Different Stages Of Germination Research Articles

Identification and expression analysis of MADS-box gene family in Docynia delavayi (Franch.) Schneid.

MADS-box gene family is a significant transcription factor family that plays a crucial role in regulating plant growth, development, signal transduction, and other processes. In order to study the characteristics of MADS-box gene family in Docynia delavayi (Franch.) Schneid. and its expression during different stages of seed germination, this study used seedlings at different stages of germination as materials and screened MADS-box transcription factors from the transcriptome database of D. delavayi using bioinformatics methods based on transcriptome sequencing. The physical and chemical properties, protein conservative motifs, phylogenetic evolution, and expression patterns of the MADS-box transcription factors were analyzed. Quantitative real-time PCR (qRT-PCR) was used to verify the expression of MADS-box gene family members during different stages of seed germination in D. delavayi. The results showed that 81 genes of MADS-box gene family were identified from the transcriptome data of D. delavayi, with the molecular weight distribution ranged of 6 211.34-173 512.77 Da and the theoretical isoelectric point ranged from 5.21 to 10.97. Phylogenetic analysis showed that the 81 genes could be divided into 15 subgroups, among which DdMADS27, DdMADS42, DdMADS45, DdMADS46, DdMADS53, DdMADS61, DdMADS76, DdMADS77 and DdMADS79 might be involved in the regulation of ovule development in D. delavayi. The combination of the transcriptome data and the qRT-PCR analysis results of D. delavayi seeds indicated that DdMADS25 and DdMADS42 might be involved in the regulation of seed development, and that DdMADS37 and DdMADS38 might have negative regulation effects on seed dormancy. Previous studies have reported that the MIKC* subgroup is mainly involved in regulating flower organ development. For the first time, we found that the transcription factors of the MIKC* subgroup exhibited a high expression level at the early stage of seed germination, so we speculated that the MIKC* subgroup played a regulatory role in the process of seed germination. To verify the accuracy of this speculation, we selected DdMADS60 and DdMADS75 from the MIKC* subgroup for qRT-PCR experiments, and the experimental results were consistent with the expression trend of transcriptome sequencing. This study provides a reference for further research on the biological function of D. delavayi MADS-box gene family from the perspective of molecular evolution.

Spermosphere Bacteria Promote Ormosia henryi Seed Germination by Activating Metabolic Pathways

Spermosphere microorganisms are affected by seed exudates (carbon deposits) and interact with each other around the germinating seeds. The interactions in this region will have an important impact on seed germination, plant growth, and development. Ormosia henryi Prain is a rare and endangered tree species with hardseed in China that generally shows low levels of natural regeneration. However, good levels of natural regeneration were observed under a few mother trees in Guanling County (Guizhou Province, China). Based on this phenomenon: we raise the question: Is the seed germination of Ormosia henryi in high-level natural regeneration areas driven by the spermosphere microbe and what is the driving mechanism? The purpose of this study was to explore the promoting role of spermosphere bacteria in different stages of germination, so as to provide a theoretical basis for screening strong colonization ability spermosphere functional bacteria. Thus, in this study, we designed a germination experiment simulating the natural soil (sterilized and unsterilized) of Guanling County. We used 16S rRNA gene sequencing and widely targeted multiple reaction monitoring (MRM) metabolomic analysis to analyze bacterial microbial diversity and the metabolic profiles of seed exudates at the four stages of seed germination: imbibition, radicle protrusion, radicle elongation, and cotyledon protrusion. The results indicated that some spermosphere bacteria: taxa of Bacillus, Acidobacteriales, Nitrospira, and Ellin6067, and other functional bacteria may play important roles in promoting seed germination depending largely on their functional characteristics and abundance. The critical period for spermosphere bacteria to promote seed germination was at the radicle protrusion stage in the unsterilized soil group, levels of metabolites such as sugars and amino acids of seed exudates were increased in this stage, more respiratory pathways were activated and seed respiratory metabolism was enhanced. Spermosphere bacteria of wild Ormosia henryi in the Guanling area appeared to promote the germination of O. henryi seeds, mainly through activating galactose metabolism and the lysine degradation pathway to provide energy for germination.

Solid-state NMR molecular snapshots of Aspergillus fumigatus cell wall architecture during a conidial morphotype transition

While establishing an invasive infection, the dormant conidia of Aspergillus fumigatus transit through swollen and germinating stages, to form hyphae. During this morphotype transition, the conidial cell wall undergoes dynamic remodeling, which poses challenges to the host immune system and antifungal drugs. However, such cell wall reorganization during conidial germination has not been studied so far. Here, we explored the molecular rearrangement of Aspergillus fumigatus cell wall polysaccharides during different stages of germination. We took advantage of magic-angle spinning NMR to investigate the cell wall polysaccharides, without employing any destructive method for sample preparation. The breaking of dormancy was associated with a significant change in the molar ratio between the major polysaccharides β-1,3-glucan and α-1,3-glucan, while chitin remained equally abundant. The use of various polarization transfers allowed the detection of rigid and mobile polysaccharides; the appearance of mobile galactosaminogalactan was a molecular hallmark of germinating conidia. We also report for the first time highly abundant triglyceride lipids in the mobile matrix of conidial cell walls. Water to polysaccharides polarization transfers revealed an increased surface exposure of glucans during germination, while chitin remained embedded deeper in the cell wall, suggesting a molecular compensation mechanism to keep the cell wall rigidity. We complement the NMR analysis with confocal and atomic force microscopies to explore the role of melanin and RodA hydrophobin on the dormant conidial surface. Exemplified here using Aspergillus fumigatus as a model, our approach provides a powerful tool to decipher the molecular remodeling of fungal cell walls during their morphotype switching.

De novo transcriptome assembly of the cotyledon of Camellia oleifera for discovery of genes regulating seed germination

BackgroundCamellia oleifera (C.oleifera) is one of the most important wood oil species in the world. C.oleifera was propagated by nurse seedling grafting. Since the morphology of rootstocks has a significant impact on grafting efficiency and seedling quality, it is necessary to understand the molecular mechanism of morphogenesis for cultivating high-quality and controllable rootstocks. However, the genomic resource for this species is relatively limited, which hinders us from fully understanding the molecular mechanisms of seed germination in C.oleifera.ResultsIn this paper, using transcriptome sequencing, we measured the gene expression in the C.oleifera cotyledon in different stages of development and the global gene expression profiles. Approximately 45.4 gigabases (GB) of paired-end clean reads were assembled into 113,582 unigenes with an average length of 396 bp. Six public protein databases annotate 61.5% (68,217) of unigenes. We identified 11,391 differentially expressed genes (DEGs) throughout different stages of germination. Enrichment analysis revealed that DEGs were mainly involved in hormone signal transduction and starch sucrose metabolism pathways. The gravitropism regulator UNE10, the meristem regulators STM, KNAT1, PLT2, and root-specific transcription factor WOX11 all have higher gene expression levels in the CAM2 stage (seed soaking), which indicates that the cotyledon-regulated program for germination had initiated when the seeds were imbibition. Our data showed differentially reprogrammed to multiple hormone-related genes in cotyledons during C.oleifera seed germination.ConclusionCotyledons play vital roles, both as the main nutrient provider and as one primary instructor for seed germination and seedling growth. Together, our study will significantly enrich the genomic resources of Camellia and help us understand the molecular mechanisms of the development in the seed germination and seedling growth of C.oleifera. It is helpful to culture standard and superior quality rootstock for C.oleifera breeding.

Microscopy Techniques for Interpreting Fungal Colonization in Mycoheterotrophic Plants Tissues and Symbiotic Germination of Seeds.

Structural botany is an indispensable perspective to fully understand the ecology, physiology, development, and evolution of plants. When researching mycoheterotrophic plants (i.e., plants that obtain carbon from fungi), remarkable aspects of their structural adaptations, the patterns of tissue colonization by fungi, and the morphoanatomy of subterranean organs can enlighten their developmental strategies and their relationships with hyphae, the source of nutrients. Another important role of symbiotic fungi is related to the germination of orchid seeds; all Orchidaceae species are mycoheterotrophic during germination and seedling stage (initial mycoheterotrophy), even the ones that photosynthesize in adult stages. Due to the lack of nutritional reserves in orchid seeds, fungal symbionts are essential to provide substrates and enable germination. Analyzing germination stages by structural perspectives can also answer important questions regarding the fungi interaction with the seeds. Different imaging techniques can be applied to unveil fungi endophytes in plant tissues, as are proposed in this article. Freehand and thin sections of plant organs can be stained and then observed using light microscopy. A fluorochrome conjugated to wheat germ agglutinin can be applied to the fungi and co-incubated with Calcofluor White to highlight plant cell walls in confocal microscopy. In addition, the methodologies of scanning and transmission electron microscopy are detailed for mycoheterotrophic orchids, and the possibilities of applying such protocols in related plants is explored. Symbiotic germination of orchid seeds (i.e., in the presence of mycorrhizal fungi) is described in the protocol in detail, along with possibilities of preparing the structures obtained from different stages of germination for analyses with light, confocal, and electron microscopy.

Changes in antioxidant system and sucrose metabolism in maize varieties exposed to Cd

Different maize varieties respond differentially to cadmium (Cd) stress. However, the physiological mechanisms that determine the response are not well defined. Antioxidant systems and sucrose metabolism help plants to cope with abiotic stresses, including Cd stress. The relationship of these two systems in the response to Cd stress is unclear. Seed is sensitive to Cd stress during germination. In this study, we investigated changes in the antioxidant system, sucrose metabolism, and abscisic acid and gibberellin concentrations in two maize varieties with low (FY9) or high (SY33) sensitivities to Cd under exposure to CdCl2 (20 mg L−1) at different stages of germination (3, 6, and 9 days).The seed germination and seedling growth were inhibited under Cd stress. The superoxide, malondialdehyde, and proline concentrations, and the superoxide dismutase, peroxidase, catalase, and lipoxygenase activities increased compared with those of the control (CK; without Cd). The expression levels of three genes (ZmOPR2, ZmOPR5, and ZmPP2C6) responsive to oxidative stress increased differentially in the two varieties under Cd stress. The activity of the antioxidant system and the transcript levels of oxidative stress–responsive genes were higher in the Cd-tolerant variety, FY9, than in the sensitive variety, SY33. Sucrose metabolism was increased under Cd stress compared with that of the CK and was more active in the Cd-sensitive variety, SY33. These results suggest that the antioxidant system is the first response to Cd stress in maize, and that sucrose metabolism is cooperative and complementary under exposure to Cd.

Biochemical and transcriptomic analyses of the symbiotic interaction between Cremastra appendiculata and the mycorrhizal fungus Coprinellus disseminatus

BackgroundCremastra appendiculata is a rare terrestrial orchid with a high market value as an ornamental and medicinal plant. However, the species depends entirely on fungi for seed germination under natural conditions. In a previous study, we have successfully isolated and identified the mycorrhizal fungus Coprinellus disseminatus which was able to induce the germination of C. appendiculata seeds. We then speculated that C. disseminatus may do so by breaking the testa imposed dormancy of the seeds. In this study, biochemical and transcriptomic analyses were used to characterize the germination of C. appendiculata seeds, collected at different stages of germination, as affected by C. disseminatus.ResultsThe lignocellulose in the seeds coat of C. appendiculata was degraded by the mycorrhizal fungus resulting in facilitated absorption of water. The rate of decline in lignin content was 67 and 73% at 6 and 12 days after sowing, respectively. The water content increased from 13 to 90% during symbiosis. A total of 15,382 genes showing significantly different levels of expression (log2 FPKM≥2.0, Qvalue≤0.05) were successfully identified among all libraries, where the highest number of DEGs was shared between 6 days versus 0 day after symbiotic germination. Gene annotation results suggested that 15 key genes related water-status, such as DHN gene family and Xero 1 were down-regulated. The genes zeaxanthin epoxidase ZEP, 9-cis-epoxycarotenoid dioxygenase NCED3 and β-carotene hydroxylase involved in the biosynthesis of abscisic acid (ABA) were significantly down-regulated in 6 days as compared to 0 day after symbiotic germination.ConclusionsThis work demonstrates that mycorrhizal fungus C. disseminatus can stimulate C. appendiculata seeds germination through a mechanism of breaking the testa imposed dormancy and inducing water absorption of the embryo.

Involvement of the MetO/Msr System in Two Acer Species That Display Contrasting Characteristics during Germination.

The levels of methionine sulfoxide (MetO) and the abundances of methionine sulfoxide reductases (Msrs) were reported as important for the desiccation tolerance of Acer seeds. To determine whether the MetO/Msrs system is related to reactive oxygen species (ROS) and involved in the regulation of germination in orthodox and recalcitrant seeds, Norway maple and sycamore were investigated. Changes in water content, MetO content, the abundance of MsrB1 and MsrB2 in relation to ROS content and the activity of reductases depending on nicotinamide adenine dinucleotides were monitored. Acer seeds differed in germination speed—substantially higher in sycamore—hydration dynamics, levels of hydrogen peroxide, superoxide anion radicals (O2•−) and hydroxyl radicals (•OH), which exhibited peaks at different stages of germination. The MetO level dynamically changed, particularly in sycamore embryonic axes, where it was positively correlated with the levels of O2•− and the abundance of MsrB1 and negatively with the levels of •OH and the abundance of MsrB2. The MsrB2 abundance increased upon sycamore germination; in contrast, it markedly decreased in Norway maple. We propose that the ROS–MetO–Msr redox system, allowing balanced Met redox homeostasis, participates in the germination process in sycamore, which is characterized by a much higher speed compared to Norway maple.

The oleosin gene family in coffee: Analysis of oleosomes, oleosins and lipid-related gene expression during germination of Coffea arabica seeds

Little information is available about lipid metabolism in Coffea arabica L. seeds, which have an oil content between 11 and 16% w/w. This is surprising given the involvement of lipids in the physical, chemical, and physiological properties of the coffee beverage. To shed light on this subject, freshly harvested C. arabica seeds were sampled at different stages of germination. Oleosomes (the organelles responsible for lipid storage) were extracted and their structure was observed by differential interference-contrast microscopy. Triacylglycerol content of oleosomes was analyzed by thin-layer chromatography, and the protein fraction was evaluated by gel electrophoresis. A review of available genomic data on oleosin and lipase genes was performed, and expression of genes related to lipid metabolism was measured by Real-Time PCR.The results drew a coherent picture of coffee seed germination, revealing that oleosins have the highest concentration after 14 days of germination, followed by a marked decrease. This was backed up by molecular observations, showing how oleosin genes were downregulated after 6–9 days of germination, while genes related to lipid degradation were upregulated from day 9 onwards. Morphological evidence supported these findings, as after 14 days embryo growth became significant and cotyledons started to invade the endosperm and metabolize seed reserves. The present research will be an important first step in better understanding the biology of oleosomes in Coffea arabica L. and in elucidating the role of lipid metabolism in green coffee quality.

Análise biométrica multidimensional com tratamentos pré-germinativos em sementes e caracterização morfológica de plântulas de Mimosa bimucronata (De Candolle) Otto Kuntze

Mimosa bimucronata is a plant species that can be used in reforestation programs, and initial studies for the production of seedlings are needed. Due to the importance of this plant species and the scarcity of information on seed germination, this research was carried out with the objective of evaluate i) the biometric characteristics of the seeds by multivariate analysis, ii) the effect of seed pretreatment on germination, and iii) to characterize morphologically the different stages of germination and seedling. The biometry and seed mass and eight pre-germination treatments were analyzed: control; sulfuric acid (98%) for two; five and 10 min; hot water at 70 to 80 °C for 2 min; mechanical scarification and distilled water for 24 h. The different stages of seed development were evaluated until seedling formation. The seeds were exposed to two environments - laboratory and greenhouse. The percentage of germination and emergence were evaluated; germination and emergence speed index; and average time of germination and emergence. According to the multivariate analysis, the seeds were classified into three groups, with classes II and III being the most suitable for seedling production. Germination with sulfuric acid and sandpaper were the most indicated. The species presents 10 stages of development.

Early detection of germinated wheat grains using terahertz image and chemometrics.

In this paper, we propose a feasible tool that uses a terahertz (THz) imaging system for identifying wheat grains at different stages of germination. The THz spectra of the main changed components of wheat grains, maltose and starch, which were obtained by THz time spectroscopy, were distinctly different. Used for original data compression and feature extraction, principal component analysis (PCA) revealed the changes that occurred in the inner chemical structure during germination. Two thresholds, one indicating the start of the release of α-amylase and the second when it reaches the steady state, were obtained through the first five score images. Thus, the first five PCs were input for the partial least-squares regression (PLSR), least-squares support vector machine (LS-SVM), and back-propagation neural network (BPNN) models, which were used to classify seven different germination times between 0 and 48 h, with a prediction accuracy of 92.85%, 93.57%, and 90.71%, respectively. The experimental results indicated that the combination of THz imaging technology and chemometrics could be a new effective way to discriminate wheat grains at the early germination stage of approximately 6 h.

Application of Sulfentrazone in Stages of Germination of IAC 90 Cassava Cuttings in Clay Soils and Sandy

Weeds interfere dramatically in the productive potential of cassava; however, information regarding herbicides that are selective to crops is still scarce. Thus, the aim in this study was to assess the initial growth of IAC 90 cassava plants after the application of sulfentrazone at different stages of germination of cassava in clayey and sandy soils. Three experiments were simultaneously deployed: the first experiment consisted in the application of sulfentrazone in the non-germinated stage of cassava cuttings; the second one in the stage of germinated cassavas cuttings (0.9 cm shoots); and the third one in applications in the stage of cassava cuttings with buds emerging (6.5 cm shoots and emerging from the soil). For each experiment the experimental design in randomized blocks was used in the 2 x 5 factorial arrangement with four replications. The factors were composed of two soils (sandy and clayey) and five doses of sulfentrazone (0, 250, 500, 750 and 1,000 g ha-1). It was found that depending on the herbicide dose, development stage of the buds of cassava cuttings and the type of soil, damage can occur in the initial development of the IAC 90 cassava plants. The greatest potential of sulfentrazone selectivity has occurred in applications in the non-germinated cassava cuttings stage and in doses lower than 500 g ha-1 in the clayey soil.

Physiological and proteomic analyses on artificially aged Brassica napus seed.

Plant seeds lose their viability when they are exposed to long term storage or controlled deterioration treatments, by a process known as seed aging. Based on previous studies, artificially aging treatments have been developed to accelerate the process of seed aging in order to understand its underlying mechanisms. In this study, we used Brassica napus seeds to investigate the mechanisms of aging initiation. B. napus seeds were exposed to artificially aging treatment (40°C and 90% relative humidity) and their physio-biochemical characteristics were analyzed. Although the treatment delayed germination, it did not increase the concentration of cellular reactive oxygen species (ROS). Comparative proteomic analysis was conducted among the control and treated seeds at different stages of germination. The proteins responded to the treatment were mainly involved in metabolism, protein modification and destination, stress response, development, and miscellaneous enzymes. Except for peroxiredoxin, no changes were observed in the accumulation of other antioxidant enzymes in the artificially aged seeds. Increased content of abscisic acid (ABA) was observed in the artificially treated seeds which might be involved in the inhibition of germination. Taken together, our results highlight the involvement of ABA in the initiation of seed aging in addition to the ROS which was previously reported to mediate the seed aging process.

In-depth proteomic analysis of rice embryo reveals its important roles in seed germination.

Seed germination is a complex physiological process that allows the seed embryo to grow and develop into a photosynthetic organism. The two major constituents of rice seed include the embryo and endosperm, with embryo being of much significance despite its small size. In this study, we conducted a systematic proteomic analysis of the embryos dissected from rice seed at different stages of germination through a combination of gel-based and gel-free strategies. In total, 343 differentially expressed proteins were identified. Among them, 191 were decreased and 152 were increased in terms of expression levels. All these proteins could be sorted into 11 functional groups based on MapMan analysis. Some starch biosynthesis-related enzymes such as starch branching enzyme, granule-bound starch synthase 1 and starch synthase increased during the early stage of germination and then decreased at the late stage, which was similar to the expressional patterns of glycolysis-related enzymes. However, tricarboxylic acid cycle-related enzymes only increased at the later stage. It was also found that sucrose might be an important intermediate for the biosynthesis of starch in embryos. Furthermore, gel-based proteomic analysis of the dissected endosperm showed that the biological processes in the endosperm were heavily regulated by the embryo. This study could provide some new insights into the distinct roles of the embryo and endosperm in rice seed germination.

The window of desiccation tolerance shown by early-stage germinating seedlings remains open in the resurrection plant, Xerophyta viscosa.

Resurrection plants are renowned for their vegetative desiccation tolerance (DT). While DT in vegetative tissues is rare in angiosperms, it is ubiquitous in mature orthodox seeds. During germination, seedlings gradually lose DT until they pass a point of no return, after which they can no longer survive dehydration. Here we investigate whether seedlings of the resurrection plant Xerophyta viscosa ever lose the capacity to establish DT. Seedlings from different stages of germination were dehydrated for 48 hours and assessed for their ability to recover upon rehydration. While a transient decline in the ability of X. viscosa seedlings to survive dehydration was observed, at no point during germination was the ability to re-establish DT completely lost in all seedlings. Pre-treatment of seedlings with PEG or sucrose reduced this transient decline, and improved the survival rate at all stages of germination. Additionally, we observed that the trait of poikilochlorophylly (or loss of chlorophyll) observed in adult X. viscosa leaves can be induced throughout seedling development. These results suggest that the window of DT seen in germinating orthodox seeds remains open in X. viscosa seedlings and that vegetative DT in Xerophyta species may have evolved from the ability to retain this program through to adulthood.

Β-Amylase from Starchless Seeds of Trigonella Foenum-Graecum and Its Localization in Germinating Seeds

Fenugreek (Trigonella foenum-graecum) seeds do not contain starch as carbohydrate reserve. Synthesis of starch is initiated after germination. A β-amylase from ungerminated fenugreek seeds was purified to apparent electrophoretic homogeneity. The enzyme was purified 210 fold with specific activity of 732.59 units/mg. Mr of the denatured enzyme as determined from SDS-PAGE was 58 kD while that of native enzyme calculated from size exclusion chromatography was 56 kD. Furthermore, its identity was confirmed to be β-amylase from MALDI-TOF analysis. The optimum pH and temperature was found to be 5.0 and 50°C, respectively. Starch was hydrolyzed at highest rate and enzyme showed a Km of 1.58 mg/mL with it. Antibodies against purified Fenugreek β-amylase were generated in rabbits. These antibodies were used for localization of enzyme in the cotyledon during different stages of germination using fluorescence and confocal microscopy. Fenugreek β-amylase was found to be the major starch degrading enzyme depending on the high amount of enzyme present as compared to α-amylase and also its localization at the periphery of amyloplasts. A new finding in terms of its association with protophloem was observed. Thus, this enzyme appears to be important for germination of seeds.

Spatial and temporal gene expression during seed germination of Brassica napus

A cDNA-amplified fragment length polymorphism (cDNA-AFLP) approach was used to analyze the spatio-temporal transcript profilings in cotyledons, radicle and hypocotyl of Brassica napus (B. napus) seed during different stages of germination. In total, 75 differentially expressed transcript-derived fragments (TDFs) in cotyledons, 145 in radicle and 145 in hypocotyl were sequenced. Base on BLAST analyses, 37 TDFs in cotyledons, 66 in radicle and 72 in hypocotyl were homologous to genes with known function in the database, respectively. The above known transcripts (175 in total) in three tissues were classified into categories based on their biochemical functions. These genes belong to 14 different functional categories and most of them correspond to metabolic processes, followed by transcription, signal transduction, translation and transport. However, many classes were expressed specifically, including regulation of metabolism and protein function genes in cotyledons, cell rescue and defense genes in radicle, signal transduction genes in hypocotyl. Besides, the expression of eleven identified genes was verified by semi-quantitative reverse transcription polymerase chain reaction in different tissues. The results showed differential gene expression at various germination stages and characteristic gene expression in different seed tissues. Analysis of gene expression profile of B. napus seed is useful for further investigating the seed germination mechanism.

Biochemical composition of broccoli seeds and sprouts at different stages of seedling development

SummaryThe proximate composition, amino acids content, fatty acid profile, total polyphenols, total flavonoids, DPPH radical scavenging capacity and sulforaphane content in broccoli seeds and sprouts were evaluated in this study. The results showed significant differences in the nutritional composition and the phytochemical level in the different stages of germination. Aspartic acid, glutamic acid, serine‐histidine, proline and threonine were found in larger concentrations. Polyphenols and flavonoids content ranged from 9 to 77 mg GAE g−1 and 33 to 117 μg QE g−1, respectively. DPPH radical scavenging capacity ranged from 174 to 674 μmol TE g−1. The sulforaphane content in seeds and sprouts ranged from 273 to 3632 μg g−1–the highest values were found in seeds and the 8‐ and 11‐day‐old sprouts. The results suggest that the broccoli sprouts are a rich source of nutrients and phytochemicals, these have a high potential as functional food.

Alpha and Beta amylase activity of Fagopyrum esculentum (Buckwheat): A Medicinal Plant

Background and Objectives: Fagopyrum esculentum, common buckwheat popularly known as mithe fapar is one of the staple food crops of the mountain region. Traditionally, it is used to treat constipation and bowel upsets. It is also used by diabetic in different parts of Nepal and India. Due to its high nutritive and medicinal value, medical scientist and researchers are interested in developing this as pharmaceutical plant. In this regard department of biochemistry, College of Applied Education and Health Sciences, C.C.S. University, Meerut, India is working to analyse the biochemical composition and benefits of this plant. So, as a part of a multidimensional project of analyzing various components and their impact on health and diseases, here we are reporting the amylase activity during germination of seed in Buckwheat (Fagopyrum esculentum) plant. Methodology: Common buckwheat (Fagopyrum esculentum) seeds were taken and germinated in dark at room temperature from 0 hours to 192 hours. Biochemical analysis for total amylase, alpha and beta amylase activities was measured by the standard method designed by Bernfeld (1955). Results: The seeds of buckwheat showed high level of amylolytic activity during different stages of germination. At 0 hours, negligible amylase activity was found. The first amylase activity was found at 24 hours and increases up to 96 hours. After 96 hours the total amylase activity starts decreasing and becomes almost negligible at 192 hours. Alpha and Beta –Amylase activity is reported separately. Conclusion: The amylases from the buckwheat showed different level of enzymatic activity during seed germination. Alpha amylase contributed a larger account to total amylase activity. The activity starts increasing and becomes maximum at 96 hours and starts decreasing and becomes lowest at 192 hours suggesting that alpha amylase plays a important role in starch metabolism in developing as well as geminating seeds which can be used for the drug discovery and treatment of several diseases like diabetes, polycystic ovary syndrome, constipation, bowel upsets, obesity and others. DOI: http://dx.doi.org/10.3126/jmcjms.v1i1.7887 Janaki Medical College Journal of Medical Sciences (2013) Vol. 1 (1):53-58

Analysis of tyrosine phosphorylation and phosphotyrosine-binding proteins in germinating seeds from Scots pine

Protein tyrosine phosphorylation in angiosperms has been implicated in various physiological processes, including seed development and germination. In conifers, the role of tyrosine phosphorylation and the mechanisms of its regulation are yet to be investigated. In this study, we examined the profile of protein tyrosine phosphorylation in Scots pine seeds at different stages of germination. We detected extensive protein tyrosine phosphorylation in extracts from Scots pine (Pinus sylvestris L.) dormant seeds. In addition, the pattern of tyrosine phosphorylation was found to change significantly during seed germination, especially at earlier stages of post-imbibition which coincides with the initiation of cell division, and during the period of intensive elongation of hypocotyls. To better understand the molecular mechanisms of phosphotyrosine signaling, we employed affinity purification and mass spectrometry for the identification of pTyr-binding proteins from the extracts of Scots pine seedlings. Using this approach, we purified two proteins of 10 and 43kDa, which interacted specifically with pTyr-Sepharose and were identified by mass spectrometry as P. sylvestris defensin 1 (PsDef1) and aldose 1-epimerase (EC:5.1.3.3), respectively. Additionally, we demonstrated that both endogenous and recombinant PsDef1 specifically interact with pTyr-Sepharose, but not Tyr-beads. As the affinity purification approach did not reveal the presence of proteins with known pTyr binding domains (SH2, PTB and C2), we suggest that plants may have evolved a different mode of pTyr recognition, which yet remains to be uncovered.