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In order to understand the effects of ultraviolet-B (UV-B) radiation and salinity stress on visual quality and morphological parameters, one-year-old seedlings of rosemary (Rosmarinus officinalis L.) were grown under three UV-B levels (0, 4.32, 6.05 kJ m-2 d-1) and four salinity regimes (Control, 50, 100 and 150 mM NaCl) under greenhouse conditions. Results showed that, enhanced UV-B radiation increased visual quality, growth index, plant biomass, shoot/root ratio, number of branches and leaves, leaf fresh and dry weight, leaf area index, specific leaf area, leaf thickness. UV-B radiation had no significant effect on root length, however, shoot height, length of axillary shoots, length of inter node, leaf area, leaf length and width and concentration of chlorophyll b were negatively impacted by UV-B radiation. On the other hand, salinity caused significant decrease in plant biomass, root length, shoot height, shoot/root ratio, length of axillary shoots, length of inter node, number of branches and leaf area, leaf area index, leaf length and width, leaf fresh and dry weight, specific leaf area and concentration of Photosynthetic pigments. But, number leaves and leaf thickness significantly increased under NaCl salinity treatments. Visual quality and growth index were hardly affected by increasing salinity until 100 mM, but declined clearly at 150 mM NaCl salinity. Interaction between UV-B irradiation and salinity showed that, pre-treatment with UV-B irradiation alleviated harmful effects of NaCl and improved visual quality rosemary plants.

This chapter includes the methods for tissue and cell culture of saffron. Saffron is a triploid plant, and therefore vegetative reproduction is a common way for its propagation. In traditional culture, few cormlets are produced each year and thus the production rate is low. Tissue culture methods such as micropropagation, somatic embryogenesis, organogenesis, and in vitro cormogenesis are necessary for mass and free pathogen reproduction of this plant. All of these methods are discussed in the preliminary sections of this chapter. Tissue culture techniques can be very effective and useful methods for genetic modification and to produce new varieties of saffron. Protoplast culture to produce somatic hybrids and stigma-like structure culture to produce stigmas and their compounds that are used in saffron tissue culture and also related research is included at the end of this chapter.

Introduction: Anthurium is a popular genus of the Araceae (order Spathiflorae).The flower consists of a protruding spadix containing numerous florets, subtended by a brightly colored modified leaf, the spathe. Anthuriums are bisexual and protogynous.Anthuriumscherzerianum as the most important species ofAnthurium genus is a potted perennial plant. Due to having beautiful, attractive and long-life flowers, A. scherzerianum can be used for the production of pot and cut flowers. Tissue culture is suggested as the most commonly method in order to rapid propagation and removing disease in a short period of time. This method also recommended for Anthuriumbecause of problems in classical propagation method of this flower..The three basic propagation methods for Anthuriumare propagation by seed, traditional vegetative and tissue culture.Micropropagation of Anthurium is using forcommercial production. Materials and Methods: In this study, the effect of plant growth regulators and explants on indirect regeneration of A. scherzerianumdetermined in separate experiments. In the first experiment, callogenesis was done by leaf explants on MS medium containing growth regulators, BA in three concentrations (0.5, 1.25 and 2 mg/l) in combination with 2, 4-D (0.5, 1.25 and 2 mg/l) or NAA (0.5, 1.25 and 2 mg/l) and the combinations of TDZ (0.5, 1.25 and 2 mg/l) with 2, 4-D (0, 0.5 mg/l). In the second experiment, regeneration was done on MS medium containing 0.75 mg/l BA with 0.05 mg/l 2, 4-D and 0.1 mg/l NAA and also in combination with TDZ (0.75mg/l). For rooting, MS medium containing different concentrations of IBA and IAA (0, 0.2 and 1 mg/l) were used. Callus induction, regeneration and rooting experiments were done based on completely randomized design, with 12, 6 and 6 replications, respectively.Data from all the schemes used in this study were analyzed with SAS statistical software. The comparison of means using Duncan's multiple range test was evaluated at the 5% level. Results and Discussion: Analysis of variance showed that the effect of explant type and hormone combinations was significant on the percentage of callogenesis, callus volume and survival percentage. The interaction effect of explant type and combination of hormones was also significant on percentage of callogenesisand the volume of callus. Means comparisons showed that the highest callogenesis, viability and callus volume were achieved on MS medium containing 2 mg/l of BA and 0.5 mg/l of 2, 4-D. Petiole explants, also produced the highest percentage of callus (95%), survival rate (96%) and callus with dimensions of 6 mm2. Callus formation in leaf vein explants was higher than others. The effect of explant type and hormone combinations on regeneration, number of branches, number of leaves and leaf length was significant.The interaction of explant and hormone combinations on regeneration, number of branches, number of leaves and leaf length was also significant. Moreover, results of regeneration experiment indicated that the maximum number of shoots (6.9) and the maximum shoot length (5 cm), number of leaves (18) and the leaf length (2.8 mm) were achieved in 0.75 mg/l BA mg/l of and 0.05 mg/l 2, 4-D. In this study, petiole explants were also regenerated earlier than leaf explants.The effect of hormone combinations and concentrations was significant on rooting specially on the number of roots and root length.Furthermore, results of rooting experiment revealed that the highest rooting percentage (95%), the maximum number of roots (4.5 per plantlet) and the longest roots (3.5 cm) were produced in the medium containing 0.2 mg/l of IBA. Finally, the rooted plantlets were adapted (90%) in vivo condition by placing them on a mixture of cocopeat and perlite (2:1) substrate. Conclusion: In this study callugensis, regeneration and rooting of A.scherzerianum’s petiole and leaf explants were studied and different levels of plant growth regulators used for callugensis and regeneration. In this study petiole explants showed the highest callugenesis and regeneration. MS medium containing BA (2 mg/l) and 2, 4-D (0.5 mg/l), was the best for callugenesis. Also the highest percentage of regeneration was observed in medium containing BA (0.75 mg/l) and 2, 4-D (0.05 mg/l). Moreover low concentration (0.2 mg/l) of auxin has a better effect on rooting than high levels (1mg/l) so that the highest rooting percentage was produced in medium containing IBA (0.2 mg/l) and the lowest rooting percentage was produced in medium containing IAA (1 mg/l). Anthurium plantlets acclimized is cocopeat and perlite substrate (2: 1) with 90% acclimation.

There is an urgent need to develop novel drugs for osteoporosis which occurs due to estrogen deficiency. Phytoestrogens derived from medicinal plants would be the best alternative to chemical drugs with harmful side effects. The main purpose of the present study was to investigate the effect of ferutinin compared to 17&beta, estradiol (E2) on bone mineralization of zebrafish larvae. Regarding the lack of publications, the histology analysis was performed after exposure to E2 to find effective treatment on bone mineralization of developing zebrafish larvae. Then, the larvae were exposed to four concentrations of ferutinin at three time points to assess the mortality, the expression of some related genes and histology of the ceratohyal and hyomandibular of treated larvae. The RT-PCR result of the treatment groups demonstrated the similar expression pattern in the larvae which were exposed to 1.25 &mu, g/mL of ferutinin and 2 &micro, M of E2 at 2 dpf, which confirmed the result of histology analysis. In addition, RT-qPCR of high concentration of ferutinin and E2 demonstrated that bmp2a/b and esr1 were downregulated and upregulated when the larvae were exposed to 5 &mu, g/mL of ferutinin and 10 &micro, M of E2, respectively.

Variegation in flower color is commonly observed in many plant species and also occurs on petunia (Petunia hybrida) as an ornamental plant. Variegated plants are highly valuable in the floricultural market. To gain a global perspective on genes differentially expressed in variegated petunia flowers, we investigated the expression of chalcone synthase (chs) and chalcone isomerase (chi) as two essential genes in biosynthesis pathway of pigment production. Also, we measured the concentration level of total flavonoids, naringenin chalcone and naringenin to evaluate the probably relationship between the expression profile of chs and chi genes and the concentration of mentioned pigments. The results indicated that chalcone synthase and chalcone isomerase expression had different profile in different petal color of Petunia hybrida. Because red flower color in petunia is related to the synthesis of pelargonidin-based (orange to red) pigments, our results suggest that the low chalcone synthase and chalcone isomerase expression levels in white petals reduce dihydrokaempferol formation, thereby inhibiting pelargonidin production. In contrast, the high expression levels of these genes observed in red petals ensure sufficient anthocyanin yields to make flowers red.

Foot and Mouth Disease (FMD) is a very dangerous livestock disease which causes a serious loss in the production of milk and meat. Therefore, producing an effective recombinant subunit vaccine virus this disease is of great importance. Transient gene expression is a valuable tool to reach rapid and acceptable recombinant vaccine. An Agrobacterium-mediated transient gene expression assay was carried out in spinach (Spinacia oleracea) leaves for expression of a chimeric gene encoding a part of capsid protein of Foot and Mouth Disease virus called VP1. The plant leaves were transformed via agroinfiltration procedure. The presence of foreign gene and its expression in transformed plants were confirmed through polymerase chain reaction (PCR), real time PCR, protein Dot blot and ELISA. The results obtained in this examination showed quite a high level of gene expression in spinach leaves, showing that transient gene expression can be applied as an effective and time-saving procedure for the production of recombinant proteins. The procedures for transformation, detection of recombinant protein and its application for molecular experiments are described in the study.

Saffron (Crocus sativus L.) is the most valuable food additive in the world which little production and high price of it caused some adulterations such as plant and chemical material similar to saffron. There are several methods for detecting these fraudulent based on morphological and chemical tests but they are not effective in some cases. In this research a novel molecular method based on ITS-2 marker is introduced. A common forward primer based on 5.8s rDNA for all plant frauds such as safflower, corn stigmas, pomegranate, turmeric and capsicum slices was designed then specific reverse primers based on ITS-2 for any frauds have been designed for polymerase chain reaction. Related ITS-2 bands were amplified in any adulterations in saffron. Specific primer for camel and cow meet fibers was designed based on cytochrome b gene and could amplified the related bands. Multiplex PCR with all of these primers could amplify all of the bands related to any adulterations. Furthermore, using 20% polyacrylamide gel lead to good segregation of bands. This method can be used successfully for detection of low percentage (1%) of fraudulent in saffron. So this marker can be used efficiently for detection of these frauds in commercial saffron.

Expressed sequence tags simple sequence repeats (EST-SSRs) are important sources for investigation of genetic diversity and molecular marker development. Similar to genomic SSRs, the EST-SSRs are useful markers for many applications in genetics and plant breeding such as genetic diversity analysis, molecular mapping and cross-transferability across related species and genera. In spite of low polymorphism, these markers show variation in the expressed part of the genome. In this study, Medicago truncatula EST-SSRs were used for investigation of transferability between M. truncatula and some chickpea (Cicer arietinum L.) genotypes, also genetic diversity between used chickpea genotypes was determined. In this research, 650 M. truncatula ESTs were searched to find simple sequence repeats (SSRs). A total of 131 EST-SSRs were contained di-and trinucleotide motif SSRs. In this study, thirty pairs of primers were designed to amplify over 10 chickpea genotypes. Thirteen primer pairs (43%) generated reproducible bands in at least one chickpea genotype that eight bands (61.5%) were polymorphic in the chickpea genotypes. A total of 24 alleles were amplified with an average of 3 alleles per primer. The average of polymorphism information content (PIC) was 78.75% and transferability across M. truncatula and C. arietinum was 43.32%. The results indicate that the developed EST-SSR markers from M. truncatula as a model plant are valuable genetic markers for legume species such as chickpea. In addition to suitability of EST-SSR markers for genetic diversity analysis, their broad range of transferability also proved their potential for comparative genomics studies.

In this paper, the effects of a number of growth regulators as well as supplements to the Murashige and Skoog (MS) basal medium were evaluated on the regeneration of Alstroemeria rhizome explants. In the first experiment the effects of three cytokinins (BA, TDZ and 2IP each at 0.5, 1 and 2 mg/l) in combination with NAA (0.2 mg/l), followed by another PGR combination of 2IP (at 0.5, 1 and 2 mg/l) with NAA (0 and 0.2 mg/l), on regeneration of rhizome-derived explants, was investigated. Through the second experiment, the effects of a number of supplements, including glucose (30 g/l as the alternative for sucrose), casein hydrolysate (1 g/l), asparagine and glutamine, (each at 30 mg/l) added to MS medium, containing 1 mg/l BA and 0.2 mg/l NAA, was examined on rhizome explants� regeneration. Among the tested cytokinins, BA induced better regeneration of rhizome explants, resulting in a higher number of shoots compared to the other cytokinins. A medium supplemented with 1 mg/l BA and 0.2 mg/l NAA proved to be the most effective, with an average of 4.16 regenerated shoots per explant. In the second PGR combination, addition of NAA at 0.2 mg/l improved regeneration, compared to NAA-free treatments. In the second experiment, glucose substitution for sucrose improved regeneration with an average of 5.10 regenerated shoots per explant, compared to 4.16 shoots in sucrose-containing medium; whereas glutamine and asparagine (with 2.66 shoots) and casein hydrolysate (with 3.80 shoots) showed a negative influence on rhizome explants� regeneration.

Leaves, one year old stems and buds were used as explants. WPM, B5, MS, half and one-fourth strength MS with 2,4-?D, NAA, IBA, IAA and picloram 2, 5 and 10 mg/l were used for callus induction. Investigation into the seasonal effect on callus regeneration and explants browning was carried out in mid-April until September. Results showed that washing with sterile distilled water for one hour after surface sterilization was the best treatment for browning inhibition of explants than other treatments. Leaf explants have best callus formation. MS with 8 g/l agar had strong effect on callus formation. Culture for callus formation in May and June found to be higher than other months. 2,4-D, NAA and 10 mg/l picloram significantly enhanced callus formation. Leaf and bud explants sampling in April to May did not show any endophyte contamination but leaf explants in June until September, specially the one year old stems, showed intensive contamination. Key words: Seasonal effect, Browning inhibitor, Berberis vulgaris D. O. I. 10.3329/ptcb.v21i2.10239 Plant Tissue Cult. & Biotech. 21(2): 161-168, 2011 (December)

The synthesis of chitinases is one of the strategies evolved in higher plants to resist stresses. To investigate the role of chitinases in the interaction between chickpea (Cicer arietinum L.) and pathogens, the expression of two classes of chitinase, I and III, was monitored in infected and non-infected plants with different disease resistance (resistant MCC10 and sensitive MCC724 genotypes). The expression levels of these genes in chickpea were studied at different times after inoculation with Fusarium oxysporum f. sp. ciceri isolate 85. Semi-quantitative reverse transcription PCR was used to study the differential induction of the chitinase genes. Expression levels of chitinase I in non-infected and infected MCC724 plants were higher than those in MCC10. Since chitinase III may play an effective role in plant defense, its expression levels were quantified by reverse transcription PCR. Chitinase III was induced in MCC724 within 6 h and in MCC10 in 48 h after inoculation. However, the expression levels of chitinase III in MCC10 non-infected plants were significantly higher than in MCC724. The results showed that chitinase III can be effective to enhance resistance to F. oxysporum f. sp. ciceri isolate 85.

Saffron (Crocus sativus) is the most valuable and indigenous crop in Iran. The stigmas of flower are used as a popular natural flavouring, colouring and medicinal agent. However, the market suffers from frauds in this plant such as mixing with safflower petals due to high profit. Identification of these frauds with conventional and biochemical methods is difficult and low sensitive. Therefore, application of molecular markers such as random amplified polymorphic DNA (RAPD)/sequence characterized amplified regions (SCAR) is being considered as an alternative. In this study, DNA was extracted from dry stigmas of 5 Saffron accessions and dry petals of 7 safflower cultivars. RAPD reactions with ten 15-mer random primers resulted in two specific monomorphic bands (500 and 700 bp) for safflower, while they were absent in saffron accessions. PCR analysis with specific SCAR primers amplified two specific bands (414 and 589 bp) for safflowers in different combinations of saffron stigmas and safflower petals. This was the case with very low rates or 1% of safflower. Therefore, this method seems to be suitable for fraud identification of safflower petals in commercial saffron samples.

Saffron (Crocus sativus L.) is an indigenous and the most valuable and marginal plant in Iran. However, limited knowledge exists on its molecular biology. The importance of this plant is due to the color, flavor and medicinal properties of its red stigmas. Saffron stigmas contain a high amount of carotenoids such as crocetin and crocin. Two genes, bch and pds, have essential role in carotenoid production. In this study, the effect of four different irrigation regimes was evaluated on the expression of bch and pds genes. Semi-quantitative RTPCR showed no significant difference in the expression levels of genes of interest related to the internal standard (18S rRNA). Results of Real-Time PCR assays showed that the expression of bch and pds genes were affected by irrigation treatments as their expression decreased in irrigated plants in comparison to non irrigated ones, exept for one irrigation treatment (one irrigation in September) where the pds gene expression showed higher level. However, the expression profile of the genes was almost the same in all treatments. The comparison between results of two techniques indicated that the Real-Time PCR is more accurate for determination of the level of transcript in the Iranian saffron. It was interesting that by decreasing of irrigation, the expression level of these two genes increased indicating that abiotic stress and drought can affect on the gene expression relating to the saffron color.

This study surveyed a rapid, efficient and reproducible protocol for in vitro shoot regeneration and rooting by different explants and different concentration of BAP. Due to optimization of shoot regeneration, two media including of MS and modified MS (MS salts with double concentration of CaC21, and B5 vitamins), different explants such as decapitated embryo axes, epicotyls and cotyledonary nodes and different concentrations of BAP hormone (1, 1.5, 2, 2.5, 3 and 4 mg L(-1)) in two genotypes (Gachsaran and Flip. 92-12 L) were used. The results showed that modified MS is a suitable medium for in vitro shoot regeneration of lentil. High levels of BAP caused increasing of shoot regeneration in lentil genotypes. Three milligram per liter of BAP induced the highest level of shoot regeneration. In addition, decapitated embryo explants were the best explants for highest shoot regeneration (5.8) (p0.05). However, increasing of hormone concentration from 2 to 3 and 4 mg L(-1) caused decreasing in the number of shoots, so 2 mg L(-1) of BAP was best. For rooting, the in vitro-in vivo method of rooting was better than only in vitro method. The shoots regenerated in 2 mg L(-1) BAP had higher rooting percentage than the shoots were regenerated in 3 and 4 mg L(-1) BAP. These results indicate on the inhibitory effect of high concentration of BAP on root induction. But the genotype didn't have any significant effect on rooting percentage and length of roots.