SummaryOpium poppy (Papaver somniferum) is an important medicinal plant producing benzylisoquinoline alkaloids (BIA). MicroRNAs (miRNAs) are endogenous small RNAs (sRNAs) of approximately 21 nucleotides. They are noncoding, but regulate gene expression in eukaryotes. Although many studies have been conducted on the identification and functions of plant miRNA, scarce researches on miRNA regulation of alkaloid biosynthesis have been reported. In this study, a total of 316 conserved and 11 novel miRNAs were identified in opium poppy using second-generation sequencing and direct cloning. Tissue-specific regulation of miRNA expression was comparatively analysed by miRNA microarray assays. A total of 232 miRNAs were found to be differentially expressed among four tissues. Likewise, 1469 target transcripts were detected using in silico and experimental approaches. The Kyoto Encyclopedia of Genes and Genomes pathway analyses indicated that miRNA putatively regulates carbohydrate metabolism and genetic-information processing. Additionally, miRNA target transcripts were mostly involved in response to stress against various factors and secondary-metabolite biosynthesis processes. Target transcript identification analyses revealed that some of the miRNAs might be involved in BIA biosynthesis, such as pso-miR13, pso-miR2161 and psomiR408. Additionally, three putatively mature miRNA sequences were predicted to be targeting BIA-biosynthesis genes.

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Sternbergia fischeriana is an endangered geophyte and therefore in vitro micropropagation of this plant will have great importance for germplasm conservation and commercial production. Bulb scale and immature embryo explants of S. fischeriana were cultured on different nutrient media supplemented with various concentrations of plant growth regulators. Immature embryos produced higher number of bulblets than bulb scales. Large numbers of bulblets were regenerated (over 80 bulblets/explants) from immature embryos on Murashige and Skoog (MS) medium supplemented with 4 mg l )1 6-benzylaminopurine (BA) and 0.25 mg l )1 a-naphthaleneacetic (NAA) or 2 mg l )1 2,4-dichlorophenoxyacetic acid (2,4-D D ) after 14 months of culture initiation. Regenerated bulblets were kept at 5°C for 5 weeks and then transplanted to a potting mixture.

Seven genotypes of winter durum wheat {Triticum durum Desf,) were cultured to establish an efficient method of callus formation and plant regeneration from mature embryo culture, and to compare the responses of immature and mature embryo cultures. Immature embryos were aseptically dissected from seeds and placed, with the scutellum upwards, in dishes containing Murashige and Skoog's (MS) mineral salts and 2mg 2,4-dichlorophenoxyacetic acid (2,4-D) per litre, Calli and regenerated plants were maintained on 2,4-D-free medium. Mature embryos were moved slightly on the imbibed seeds. For callus formation, the seeds with moved embryos were placed, furrow downwards, in dishes containing 8 mg 2,4-D per litre. The developed calli and regenerated plants were maintained on the MS medium. Plants regenerated from both embryo cultures were vernalized and grown to maturity in soil. Variability was observed among the wheat genotypes tested for various culture responses in both explant cultures. Callus induction rate and regeneration capacity of callus were independent of each other. Mature embryos have a low frequency of callus induction but a high regeneration capacity. Considering availability, rapidity and reliability, this form of mature embryo culture can be used as an alternative method for immature embryo culture.

Cowpea is an important grain legume crop. The study reports an efficient in vitro multiplication and shoots regeneration protocol from preconditioned embryonic axes of the Turkish cowpea cultivar Akkiz. The embryonic axes were preconditioned with 10 mg/l BA on agar solidified MS medium for 5 days. Thereafter they were cultured on MS medium containing 0.25, 0.50, 0.75 and 1.00 mg/l BA with or without 0.10 mg/l NAA. Mean frequency (%) of shoot regeneration, number of shoots per explant and shoot length decreased with each increase in BA concentration used singly. However, a positive increase was recorded in all parameters in the presence of 0.10 mg/l NAA in the regeneration medium. A maximum mean number of 10.33 shoots per explant was recorded on an MS medium containing 1.00 mg/l BA -0.1 mg/l NAA. Regenerated shoots were rooted on an MS medium containing 0.50 mg/l IBA. Rooted plants were acclimatized at room temperature in soil mix contained in pots where they were subjected to an intermittent mistwater spray for 24 h that maintained 90% relative humidity during the first few days which was gradually reduced to 40% for 10 days. All plants flowered and set seeds in a greenhouse after 3 months.