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To study the effects of foliar application of putrescine (distilled water (0), 0.75, 1.5, and 2.25 mM) and water deficit stress (20%, 40%, 60%, and 80% available soil water depletion (ASWD)) on the physiological, biochemical, and molecular attributes of Salvia officinalis L., a factorial experiment was performed in a completely randomized design with three replications in the growth chamber. The results of Real-Time quantitative polymerase chain reaction (qRT-PCR) analysis showed that putrescine concentration, irrigation regime, and the two-way interaction between irrigation regime and putrescine concentration significantly influenced cineole synthase (CS), sabinene synthase (SS), and bornyl diphosphate synthase (BPPS) relative expression. The highest concentration of 1,8-cineole, camphor, α-thujone, β-thujone, CS, SS, and BPPS were obtained in the irrigation regime of 80% ASWD with the application of 0.75 mM putrescine. There was high correlation between expression levels of the main monoterpenes synthase and the concentration of main monoterpenes. The observed correlation between the two enzyme activities of ascorbate peroxidase (APX) and catalase (CAT) strongly suggests they have coordinated action. On the other hand, the highest peroxidase (PO) and superoxide dismutase (SOD) concentrations were obtained with the application of 0.75 mM putrescine under the irrigation regime of 40% ASWD. Putrescine showed a significant increase in LAI and RWC under water deficit stress. There was an increasing trend in endogenous putrescine when putrescine concentration was increased in all irrigation regimes. Overall, the results suggest that putrescine may act directly as a stress-protecting compound and reduced H2O2 to moderate the capacity of the antioxidative system, maintain the membrane stability, and increase secondary metabolites under water deficit stress.

An effective culture system for producing embryos was developed from isolated microspores of sweet pepper (Capsicum annuum L.). The present work aimed to define exogenous arabinogalactan proteins (AGPs) on the embryogenesis of sweet pepper under the growth environments of donor plants. The experiments were performed with the cultivars Arancia, California wonder, Californium, Camelot, Inspiration, Plato, and Taranto under three different growth environments of the donor plants. The impacts of the different numbers of sweet pepper ovaries (0, 5, 10, and 15) were evaluated on microspore embryogenesis of cultivars in a factorial design with five replications. The simultaneous application of 10 ovaries in stress (sucrose starvation; temperature) and induction media resulted in the highest embryogenesis of the seven aforementioned cultivars under field conditions. Also, the impacts of 0, 80, 120, and 200 mg L−1 gum arabic were studied on embryogenesis in stress and induction media. The treatments containing 120 mg L−1 gum arabic showed the highest embryogenesis across all cultivars, except the California Wonder cultivar in the field conditions. Also, the simultaneous impact of the number of ovaries and concentrations of gum arabic on embryogenesis were studied. A combination of ten ovaries and 120 mg L−1 gum arabic generated the highest embryogenesis in the majority of cultivars. The results revealed that ovaries of sweet pepper and gum arabic had a significant impact on the efficiency of microspore embryogenesis in sweet pepper plants under field conditions.

The phenotypic diversity of the Iranian orchids has sharply decreased during the last 15 years and is currently threatened with local extinction and global climatic changes. Harvest of tuberous orchids has recently been raised in Iran owing to the high prices of orchid's tubers and international demand from neighboring countries. This has caused competition among orchid harvesters in the country so that the orchids are not even allowed to produce seeds. The present study was done aimed at protecting and identifying the phenotypic diversity within Orchis mascula genotypes in Abr forest, which is a natural habitat of Orchis mascula in Shahrood area, Northeast Iran. The morphological variability of 101 collected samples of early purple orchids was analyzed based on a total of 57 morphological traits. In terms of the quantitative traits, the highest and lowest coefficients of variation (CV%) were observed in the number of side tubers (200%) and petal length (14.43%), respectively. When it comes to the qualitative traits, the highest and lowest CVs were observed in lip color (70.90%) and the flower form on the stem (0%), respectively. Among the studied traits, 11 attributes showed a CV value of less than 20%. The highest positive correlations were observed between tuber fresh weight and tuber dry weight (r = 0.908). Applying principal component analysis (PCA) indicated three main components that explained 45.81% of the total variance. The clusters number four and seven could arguably be suitable sources of parents in breeding crosses for tuber fresh and dry weight traits. In summary, this study confirmed that Abr forest would be one of the rich diversity areas for Orchis mascula and our findings might be satisfactorily applied for breeding, conservation, domestication and development of new traits depending on different breeding purposes.

Redroot Pigweed (Amaranthus retroflexus L.) is an important weed that is highly competitive with common bean. The aims of this study were to evaluate the relative expression of a number of antioxidant enzyme and light response genes and investigate the activity of antioxidant enzymes, anthocyanins and photosynthetic pigments of several red common bean cultivar/lines (Sayad, Derakhshan, and Line D81083) in third trifoliate leaf (V4) and pod formation (R7) stages under Redroot Pigweed free and infested conditions. At V4, enzymatic activity and gene expression decreased while with the increase of common bean competition with weeds, the expression of genes and enzymes activity increased. The activity of antioxidant enzymes increased in the R7 as compared to the V4, indicating the production of reactive oxygen species (ROS) upon competition with weed and increased enzymatic activity for the elimination of ROS. Sayad and D81083 exhibited the highest catalase contents under weed interference whereas demonstrated the lowest catalase enzyme amount under weed-free conditions. Thus, response of common bean to competition with weed depends on bean growth stage and genotype.

Echinacea purpurea L. (2n = 2x = 22), is one of the most popular medicinal plants in developed countries containing important secondary metabolites including caffeic acid derivatives, amongst which chicoric acid is the most significant. Chicoric acid has positive effects on human immune system. Hairy roots culture of E. purpurea is an efficient method for producing chicoric acid, because of their rapid growth and higher chicoric acid content compared to an intact plant. In this research, the effects of simultaneous changes in [sucrose and NH4NO3] and [sucrose and Ca(NO3)2] concentrations, as well as the effect of change in KH2PO4 concentration, were studied on the dry weight (DW) of diploid hairy roots of Echinacea purpurea in woody plant medium (WPM). Additionally, the effects of the individual and simultaneous application of phenylalanine (the precursor of phenolic compounds) and methyl jasmonate (the inducer of phenolic compounds synthesis pathway) were investigated on chicoric acid content in diploid and tetraploid Echinacea purpurea hairy roots. The highest DW of diploid hairy roots (0.525 g per 60 mL) was produced by the treatment including 500 mg L−1 NH4NO3 and 60 g L−1 sucrose. Furthermore, results revealed that the tetraploid hairy roots treated with 1 mM phenylalanine for 72 h produced the highest amount of chicoric acid (14.86 mg g−1 DW), while in diploid hairy roots the highest amount of chicoric acid (8.89 mg g−1 DW), was produced by applying 5 mM phenylalanine for 72 h.

Diosgenin as a triterpene with numbers of pharmaceutical applications has been identified in Trigonella foenum-graceum. In this survey, in order to scale up the amount of diosgenin in Fenugreek as a promising alternative of yam, ∆

Mulberry (Morus spp.) has always been a favorite plant of gardners, farmers and researchers because of its edible fruits, medicinal properties and timber. We have investigated the morphological, physiological and chemical variabilities of white (Morus alba), red (Morus rubra) and black (Morus nigra) mulberry genotypes grown in four regions of Iran. Among the genotypes assessed, fresh fruit weight varied between 3.2 and 87.9 g, total chlorophyll ranged from 4.3 to 13.4 mg/g fresh weight and vitamin C content ranged from 5.5 to 64 mg per 100 g. The highest and lowest antioxidant activity was 275 and 1575 mg ascorbic acid equivalent (AAE) 100 g−1 fresh weight, respectively. The population analysis indicated that the Gorgan population, in terms of morphological and physiological traits, and Shahrood population, in terms of chemical traits, had the highest coefficient of variation. Correlation analysis showed that fruit size and taste were positively correlated with foliar traits and that phenolic content was positively correlated with both antioxidant activity and total soluble solids (TSS). Of the indices assessed, the Berger–Parker index was the most valuable because it was able to identify significant differences between populations in terms of the studied traits. The results of this study confirm that Iran is one of the mulberry phenotypic and biochemical diversity areas and provide information of possible use for the development, conservation and selection of parents for cross-combinations in breeding programs.

Thrombolytic and fibrinolytic therapies are effective treatments to dissolve blood clots in stroke therapy. Thrombolytic drugs activate plasminogen to its cleaved form plasmin, a proteolytic enzyme that breaks the crosslinks between fibrin molecules. The FDA-approved human tissue plasminogen activator Reteplase (rPA) is a non-glycosylated protein produced in E. coli. rPA is a deletion mutant of the wild-type Alteplase that benefits from an extended plasma half-life, reduced fibrin specificity and the ability to better penetrate into blood clots. Different methods have been proposed to improve the production of rPA. Here we show for the first time the transient expression in Nicotiana benthamiana of rPA fused to the immunoglobulin fragment crystallizable (Fc) domain on an IgG1, a strategy commonly used to improve the stability of therapeutic proteins. Despite our success on the expression and purification of dimeric rPA-Fc fusions, protein instability results in high amounts of Fc-derived degradation products. We hypothesize that the “Y”- shape of dimeric Fc fusions cause steric hindrance between protein domains and leads to physical instability. Indeed, mutations of critical residues in the Fc dimerization interface allowed the expression of fully stable rPA monomeric Fc-fusions. The ability of rPA-Fc to convert plasminogen into plasmin was demonstrated by plasminogen zymography and clot lysis assay shows that rPA-Fc is able to dissolve blood clots ex vivo. Finally, we addressed concerns with the plant-specific glycosylation by modulating rPA-Fc glycosylation towards serum-like structures including α2,6-sialylated and α1,6-core fucosylated N-glycans completely devoid of plant core fucose and xylose residues.

The corn poppy (Papaver rhoeas L.) is an important horticultural plant and one of the most well-known members of the Papaveraceae. The species is also used for the treatment of various diseases in folk medicine. Using a biotechnological approach, this species can be further improved. However, the corn poppy is a recalcitrant plant species for tissue culture. Here, we investigated what effects explant type and different auxins and cytokinins had on callus induction and shoot formation. Root, hypocotyl, cotyledon, petiole, and leaf explants were cultured on Murashige and Skoog (MS) medium supplemented with different concentrations and combinations of the auxins picloram, naphthaleneacetic acid, and 2,4-dichlorophenoxyacetic acid (0.5 and 1.0 mg L−1) and the cytokinins 6-Benzylaminopurine (BAP) and kinetin (0.1 and 0.5 mg L−1). There was a significant difference in callus induction among the explants and combinations of plant growth regulators at p = 0.001 and p = 0.05, respectively. The best results for callus induction were observed in petiole and leaf explants on MS medium containing 1.0 mg L−1 picloram and 0.1 mg L−1 BAP (89.2%). Adventitious shoots frequently formed when root explants were cultured on MS medium with 1.0 mg L−1 picloram and 0.5 mg L−1 BAP. Over 70.8% of the root explants successfully produced adventitious shoots, but the other explants did not exhibit any shoot formation. Therefore, we found the optimal culture media for regulating dedifferentiation (callus induction) and differentiation (shoot formation) by using several explants.

ISSR markers were applied to evaluate the genetic diversity and differentiation of 270 individuals of 27 Iranian C. melo landraces of various varietal groups include vars. inodorous, cantalupensis, reticulatus, ameri, dudaim. Genetic diversity among the studied genotypes obtained by GeneAlex analysis (H = 0.08, I = 0.12, Na = 0.77, PPL = 22.6%). Cluster analysis divided Iranian melon landraces into two main cluster. Non-sweet genotype (dudaim group) was well separated from sweet genotypes (inodorous, ameri, reticulatus, cantalupensis). The most similar genotypes were BANI and TONI (0.95) and the most dissimilar ones were GER and TS (0.58). AMOVA result showed that the percentage of genetic variation among and within Iranian melon is 69% and 31%, respectively. All landraces evaluated based on 10 morphological traits which revealed the diversity of melon varietal groups. Bayesian analysis assigned ten landraces to Pop 1, eight landraces to Pop 2 and nine melon landraces to Pop 3. Bayesian and UPGMA cluster analyses demonstrated the almost related results. Our results indicated that ISSR markers technique alongside polyacrylamide gel analysis could be helpful to discriminate varieties of melon. Keywords: Orthogonal contrasts analysis, Morphological traits, AMOVA, Bayesian analysis, UPGMA cluster analysis

Artemisinin, a sesquiterpene lactone produced by some Artemisia species, is an efficacious anti-malarial drug, effective against cancer, hepatitis, and schistosomiasis. A. annua is a main source of artemisinin while other Artemisia species produce less artemisinin content. The aim of the current study was to identify the limiting factor of artemisinin biosynthesis in studied Artemisia species, compared to A. annua. The specialized 10-celled biseriate glandular trichomes on the leaves, stems, and inflorescences of some Artemisia species are as a site of artemisinin synthesis. The leaves of five Artemisia species, having different artemisinin contents were assessed in terms of the glandular trichomes density, and area per leaf, and the expression of artemisinin biosynthesis genes and two genes (Aa-TTG1 and Aa-TFAR1) involved in trichome formation. This study identified one novel plant sources of artemisinin (A. deserti, 5.30 mg g−1 DW) that statistically performed as well as A. annua of Iran (6.27 mg g−1 DW), but inferior to A. annua cv. Anamed (14.50 mg g−1 DW) at the flowering stage. A. deserti had the highest trichome area per leaf area accompanied with a high expression of Aa-ADS, Aa-AlDH1, Aa-CYP71AV1, Aa-TTG1, and Aa-TFAR1 genes. A. persica with low artemisinin content had a high density of glandular trichome, high expression of TTG1 and TFAR1, but low expression of artemisinin biosynthetic genes. A. khorassanica with no artemisinin content had a very low density of glandular trichome and gene expression. The artemisinin content of A. deserti is significantly as same as A. annua of Iran and inferior to A. annua cv. Anamed despite having the highest glandular trichome area per leaf, and high relative expression of Aa-ADS, Aa-ALDH1, Aa-CYP71AV1, Aa-TTG1, and Aa-TFAR1. We suggest that it is related to the preferential oxidation of artemisinic aldehyde to artemisinic acid than the reduction of the artemisinic aldehyde to dihydroartemisinic aldehyde, due to the very high expression of Aa-ALDH1 and Aa-CYP71AV1, and the low expression of Aa-DBR2. It is possible to develop high artemisinin producer plant by overexpression of Aa-DBR2 in A. deserti. It is concluded that there is a relationship between the enhancement of artemisinin content and increased expression of some genes.

Extraction of high quality RNA is a critical step in molecular genetics studies. Hazelnut is one of the most important nuts plants in the world. The presence of the taxol and other taxanes in hazelnut plant necessitates explaining their biosynthesis pathway and identifying the candidate genes. Therefore, an easy and practical method is necessary for RNA extraction from hazelnuts. Hazelnut has high levels of phenolic compounds. High amounts of polyphenolic and polysaccharide compounds in plants could be causing problems in RNA extraction procedures. To avoid these problems, a simple and efficient method can be used based on cetyltrimethylammonium bromide (CTAB) extraction buffer and lithium chloride for extraction of high quality RNA from different parts of hazelnut plant. Using this method, a high-quality RNA sample (light absorbed in the A260/A280 was 2.04

Salinity is among the most important abiotic stresses affecting crop production throughout the earth. Halophyte plants can sustain high salinity levels, therefore elucidating molecular mechanisms underlying their salinity resistance is beneficial for crop improvement. Aeluropus littoralis , a halophyte weed, is a great genetic resource for this purpose. Isolated expressed sequence taq (EST) sequences from A. littoralis under salinity stress, have given us the chance to find and analyze transcripts of genes involved in response to salinity. Transcriptome analyses indicated the expression levels of mRNAs corresponding to 10 of sequences were increased under treatments. All mRNAs were significantly induced under salt treatment with the highest peaks observed at different hours of treatments. Moreover, the full-length cDNA of vacuolar H + -pyrophosphatase ( VP ) was isolated utilizing 3′ and 5′ rapid amplification of cDNA ends polymerase chain reaction (RACE-PCR) and characterized (GenBank accession number of KT253223.1). The extracted full-length of VP was 2732 bp, which contained ORF of 2292 bp encoding 763 amino acids.

INTRODUCTION Throughout history, thousands of medicinal and aromatic plants have been widely utilised by people worldwide. Owing to them possessing of valuable compounds with little side effects in comparison with chemical drugs, herbs have been of interest to humans for a number of purposes. Diosgenin, driven from fenugreek, Trigonella foenum-graecum L., has extensively drawn scientist's attention owing to having curable properties and being a precursor of steroid hormones synthesis. Nonetheless, complete knowledge about the biosynthesis pathway of this metabolite is still elusive. OBJECTIVE In the present research, we isolated the full-length CDS of 14 genes involving in diosgenin formation and measured their expression rate in various genotypes, which had illustrated different amount of diosgenin. METHODOLOGY The genes were successfully isolated, and functional motifs were also assessed using in silico approaches. RESULTS Moreover, combining transcript and metabolite analysis revealed that there are many genes playing the role in diosgenin formation, some of which are highly influential. Among them, ∆24 -reductase, which converts cycloartenol to cycloartanol, is the first-committed and rate-limiting enzyme in this pathway. Additionally, no transcripts indicating to the presence or expression of lanosterol synthase were detected, contradicting the previous hypothesis about the biosynthetic pathway of diosgenin in fenugreek. CONCLUSION Considering all these, therefore, we propose the most possible pathway of diosgenin. This knowledge will then pave the way toward cloning the genes as well as engineering the diosgenin biosynthesis pathway.

Drought is a worldwide problem seriously influencing production of agricultural plants such as coriander, but development of tolerant genotypes is inhibited by a lack of effective selection criteria. The objective of this study was to interpret the relationship between fruit yield and related traits in coriander to evaluate genotype by trait interaction by considering genotype × irrigation interaction. According to this aim, 16 Iranian endemic coriander genotypes were grown in a glasshouse under well watered (WW) and water stressed (WS) conditions. Structural equation modeling (SEM) was used, based on genotypic correlation coefficients and genotypic variance estimates obtained from combined analysis of WW and WS conditions and genotype-trait (GT) biplot analysis. Applying both types of analyze to the multiple trait data revealed that SEM and DC + YPr data based GT biplot results are similar. The GT biplot graphically displayed the interrelationships among traits and facilitated visual comparison of genotypes for selection. Predictors in the causal diagram could explain 70.8%, 80.9%, 100% and 41.7% of the total variation in fruit weight per plant (FWPP), fruit number per plant (FNPP), day to end of flowering (DTEOF) and umbel number per plant (UNPP), respectively. Also, the GT biplots explained 81.2% for WW data, 74% for WS data and 79.7% for DC + YPr data. It was found that selection for high SPAD chlorophyll content in the grain filling stage (SCCIGFS), UNPP, FNPP and branch number per plant (BNPP), and low shoot dry weight per plant (SDWPP) and DTEOF should be considered as priorities in breeding programs for coriander aiming for more productive and drought tolerant genotypes. Furthermore, G13 (TN-59-353) can be used as a drought tolerance donor in breeding programs.

The aim of this study was to evaluate molecular and phytochemical changes in four landraces of Rosa damascena with different in color and floral developmental stages. Data showed the highest amount of anthocyanin content in stage B, while it was reduced at stage C during flower development in all landraces. Interestingly, increasing in cellular pH was occurred gradually in all landraces during flower development. Based on essential oil analysis, 2-Phenylethanol was detected in all landraces, except for Lavasanat. Molecular expression pattern of RhPAR gene showed high expression rate in all landraces at stage B with 6 time more than its control, except for Lavasanat. Similarly, the change in the expression rate was occurred in RhMYB1 and the transcript level of RhANS was significantly higher in Kamoo and Minab in compared to Ghalhar, which is consistent with the anthocyanin content. Increasing the RhPAR relative expression could be implied to promoting effect of RhMYB1 which resulted in enhancing the 2-PE component production in the petals of Rosa damascena and expression of RhANS, as a key gene in anthocyanin biosynthesis pathway.

Induction of hairy roots stemmed from Agrobacterium rhizogenes transformation, is a key step to reach mass production of secondary metabolites in medicinal plants. Accordingly, this study is aimed to investigate potential impact of different A. rhizogenes strains, including ATCC15834, R1000, A4 and C58, on diosgenin biosynthesis in high- “Boshruyeh” (23.8 mg/gDW) and low “Hamadan” (6.4 mg/gDW) diosgenin-producing genotypes of fenugreek. Interestingly, comparing the metabolite extracted from both leaves and hairy roots of these genotypes suggested that hairy roots are promising platform to produce scalable amount of diosgenin, producing 143.96 mg/gDW in high diosgenin producing hairy roots (A4-mediated hairy roots induction in “Boshruyeh” (BA4)), while 23.8 mg/gDW in high diosgenin producing genotype. Transcript abundance of 10 genes involved in diosgenin biosynthesis pathway as well as three rate-limiting genes; i.e 1-deoxy-D-xylulose-5-phosphate synthase (DXS), 1-deoxy-D-xylulose 5-phosphate reductoisomerase (DXR) and 4-hydroxy-3-methylbut-2-enyl diphosphate reductase, functioning in plastid 2-C-methyl-D-erythritol 4-phosphate (MEP) pathway were measured. Comparison of the transcription and metabolite analysis revealed that the more expression of Δ24-reductase gene, the higher accumulation of diosgenin in the hairy roots. In high diosge nin producing hairy roots, the expression of Δ24-reductase gene were significantly up-regulated (8-folds). Moreover, integration of metabolite and transcript results revealed that A4 has a strong capacity in boosting diosgenin accumulation in the both genotypes. These results may lead to the next step in production of diosgenin in scalable and commercial levels.

Black caraway is of great importance for its terpene compounds. Many genes are involved in the biosynthesis of secondary metabolites in medicinal plants. For this study, black caraway seeds were collected from five different regions, i.e. [Isfahan; Kerman (Khabr); Semnan; Kerman (Sirch); and Hormozgan]. The black caraway seed oil was extracted and analyzed by means of the gas chromatography method. There was a negatively significant correlation (p ≤ 0.05) observed between cuminaldehyde and gammaterpinene compounds. 3-hydroxy-3-methylglutaryl coenzyme A reductase (HMGR) and isopentenyl pyrophosphate isomerase (IPI) play an important role in the biosynthesis of secondary metabolites. Appropriate primers were designed for these genes based on the conserved regions in other plants. Amplified fragments were then sequenced. Blastn results indicated the similarity of the high RNA sequences between new sequences and other HMGR and IPI gene sequences in GenBank, and it also identified the HMGR and IPI gene sequences of B. persicum. A fragment of the HMGR gene with KJ143741 number was recorded in the gene bank. Quantitative PCR showed that the relative expression of two genes in different growth stages of B. persicum was significantly different between the germination stage and the multi-leaf stage, and also between the germination stage and the flowering stage (p

Artemisinin, an effective anti-malarial drug is synthesized in the specialized 10-celled biseriate glandular trichomes of some Artemisia species. In order to have an insight into artemisinin biosynthesis in species other than A. annua, five species with different artemisinin contents were investigated for the expression of key genes that influence artemisinin content. The least relative expression of the examined terpene synthase genes accompanied with very low glandular trichome density (4 No. mm−2) and absence of artemisinin content in A. khorassanica (S2) underscored the vast metabolic capacity of glandular trichomes. A. deserti (S4) with artemisinin content of 5.13 mg g−1 DW had a very high expression of Aa-ALDH1 and Aa-CYP71AV1 and low expression of Aa-DBR2. It is possible to develop plants with high artemisinin synthesis ability by downregulating Aa-ORA in S4, which may result in the reduction of Aa-ALDH1 and Aa-CYP71AV1 genes expression and effectively change the metabolic flux to favor more of artemisinin production than artemisinic acid. Based on the results, the Aa-ABCG6 transporter may be involved in trichome development. S4 had high transcript levels and larger glandular trichomes (3.46 fold) than A. annua found in Iran (S1), which may be due to the presence of more 2C-DNA (3.48 fold) in S4 than S1.

Papaver somniferum L. is an herbaceous, annual and diploid plant that is important from pharmacological and strategic point of view. The cDNA clones of two putative MYB and WRKY genes were isolated (GeneBank accession numbers KP411870 and KP203854, respectively) from this plant, via the nested-PCR method, and characterized. The MYB transcription factor (TF) comprises 342 amino acids, and exhibits the structural features of the R2R3MYB protein family. The WRKY TF, a 326 amino acid-long polypeptide, falls structurally into the group II of WRKY protein family. Quantitative real-time PCR (qRT-PCR) analyses indicate the presence of these TFs in all organs of P. somniferum L. and Papaver bracteatum L. Highest expression levels of these two TFs were observed in the leaf tissues of P. somniferum L. while in P. bracteatum L. the espression levels were highest in the root tissues. Promoter analysis of the 10 co-expressed gene clustered involved in noscapine biosynthesis pathway in P. somniferum L. suggested that not only these 10 genes are co-expressed, but also share common regulatory motifs and TFs including MYB and WRKY TFs, and that may explain their common regulation.

The species of Artemisia, one of the largest genera of the family Asteraceae, are frequently utilized for the treatment of diseasessuch as malaria, hepatitis, cancer, inflammation, and infections by fungi, bacteria, and viruses. Karyological studies were performedon 18 Artemisia khorassanica populations: eleven were diploid (2n = 18) and seven were tetraploid (2n = 36). The mean chromosomelengths were 3.61 and 3.84 ?m for diploids and tetraploids, respectively. Two chromosome types ("m", "sm") formed karyotype formulas"18m" for diploids and "36m" and "34m + 2sm" for tetraploids. The mean 2C DNA contents were 5.91 and 11.53 pg in diploids andtetraploids, respectively. The transcription levels of key genes involved in artemisinin production were compared in diploid (B, D, H)and tetraploid (O, P, R) A. khorassanica relative to A. annua as a standard species. No artemisinin content was detected in diploid andtetraploid A. khorassanica populations. No significant differences were detected between diploids and tetraploids in terms of DXR,HMGR, FDS, and ADS gene expression. This implies that most of the genomic amplification likely occurs in the amount of repetitiveDNA and not in unique sequences. The DBR2 gene was expressed in the diploid A. khorassanica in a low amount but silenced in theautotetraploid A. khorassanica.

Opium poppy (Papaver somniferum L.) is one of the ancient medical crops, which produces several important alkaloids such as morphine, noscapine, sanguinarine and codeine. MicroRNAs are endogenous non-coding RNAs that play important regulatory roles in plant diverse biological processes. Many plant miRNAs are encoded as single transcriptional units, in contrast to animal miRNAs, which are often clustered. Herein, using computational approaches, a total of 22 miRNA precursors were identified, which five of them were located as a clustered in pre-ribosomal RNA. Afterward, the transcript level of the precursor and the mature of clustered miRNAs in two species of the Papaveraceae family, i.e. P. somniferum L. and P. bracteatum L, were quantified by RT-PCR. With respect to obtained results, these clustered miRNAs were expressed differentially in different tissues of these species. Moreover, using target prediction and Gene Ontology (GO)-based on functional classification indicated that these miRNAs might play crucial roles in various biological processes as well as metabolic pathways. In this study, we discovered the clustered miRNA derived from pre-rRNA, which may shed some light on the importance of miRNAs in the plant kingdom.

Noscapine is an antitumor alkaloid produced in opium poppy (Papaver somniferum) and some members of the Papaveraceae family. It has been primarily used for its antitussive effects; more recently, its anticancer properties were shown. Herein, we detected an SSR embedded in the promoter region of the CYP82Y1 gene, which was found to be the first committed-step enzyme in the noscapine biosynthesis pathway, using the MISA program. Some collected ecotypes of P. somniferum were investigated for understanding of SSRs role in the regulation of gene expression and metabolite content. Quantitative PCR showed that a variation in the motif repeat number (either a decrease or increase) down-regulated the expression of the CYP82Y1 gene. Furthermore, the analysis of noscapine content suggested that a variation in the promoter region influence noscapine amount. Moreover, P. bracteatum was analyzed in both transcript and metabolite levels, and illustrated much less expression and metabolite level in comparison to P. somniferum. By exploiting the transcriptome data from the eight genera of the Papaveraceae family, we found that noscapine biosynthesis genes are present in P. bracteatum and are not shared in other genera of the Papaveraceae family. This results may explain production of a confined metabolite within a genus.

Alliinase is the key enzyme in allicin biosynthesis pathway. In the current study, the identification and sequencing of alliinase genes along with determination of allicin contents were reported for Allium species with a novel report for Iranian endemic species. The presence of different isoforms in the Allium being discovered for the first time. In bulbs tissue, the highest allicin concentration was in Allium sativum, A. umbilicatum, and A. fistolosum (1.185%, 0.367%, and 0.34%, respectively), followed by A. spititatum (0.072%), A. lenkoranicum (0.055%), A. atroviolaseum (0.36%), A. rubellum (0.041%), and A. stamineum (0.007%). The highest allicin content in the leaves and roots were in A. sativum (0.13%), and A. stamineum (0.195%), respectively. The ORFs length ranged from 1416 in A. sativum (iso-alliinase2; ISA2) to 1523 bp in A. sativum (alliinase); the identity with A. sativum (alliinase) varies from 95% to 68% for A. ampeloprasum, and A. sativum (iso-alliinase1, ISA1) respectively. These data suggested that both ISA1 and ISA2 had a high expression in the roots and bulbs compared to A. sativum as the control in all species. Note that ISA1 and ISA2 were not expressed in the leaves. The results showed that isoforms expression patterns among different tissues in Allium species were variable. The presence of various isoforms is a possible explanation for the difference between the species in terms of obtained results, especially the amount of allicin.

The goal of this study is to identify characterization of expressed sequence tag (EST)-simple sequence repeats (SSR) markers from EST library of durum wheat and functional analysis of SSR-containing EST sequences for application in comparative genomics and breeding. 19,141 sequences were analyzed among which 18,937 ESTs were selected. Consistent with MISA results, 313 EST-SSRs were yielded. The final EST-SSRs were compared to the GenBank non-redundant database using BLASTX and classified based on these functions. Results indicated that the perfect EST-SSRs are the most frequent. The TTG/CTG imperfect EST-SSR had gamma-gliadin putative function that can be appropriate for durum wheat. Also, the mononucleotides and trinucleotides were the most frequent. Findings suggested that the identified EST-SSRs could be categorized into 83 types. Motifs TTG in trinucleotides and TC in dinucleotides had the highest frequency. TTG is the new motif in durum wheat identified in this study. We identified new EST-SSRs with more than trinucleotide and detected motifs that have potential to code amino acids. Arginine was the most frequent amino acid. Enzymes had the highest frequency among predicted functions. EST-SSRs have been identified in this study can be used for developing ESS-SSR-based detection tool for durum wheat in future studies and will be a useful resource for molecular breeding, genetics, genomics, and environmental stress studies. Motifs coding amino acids could be used as a new source of functional markers and biological study. In addition to, designed new PCR primer pairs are new resources for to identify useful alleles in transcription factors, storage proteins, and enzymes which incorporated them again into the cultivated material.

A primary mechanism for controlling the development of multicellular organisms is transcriptional regulation, which carried out by transcription factors (TFs) that recognize and bind to their binding sites on promoter region. The distance from translation start site, order, orientation, and spacing between cis elements are key factors in the concentration of active nuclear TFs and transcriptional regulation of target genes. In this study, overrepresented motifs in cold and pathogenesis responsive genes were scanned via Gibbs sampling method, this method is based on detection of overrepresented motifs by means of a stochastic optimization strategy that searches for all possible sets of short DNA segments. Then, identified motifs were checked by TRANSFAC, PLACE and Soft Berry databases in order to identify putative TFs which, interact to the motifs. Several cis/trans regulatory elements were found using these databases. Moreover, cross-talk between cold and pathogenesis responsive genes were confirmed. Statistical analysis was used to determine distribution of identified motifs on promoter region. In addition, co-regulation analysis results, illustrated genes in pathogenesis responsive module are divided into two main groups. Also, promoter region was crunched to six subareas in order to draw the pattern of distribution of motifs in promoter subareas. The result showed the majority of motifs are concentrated on 700 nucleotides upstream of the translational start site (ATG). In contrast, this result isn't true in another group. In other words, there was no difference between total and compartmentalized regions in cold responsive genes.

To estimate the genetic diversity of chickpea germplasm from Iran, a total of 307 landraces from 4 regions including: northern areas (29 from Ardebil, 3 from Qazvin and 5 from Mazanderan provinces), temperate (16 from Kermanshah, 2 from Semnan, 54 from Khorasan and 20 from Kerman provinces), semi-arid (28 from Ghom and 56 from Isfahan provinces) and cold areas (15 from West Azarbayjan, 52 from Tehran and 27 from East Azarbayjan provinces) were analysed using 16 microsatellite loci. The number of alleles per microsatellite locus ranged from 8 to 29, with an average of 19.31 per locus. A high level of genetic diversity in the northern area (He = 0.76), even with a limited number of available landraces (37) compared with the other three regions (84–94), might confirm the northern Persia as part of the chickpea centre of origin. The neighbour-joining tree showed a low relationship between molecular divergence and the geographical grouping of chickpea. Moreover, cluster analyses based on molecular data showed that the northern area was separated clearly from the other three regions, indicating a physical barrier or geographical and environmental differences among these regions. A wide genetic diversity of Iranian chickpea landraces is a critical component for future selection and use of this germplasm for future breeding of chickpea.

The artemisinic aldehyde double bond reductase (DBR2) plays an important role in the biosynthesis of the antimalarial artemisinin in Artemisia annua. Artemisinic aldehyde is reduced into dihydroartemisinic aldehyde by DBR2. Artemisinic aldehyde can also be oxidized by amorpha-4,11-diene 12-hydroxylase and/or aldehyde dehydrogenase 1 to artemisinic acid, a precursor of arteannuin B. In order to better understand the effects of DBR2 expression on the flow of artemisinic aldehyde into either artemisinin or arteannuin B, we determined the content of dihydroartemisinic aldehyde, artemisinin, artemisinic acid and arteannuin B content of A. annua varieties sorted into two chemotypes. The high artemisinin producers (HAPs), which includes the '2/39', 'Chongqing' and 'Anamed' varieties, produce more artemisinin than arteannuin B; the low artemisinin producers (LAPs), which include the 'Meise', 'Iran#8', 'Iran#14', 'Iran#24' and 'Iran#47' varieties, produce more arteannuin B than artemisinin. Quantitative PCR showed that the relative expression of DBR2 was significantly higher in the HAP varieties. We cloned and sequenced the promoter of the DBR2 gene from varieties of both the LAP and the HAP groups. There were deletions/insertions in the region just upstream of the ATG start codon in the LAP varities, which might be the reason for the different promoter activities of the HAP and LAP varieties. The relevance of promoter variation, DBR2 expression levels and artemisinin biosynthesis capabilities are discussed and a selection method for HAP varieties with a DNA marker is suggested. Furthermore, putative cis-acting regulatory elements differ between the HAP and LAP varieties.