a total length of about 1030.5 cM. Using the method of composite interval mapping 2, 4, 2, 4, 1, 4 and 7 QTLs were detected for days to maturity, number of tillers, 1000-grain weight, plant height, spike length, number of grains per spike and grain yield, respectively. Ten QTLs had corresponding occurrences with the QTLs reported earlier, indicating that these QTLs are stable across genetic backgrounds. The results of this study also showed that, grain yield per plant controls with several minor genes. It can therefore be concluded that direct improvement of this trait is somehow problematic. Two major QTLs contributed by ‘Pfyner’ on chromosomes 1H and 2H (qgs-1 and qnt-2a) were found to significantly increase number of grains per spike and number of tillers, respectively. Thus, genetic improvement in grain yield would be easier through indirect selection for these QTLs than through direct selection for grain yield.

Nowadays plant cell suspension cultures have become an attractive source for secondary metabolites production. The fluorescent compounds are highly valuable in a variety of fields including environmental chemistry and medical and food industries. In this study the cell suspension cultures of black zira, an important medicinal plant in Iran, was established in MS liquid medium supplemented with 2 mg/l NAA and 0.5 mg/l BA. For isolation and identification of fluorescent compound in cell suspension culture samples, 100 gr dried cell with 100 mL dichloromethane was extracted followed by subjecting on a column chromatography and then preparative TLC. The mobile phase was selected as dichloromethane-methanol; 9:1. The TLC comparison of extracts from different parts of the plant (seed, stem, leaf and root) as well as cell suspension culture samples showed that under UV light 365, a blue fluorescent component is present in cell samples whereas nothing was found in the extracts of arial parts of the plant. It is also found that a little fluorescent component has been released into the medium. Results of this study demonstrated the fluorescent component content in callus is less than cell samples under the same conditions. This compound was identified by Nuclear Magnetic Resonance analysis as one of the coumarin derivatives (6-methoxy 7-hydroxy coumarin). Substitutions a methyl group or hydroxyl on the coumarin structure shifted the fluorescent band.

Quantitative trait loci (QTL) mapping were carried out for 10 rice cooking and eating quality traits, by using a 144 recombinant inbred lines derived from cross between Hashemi and Neamat varieties with microsatellite markers. The ten traits included amylose content (AC), gelatinization temperature (GT), and 8 parameters from the viscosity profile. A total of 23 QTLs were detected for the 10 traits, with at least one QTL and as many as 4 QTLs for each individual trait. At least one QTL for each trait detected on chromosome 6. Several QTL clusters were observed for multiple quality traits. Especially, one QTL cluster near the Alk locus on chromosome 6 includes four QTLs: qGT6, qFV6b, qCSV6 and qSBV6, which control GT, final viscosity (FV), consistency viscosity (CSV) and setback viscosity (SBV), respectively. Most of them were major QTLs. Another important QTL cluster was located at Wx locus on chromosome 6 includes 3 QTLs: qAC6, qMV6a and qBDV6, which control AC, MV (minimum viscosity) and breakdown viscosity (BDV), respectively. Moreover, these QTLs were major QTLs. Our results showed some of cooking and eating quality controlled by tow region on chromosome 6, region of Wx locus and region of Alk locus. Markers linked to these two genes can be used as marker assisted selection in breeding programmes.

One of the most commonly used methods to produce doubled haploid plants is microspore culture. In this research work, the isolated microspore culture system in two rose cultivars i.e. Apollo and Amarosa was investigated. Important factors including isolation media (AB, B), AT3 induction medium with different carbohydrate sources (sucrose, maltose or glucose) and amino acid source (lactalbumin hydrolysate) were studied. Carbon starvation and temperature (heat and cold) treatments as two important stresses alone or in combination with each other for various periods were evaluated on the induction of symmetrical (sporophytic) divisions. A mixture of different developmental stages of microspores was used to initiate the cultures but the majority of them were at late uni-cellular stage. For eliminating bacterial or fungal contaminants, buds were surface-sterilized by immersion in 70% ethanol for 15, 30, 60 Sec. or 3.5% (w/v) sodium hypochlorite solution for 5, 10, 15 min prior to microspore isolation. The best sterilization procedure was observed when microspores were sterilized with sodium hypochlorite (%3.5) for 10 minutes. Two isolation media did not show a significant difference on the viability of microspores. Among induction media tested, in cv. Amarosa, the highest viability was observed in AT3 medium supplemented by glucose. Induction media in Apollo cultivar did not show a significant difference on viability of microspores. Combination of starvation (B medium) and cold (4oC) treatments for 3 days induced formation of pro-embryos in cv. Amarosa. Present investigation reports a protocol for induction of embryogenic developement in rose (Rosa hybrida) microspores.

Another mechanism for reduction the effects of deoxynivalenon toxin (DON) is target site (RPL3 protein) manipulation. In this study amino acid residue 258 is changedFrom tryptophan to cyctein and 259 from histidine to tyrosine in tomato ribosomal protein L3 (LeRPL3) cDNA through Site Directed Mutagenesis (SDM). Transgenic tobacco plants expressing these modified LeRPL3 cDNAs were tested for ability of leaf discs to regenerate and produce callus in the presence of DON. Significant differences in callus induction and ability to undergo regeneration was seen in transformed lines as compared to non-transformed tobacco plants in DON assays. Among the mutant types, marked difference with respect to resistance against DON was observed with regenerants expressing LeRPL3WC/HY; and plants expressing LeRPL3H259Y giving better response than transforments expressing LeRPL3W258C. The results indicate the possibility of increase in DON tolerance (and Fusarium head blight resistance respectively) among the plants based on expression of engineered RPL3.