مجله فنآوری زیستی در کشاورزی
سال هفدهم شماره 1 (بهار و تابستان 1397)

Biological control of plant diseases is one of the effective strategies to reduce harm effects of pesticides on human health and environment. T. harzianum is one of the most successful biological control agents which have antagonistic activity against many of pathogenic fungi. Biocontrol activity of T. harzianum is linked to the amount and type of hydrolase enzymes. In the authors' previous work, chimeric chitinase was constructed by protein engineering and the recombinant protein was transferred to the T. harzianum. In the present study, the biocontrol activity and colonization of improved isolates were evaluated on bean plant and its pathogen (Rhizoctonia solani) in laboratory and greenhouse. The presence and stability of the chimeric protein was confirmed by molecular technique, after that biocontrol assay was done in dual culture method. The results showed that T3 with 82/53 percent inhibition compared to control, was the best biocontrol in invitro. According to the results of greenhouse tests, plants that were treated with T7, T3 and T15 in all stages of measurement (two leaf, mid-term growth, early reproductive stage and harvest stage) showed symptoms less than 40%. Root surface colonization and fungal hyphae penetration was confirmed by molecular studies using primers PF1/R3xba1.

Yellow rust is one of the major diseases of wheat that suffers a loss many farmers. The purpose of the current study, was determining the wheat proteome response to presence and non-presence of yellow rust disease using Two-dimensional gel electrophoresis. Relative resistance indices including infection type, severity of disease, contamination coefficient, surface under the disease development curve and apparent contamination rates are indicators that are used to identify the resistance or sensitivity of different cultivars. In this study, firstly these indicators studied for 14 bread wheat cultivars was obtained during three 7-day readings. The studied wheat cultivars were classified into three susceptible, resistant and semi-resistant groups using cluster analysis based on resistance indices. Accordingly, two Pishgam and Shahriar cultivars were identified as the most resistant and sensitive cultivars, respectively. Changes in the pattern of expression of proteins in two resistant (Pishgam) and susceptible (Shahriyar) cultivars by tow dimensional electrophoresis of were investigated. The results of statistical analysis showed that 27 protein spots out of 442 repeated protein spots showed a change in expression. Significant changes in total proteins, 17 spots, increased expression, and 10 spots decreased expression. In general, the results of this study showed that the difference in proteome pattern of two resistant and susceptible wheat cultivars to yellow rust could indicate a change in the expression of the enzymes and proteins involved in resistance, and possibility to resist pathogenic agents.

Sugarcane (Saccharum officinarum) is the most important industrial sugar producing plant. Both tissue culture practices and transformation are challenging and need to be optimized. A gene encoding a mutansucrase (gtfI) enzyme from Streptococcus downei bacterium was isolated and cloned in a binary expression vector and transferred to sugarcane cultivars, using gene gun and Agrobacterium-mediated transformation. Results of this study showed that Agrobacterium was not able to deliver gtfI gene to sugarcane plants cells, whereas Biolistic gene transfer was successful and resulted in almost 25.7% transformation efficiency. Distance between explant and rupture disc carrying gtfI construct had a significant effect on transformation efficiency with 9cm distance producing the higher number of transformants than 12 cm distance. A combination of sorbitol and mannitol osmoticums showed a profound effect on transformation efficiency. Furthermore, a combination of IBA and NAA auxins had a significant effect on root regeneration. Interestingly, mutansucrase was found active in transgenic sugarcane lines utilizing sucrose by almost 33%. Detailed sugar analysis of a few transgenic sugarcane plants revealed that transgenic plants had a significant lower sucrose (lower pol%) content than untransformed control plants.

In this research,two strain of Trichoderma (T. harzianum and T. viride) and their mutants were used for cellulase enzyme production. The Avicel, CMC(III), Bacterial Cellulose (Iα) and Whatman NO.1 filter paper, were used for cellulase activity assay. Their chemical structural properties were investigated by FT-IR, XRD and SEM compared to Avicel. The molecular weight of cellulase enzymes were studied using SDS-PAGE. The SEM image of substrates, showed more delicacy of BC fibers relative to Avicel and CMC. The diameter ratio of BC to Avicel is approximately 1/30 or less. The FTIR spectroscopy and XRD assessments designated that the produced CMC is carrying a carboxylic group and its crystallinity is 81.84. The crystallinity of the Avicel and BC were demonstrated that the crystallinity index of Avicel (89.15%) was more than that of bacterial cellulose (66.44%). Both species and their mutants produced varying amounts of extracellular proteins in the fermentation medium. In T. harzianum and its mutants, the highest cellulase enzyme activity were showed in Th M7 and Th M6, respectively and in T. viride and its mutants, the highest enzymatic activity of mutant strains was observed in Tv M14, Tv M15, respectively. The results showed that bacterial cellulose is a good substrate for total cellulase activity assay, because it has the highest enzyme activity compared to filter paper and avicel; so it signs a high potential of accessibility for cellulose Iα biodegradability compared to cellulose Iβ.

Catharanthus roseus is a medicinal plantthat is important because of production of many alkaloids, including anticancer drugs, Vincristine and Vinblastine. Vincristine and Vinblastine are among the most important drugs for cancer treatment, but the performance of these two compounds is relatively low and the approximate amount of them in the plant is 0.0005%. Due to the high price of these compounds and small amount in this plant, efforts to increase the production of these compounds have led to extensive researches on this plant. In this experiment to study the effect of fungal extracts and time of the alkaloids production in Catharanthus roseuse cell suspension culture, test factorial randomized complete block designed with three replications. For this purpose, after preparation of cell suspensions, extracts of fungi P. indica, T. tomentosum and fungal extracts combined of P. indica and T. tomentosum at a concentration of 1% v/v was add to the cell suspension. Sampling in the time period (0, 24, 48, 72h) after the treatment were performed, and Vinblastine and Vincristine alkaloids were measured by using HPLC method. The results showed that three fungal extracts, P. indica, T. tomentosum and mixture of the fungal extract (P + T), increases the amount of Vinblastine, compared to the control and the greatest impact has been on fungal extract T. tomentosum. The average comparison of Vincristine between fungal extracts showed that the highest amount of Vincristine in the control samples and fungal extracts decrease the amount of Vincristine.

Fig mosaic disease which is known as one of the most important impacting viral diseases of fig, is caused by several DNA and RNA viruses belonging to the different genera and families. The objective of the present study was to evaluate the presence of the most important RNA viral agents causing the disease including: Fig mosaic virus (FMV), Fig latent virus-1 (FLV-1), Fig cryptic virus (FCV), Fig fleck-associated virus (FFKaV), Fig leaf mottle associated virus-1 (FLMaV-1), Fig leaf mottle associated virus-2 (FLMaV-2), Fig leaf mottle associated virus-3 (FLMaV-3) and Fig mild mottle associated virus (FMMaV) in different fig cultivations of the country. Sample collection was done from 14 provinces and 611 symptomatic fig leaves. The cDNA of the samples was synthesized using the isolated dsRNAs with cellulose column and random primer. The molecular identification was performed using PCR and specific primers for each virus. After characterization of the infected regions, the results of identification of mixed infections by different viruses showed that the FMV. The results indicated that all of the studied provinces and more than half of the collected samples are infected with at least one of the viruses. Some of the viruses such as FMV were not detected in certain visited regions such as Fars province. The FMV and FFKaV with infection rates of 34.7% and 14.2% and FMMaV with 4.4% showed the highest and lowest infection rates, respectively. The highest level of the mixed infection was observed for FMV and the members of the Closteroviridae family.

In this research, the application of new electrophoresis method of polyacrylamide gel using ethidium bromide staining for genetic variation of sunflower genotypes using TRAP novel marker has been proposed which has advantages such as low-cost, higher speed of analysis, analysis of a large number of samples per unit time (196×2) per experiment, as well as the reuse of gels in comparison with conventional methods of silver staining and fluorescent primer labeling. Six fixed and six arbitrary primers produced 19 TRAP markers so that from 116 bands, 109 of them were polymorphic. Mean number of polymorphic bands were 5.79. DNA fragments size were between 25-920 bp. Polymorphic Information Content (PIC) of this marker were between 0.03 and 0.50 (with the average of 0.33). Primer combination (MAX3BR and Sa12) with PIC value of 0.49 was the highest polymorphic locus. The results showed that this method could be very useful not only in detecting products of the microsatellite markers, but also in other markers, such as TRAP, which produces many polymorphic bands.