The study of transcriptome sequencing for flower coloration in different anthesis stages of alpine ornamental herb (Meconopsis 'Lingholm').

Meconopsis (Papaveraceae) is an interesting alpine herb, mainly distributed in the mountainous area of southwest China and high altitude zone in Tibetan-Himalaya. Different Meconopsis species showed a flower color alteration in different anthesis stages, Meconopsis 'Lingholm' is one of the localized species whose petal color changes from purple to blue during the flowering process. In general, the blue color flower is a rare kind, and usually hard to cultivate artificially. The molecular mechanism of flower color formation and color alteration of alpine flowers were reported by many research workers. To find critical genes that regulate Meconopsis 'Lingholm' color alteration and the mechanism of environmental adaptation, the current study performed transcriptome sequencing by using Meconopsis 'Lingholm' petals from different anthesis stages. There were totally 91,615 unigenes obtained from 31.4 Gb sequencing data, and differentially expressed genes between two consecutive flowering stages were obtained. Bioinformatics studies showed genes regulating petal color alteration were activated. Moreover, the functional analysis showed that Meconopsis 'Lingholm' showed a stress response to mechanical damage, non-biological stimulation and water deficiency in the bud stage, as well as showed a stress response to the cold from cracking stage to blooming stage. Furthermore, RNA-Seq results were verified using nine randomly selected genes by qPCR, which showed same expression trend with sequencing results. During this study, 20 candidate genes identified for further studies, which included five petal color related genes and 15 environmental response genes.

Hairy root induction and benzylisoquinoline alkaloid production in Macleaya cordata.

Sanguinarine is currently widely used to replace antibiotic growth promoters in animal feeding and has demonstrated useful anticancer activity. Currently, the main source of sanguinarine is from an important medicinal plant, Macleaya cordata. To obtain a new source of sanguinarine production, we established hairy root cultures of M. cordata by co-cultivating leaf and stem explants with Agrobacterium rhizogenes. Except the co-cultivation medium, all growth media contained 200 mg/L timentin to eliminate A. rhizogenes. Through comparing the metabolic profiles and gene expression of hairy roots and wild-type roots sampled at five time points, we found that the sanguinarine and dihydrosanguinarine contents of hairy roots were far higher than those of wild-type roots, and we revealed the molecular mechanism that causes these metabolites to increase. Consequently, this study demonstrated that the hairy root system has further potential for bioengineering and sustainable production of sanguinarine on a commercial scale. To the best of our knowledge, this is the first efficient protocol reported for the establishment of hairy root cultures in M. cordata using A. rhizogenes.

Several newly discovered Mo-enriched plants with a focus on Macleaya cordata.

Phytoremediation as an alternative strategy has been a widespread attention. The screening of enriched plants and hyperaccumulators is the key of the strategy. So this study examined the status of heavy metal pollution in molybdenum (Mo) mine soils, metal accumulation in plants growing on mine, and their tolerance strategies. The analysis of 14 soils and 27 plant samples in mining area showed that Mo, zinc (Zn), and cadmium (Cd) concentrations exceeded soil safety standards and their levels varied in 27 plant samples. Mo was the heavy pollution with an average total content of 256.1 mg/kg in soils. As Mo-enriched plants, Mo concentrations of Macleaya cordata (Willd.) R. Br. and Morus australis Poir. were 704.4 and 772.4 mg/kg, respectively. M. cordata was selected as the research material, due to its high biomass. Molybdenum significantly decreased the biomass and photosynthesis of M. cordata at high concentration (> 200 µmol/L), but its biomass and photosynthesis reached the maximum after 50 µmol/L Mo treatment, respectively. Analysis of the subcellular distribution and chemical speciation showed that Mo was distributed a certain way in the extracts and that this suggested that it may be present in cell wall and soluble fraction of roots (51.9-63.9%; 26.1-44.7%) or shoots (30.0-44.4%; 47.3-56.0%) and complexed to organic acid, pectate, oxalate, and protein. This might be responsible for the adaptation of M. cordata to Mo stress. Therefore, M. cordata could serve as a potential plant to utilize for the phytoremediation of Mo-contaminated soil.

Plant germination and production of callus from the yellow hornpoppy (Glaucium flavum): the first stage of micropropagation.

The yellow hornpoppy, Glaucium flavum Cr. (Fam. Papaveraceae) is a perennial herb, distributed in the Mediterranean region, including Egypt. The plant contains many benzyl isoquinoline alkaloids from the aporphine type such as glaucine, isoboldine, 1-chelidonine, 1-norchelidonine and 3-O-methylarterenol, making it to display various medicinal activities including antitussive, anticancer, antioxidant, antimicrobial, antiviral, hypoglycemic, analgesic, antipyretic, bronchodilator and anti-inflammatory effects. The plant is now rare and endangered in the Egyptian flora due to urban sprawl. The present study looks into Glaucium flavum seeds' in vitro germination as well as the ability of the explants taken from the growing seedlings to form stable callus lines in order to enable micropropagation as a way to save the rare plant. The study also scans the production of different medicinally valuable alkaloids, particularly glaucine, in produced callus.

Asymmetric morphogenetic cues along the transverse plane: shift from disymmetry to zygomorphy in the flower of Fumarioideae.

PREMISE OF THE STUDY: Zygomorphy has evolved multiple times in angiosperms. Near-actinomorphy is the ancestral state in the early diverging eudicot family Papaveraceae. Zygomorphy evolved once in the subfamily Fumarioideae from a disymmetric state. Unusual within angiosperms, zygomorphy takes place along the transverse plane of the flower. METHODS: We investigated floral development to understand the developmental bases of the evolution of floral symmetry in Papaveraceae. We then assessed the expression of candidate genes for the key developmental events responsible for the shift from disymmetry to transverse zygomorphy, namely CrabsClaw for nectary formation (PapCRC), ShootMeristemless (PapSTL) for spur formation, and Cycloidea (PapCYL) for growth control. KEY RESULTS: We found that an early disymmetric groundplan is common to all species studied, and that actinomorphy was acquired after sepal initiation in Papaveroideae. The shift from disymmetry to zygomorphy in Fumarioideae was associated with early asymmetric growth of stamen filaments, followed by asymmetric development of nectary outgrowth and spur along the transverse plane. Patterns of PapSTL expression could not be clearly related to spur formation. PapCRC and PapCYL genes were expressed in the nectary outgrowths, with a pattern of expression correlated with asymmetric nectary development in the zygomorphic species. Additionally, PapCYL genes were found asymmetrically expressed along the transverse plane in the basal region of outer petals in the zygomorphic species. CONCLUSION: Genes of PapCRC and PapCYL families could be direct or indirect targets of the initial transversally asymmetric cue responsible for the shift from disymmetry to zygomorphy in Fumarioideae.

Diversity and evolution of CYCLOIDEA-like TCP genes in relation to flower development in Papaveraceae.

Monosymmetry evolved several times independently during flower evolution. In snapdragon (Antirrhinum majus), a key gene for monosymmetry is CYCLOIDEA (CYC), which belongs to the class II TCP gene family encoding transcriptional activators. We address the questions of the evolutionary history of this gene family and of possible recruitment of genes homologous to CYC in floral development and symmetry in the Papaveraceae. Two to three members of the class II TCP family were found in each species analyzed, two of which were CYC-like genes, on the basis of the presence of both the TCP and R conserved domains. The duplication that gave rise to these two paralogous lineages (named PAPACYL1 and PAPACYL2) probably predates the divergence of the two main clades within the Papaveraceae. Phylogenetic relationships among angiosperm class II TCP genes indicated that (1) PAPACYL genes were closest to Arabidopsis (Arabidopsis thaliana) AtTCP18, and a duplication at the base of the core eudicot would have given rise to two supplementary CYC-like lineages; and (2) at least three class II TCP genes were present in the ancestor of monocots and eudicots. Semiquantitative reverse transcription-polymerase chain reaction and in situ hybridization approaches in three species with different floral symmetry indicated that both PAPACYL paralogs were expressed during floral development. A pattern common to all three species was observed at organ junctions in inflorescences and flowers. Expression in the outer petals was specifically observed in the two species with nonactinomorphic flowers. Hypotheses concerning the ancestral pattern of expression and function of CYC-like genes and their possible role in floral development of Papaveraceae species leading to bisymmetric buds are discussed.

EST database for early flower development in California poppy (Eschscholzia californica Cham., Papaveraceae) tags over 6,000 genes from a basal eudicot.

The Floral Genome Project (FGP) selected California poppy (Eschscholzia californica Cham. ssp. Californica) to help identify new florally-expressed genes related to floral diversity in basal eudicots. A large, non-normalized cDNA library was constructed from premeiotic and meiotic floral buds and sequenced to generate a database of 9,079 high quality Expressed Sequence Tags (ESTs). These sequences clustered into 5,713 unigenes, including 1,414 contigs and 4,299 singletons. Homologs of genes regulating many aspects of flower development were identified, including those for organ identity and development, cell and tissue differentiation, cell cycle control, and secondary metabolism. Over 5% of the transcriptome consisted of homologs to known floral gene families. Most are the first representatives of their respective gene families in basal eudicots and their conservation suggests they are important for floral development and/or function. App. 10% of the transcripts encoded transcription factors and other regulatory genes, including nine genes from the seven major lineages of the important MADS-box family of developmental regulators. Homologs of alkaloid pathway genes were also recovered, providing opportunities to explore adaptive evolution in secondary products. Furthermore, comparison of the poppy ESTs with the Arabidopsis genome provided support for putative Arabidopsis genes that previously lacked annotation. Finally, over 1,800 unique sequences had no observable homology in the public databases. The California poppy EST database and library will help bridge our understanding of flower initiation and development among higher eudicot and monocot model plants and provide new opportunities for comparative analysis of gene families across angiosperm species.

Expression patterns of STM-like KNOX and Histone H4 genes in shoot development of the dissected-leaved basal eudicot plants Chelidonium majus and Eschscholzia californica (Papaveraceae).

Knotted-like homeobox (KNOX) genes encode important regulators of shoot development in flowering plants. In Arabidopsis, class I KNOX genes are part of a regulatory system that contributes to indeterminacy of shoot development, delimitation of leaf primordia and internode development. In other species, class I KNOX genes have also been recruited in the control of marginal blastozone fractionation during dissected leaf development. Here we report the isolation of class I KNOX genes from two species of the basal eudicot family Papaveraceae, Chelidonium majus and Eschscholzia californica. Sequence comparisons and expression patterns indicate that these genes are orthologs of SHOOTMERISTEMLESS (STM), a class I KNOX gene from Arabidopsis. Both genes are expressed in the center of vegetative and floral shoot apical meristems (SAM), but downregulated at leaf or floral organ initiating sites. While Eschscholzia californica STM (EcSTM) is again upregulated during acropetal pinna formation, in situ hybridization could not detect Chelidonium majus STM (CmSTM) transcripts at any stage of basipetal leaf development, indicating divergent evolution of STM gene function in leaves within Papaveraceae. Immunolocalization of KNOX proteins indicate that other gene family members may control leaf dissection in both species. The contrasting direction of pinna initiation in the two species was also investigated using Histone H4 expression. Leaves at early stages of development did not reveal notable differences in cell division activity of the elongating leaf axis, suggesting that differential meristematic growth may not play a role in determining the observed dissection patterns.

[Concentration of four alkaloids in the aerial parts of Eomecon chionantha from different mounth in year].

The concentration of four alkaloids (sanguinarine, chelerythrine, protopine and alpha-allocryptopine) in the aerial parts of snowpopy (Eomecon chionantha) collected from April to August in 1996 were determined by capillary electrophoretic method. The results show that the change of contents of alkaloids according to months is similar to that of the underground parts of snowpopy, i.e. the contents of sanguinarine and chelerythrine wrer lower in June than other months; the contents of chelerythrine were higher than that of sanguinarine and alpha-allocryptopine were higher than protopine.