مجله فنآوری زیستی در کشاورزی
سال نوزدهم شماره 1 (بهار و تابستان 1399)

Extraction of genomic DNA of high quality and quantity from some medicinal plants is difficult. It can be attributed to the presence of high amounts of specialized metabolites biosynthesized into their cells. On the other hand, molecular biological studies require successful extraction of high-quality DNA. Hence, the present study aimed to compare four modified procedures of DNA extraction including Murry and Thompson (1980), Dellaporta et al. (1983), Doyle and Doyle (1990) and Ziegenhagen et al. (1993) from five genera of Lamiaceae. Fresh and dried leaves of Rosmarinus officinalis, Lavandula officinalis, Salvia hydrangea, Dracocephalum kotschyi and Ziziphora tenuior were used for the procedures. Following measurement of the extracted DNA quantity and quality, the DNA preparations were tested by PCR amplification using two inter-simple sequence repeats (ISSR) primers. All of the procedures were capable of extracting DNA from five genera. The modified Murray and Thompson method was found to be the most efficient DNA extraction method, indicating high DNA concentration (570.57ng/µl), good-quality DNA (1.80), affordable cost and less time. The results also showed that the quality and yield of extracted DNA from fresh leaves were relatively higher than those of dried leaves. In addition, banding pattern obtained using 1% agarose gel and PCR showed the same results. According to the data, Murray and Thompson method is recommended for DNA extraction from fresh and dried leaves of five studied medicinal plants.

Tobacco streak virus (TSV) belonging to the genus Ilarvirus, is a multipartite single-stranded, positive-sense RNA virus with an extensive host range, that its damage has been reported in many crops and is introduced as a causal agent of soybean bud blight. In this study, 470 samples of soybean were collected from Golestan and Mazandaran provinces with yellowing, mosaic, leaf distortion, necrosis, chlorosis, and stunting symptoms. TSV infection of samples was tested by DAS-ELISA method and polymerase chain reaction (PCR) with specific designed primers. Results showed 9.3% and 6.2% TSV incidence for Golestan and Mazandaran provinces, respectively. Extract of infected plants was inoculated in Chenopodium quinoa and propagated in Nicotiana benthamiana and Datura stramonium and maintain in greenhouse as a source of virus. Because of soybean podding disorder in north of Iran and possible role of TSV in disorder, for evaluation of soybean cultivars response to TSV, a field experiment was conducted in a split plot design with three factors, virus inoculation, using net for prevent pest activity, and cultivars. Based on the result, TSV infection caused yellowing and stunting, and significant reductions in final height, number of nod, fresh and dry weight, and yield were observed in plants that inoculated by TSV but specific symptoms of disorder did not seen. The highest and lowest plant growth indices were seen in no virus inoculation plus use of net, and virus inoculation plus no use of net treatments, respectively.

In this study, the genetic diversity of nine different indigenous and non-indigenous luffa cultivars with the scientific name Luffa cylindrical was evaluated by analyzing the sequence of chloroplast rbcL gene. After DNA extraction and PCR using rbcL gene-specific primers, the amplified products were sequenced and aligned, and the dendrograms of phylogenetic relationships were plotted. Of the 703 sites identified in the study, 179 sites had deletions and additions (177 monomorphs and 2 polymorphs) and 524 sites without deletions and additions. Also, the average numerical value of the dN/dS ratio was approximately one. Overall, the results of this study showed that according to the dendrogram obtained from cluster analysis, and genetic distances, not only with the help of rbcL marker, the diversity within species of this plant species can be well evaluated, but these genotypes can be to some extent classified based on geographical area. It is suggested that the diversity of these genotypes be examined by other DNA barcodes and compared with the results of the present study.

Citrus greening disease is the most destructive citrus disease in most citrus-producing countries around the world and causes significant economic losses in severely affected areas. In order to reveal the genes that may be involved in disease tolerance, microarray data of gene expression of susceptible and tolerant genotypes to citrus greening disease with accession number GSE30502 was retrieved from GEO repository database NCBI and analyzed by RapidMiner Studio 7.6 by eight weighting algorithms (Information gain, Information gain ratio, Deviation, Correlation, Chi squared statistic, Gini Index, Uncertainty, Relief). After identifying key probes by weighting algorithms, heatmap and hierarchical clustering were performed to evaluate the strength of detected probes in differentiating tolerant and susceptible genotypes using Clustvis web tool. Applying weighting algorithms resulted in the identification of 145 probes. According to the heatmap, there was a significant contrast in the expression of the identified probes between the samples of the tolerant genotype and the samples of the susceptible genotype. Also, based on hierarchical clustering, samples related to tolerant genotype and susceptible genotype were placed in separate clusters. A search to determine transcription factors and protein kinases using the iTAK site resulted in the identification of five transcription factors and four different protein kinases. Besides, the abundance of transcripts of the genes involved in hypersensitivity response were higher in the tolerant genotype than in the susceptible genotype, suggesting the activation of hypersensitivity reaction as one of the principal defense strategies against citrus greening disease. The genes identified in this study can be introduced as key candidates to investigate their exact role in citrus greening disease tolerance as well as potential targets for biotechnology approaches.

In this study the the genetic diversity and population structure of six population of hyssop was evaluated using ISSR markers. The evaluated populations included 5 native populations from Iran (Arak, Shiraz, Mashhad, Isfahan1, Isfahan2) and one population from Sweden. Eighteen primers were analyzed resulting in 126 high-resolution bands, among which 119 bands were polymorphic. Cluster analysis on the basis of Jaccard genetic similarity matrix and UPGMA method divided hyssop samples into 6 groups. The method of Principal coordinate analysis (PCOA), confirmed the results of cluster analysis to a great extent. The highest genetic similarity was observed between Isfahan 1 and Mashhad populations and the lowest between Arak and Sweden populations. Diversity within populations was analyzed using the mean of Nei’s gene diversity Index (h) and Shannon Information Index (I). The highest and lowest genetic diversity within the population were observed in Isfahan 1 (h = 0.24 and I = 0.36) and Mashhad (h = 0.14 and I = 0.21), respectively. The mean of effective alleles (Ne) to observed alleles (Na) was 0.85. Based on the results of molecular analysis of variance (AMOVA), the mean indices of Gst (gene diversity) and Dst (genetic diversity between populations) were 0.38 and 0.12, respectively, indicating high genetic differentiation between populations. Molecular analysis of variance between populations also showed that 61% of the total genetic diversity in the samples was related to diversity within populations and 39% among populations. In the study of the genetic structure of the populations using STRUCTURE software, the populations were divided into four groups which Swedish population was the purest one. In this study, the employed ISSR markers showed high percentage of polymorphisms and were able to differentiate populations well. So that, this method is suggested to describe genetic differences within and among hyssop populations.

Biofertilizers (biological fertilizers) contain a sufficient number of one or more species of beneficial soil microorganisms. These fertilizers are able to convert nutrients in the soil into nutrients such as vitamins and other minerals in a biological process and to the plant roots. Therefore, this experiment was conducted to investigate the effect of Fulzime biofertilizer containing growth-promoting bacteria on quantitative traits of potato hybrids of Savalan and Satina cultivars under water deficit stress based on a completely randomized factorial design with three replications in Zara Gostar Arta Company in 2017. The first factor was Folsium fertilizer (zero, 30 and 60g/l) and the second factor was polyethylene glycol (zero, -3 and -6 bar). Stress levels in terms of germination percentage, stem length, number of leaves per seedling, root length, number of microtubers per seedling, weight of microtubers per plant and fulzyme levels in terms of germination percentage, stem length, number of leaves per seedling, root length and number of microtubers per seedlings had significant differences. The interaction effect of water deficit stress and fulzyme was also significant in terms of germination percentage, number of leaves per seedling and microtubers weight per plant. Severe stress significantly reduced all traits under study compared to other levels. Root length and stem length had the highest values at the use of 30 and 60g/l fulzyme, respectively. In non-stress conditions without consuming fulzyme, under mild stress conditions 30g/l and in severe stress conditions 60g/l of fulzyme had the highest germination rate. The highest number of leaves per seedling under non-stress conditions was obtained with consumption of 30g/l of fulzium. The lowest microgranular weight per plant belonged to the treatment of not using Fulzime in non-stress conditions and zero to 60g/l Fulsium fertilizer in severe stress conditions. Based on the results, the application of 30g/l of fulzium is recommended to improve the quantitative traits in the production of potato microtubers.

Mitogen-Activated Protein Kinase Kinase Kinases (MAPKKKs) are important components of MAPK cascades, which are associated with responses to various abiotic and biotic stresses such as plant pathogens. Although MAPKKKs have been analyzed in many plants, the role, function, evolution relationship, structure and expression profile of MAPKKKs have not been studied in potato. In this study, 114 MAPKKK genes (StMAPKKKs) were identified and classified in Raf, ZIK and MEKK families accoding to phylogentic analysis. The assignment of each studied gene to the families was confirmed by the analysis of their conserved motifs. Moreover, the analysis of expression profile of the StMAPKKKs transcriptome which are accessible in database of potato revealed their differentially regulations under various stresses. Furthermore, a number of eight StMAPKKK genes which have been repoted to be upregulated in RNA-seq studies of potato under the heat, cold, salinity and pathogen (Phytophthora infestans) were chosen for real-time qPCR quantitations. All selected genes showed an up-regulation in either heat and salt treatments or inoculation with Phytophthora infestans. Results indicated that StMAPKKKs genes play roles in response to different stimuli and can be utilized in stress-plant interaction studies in potato.