8(5) 2015 issue
September 2015 issue
Southern Cross Publishing Group©2015
Proteome analysis of wheat embryo (Triticum aestivum) Sensu stricto germination under osmotic stress
Ali Shayanfar, Reza Tavakkol Afshari*, Houshang Alizadeh
Department of Agronomy and Plant Breeding, College of Agriculture and Natural Resources, University of Tehran, Karaj, Iran
Seed germination is an important growth phase in the plant life cycle that can be injured by osmotic stress. Germination is divided into three phases and many processes are initiated before radical protrusion. Many metabolic activities occur, especially in Sensu stricto germination phase since the seed embryo is a live organ, so differences in protein patterns are important in this phase. This study was carried out to find out the effects of osmotic stress on protein patterns in Sensu stricto germination phase. Effects of osmotic stress (-12 bar) on T. aestivum embryo (Sardari and Qods, tolerant and susceptible cultivars, respectively) proteins were evaluated. An osmotic potential of -12 bar was selected as the critical stress level, because at this treatment, germination percentage in susceptible cultivar was too low, but in tolerant cultivar was 50% as compared to control condition. Osmotic stress decreased germination percentage and caused adverse effects on protein patterns. After 2-DE proteome analysis of wheat embryo axis, 1000 spots were reproducible, although 16 spots were expressed differentially. Five protein categories were identified by MALDI-TOF/TOF in 16 differentially expressed, including energy (19%), stress/disease /defense (25%), metabolism (37%), protein synthesis/degradation (13%), and signal transduction (6%). Therefore, it should be noted that seeds were acting as intelligent systems which has an important role in response mechanism to osmotic stress during germination, especially before radical protrusion. In this study, it was demonstrated that stress and metabolism-related proteins such as HSPs, 1-Cys peroxiredoxin, alpha amylase inhibitor, and ADP-glucose pyrophosphorylase were more than other protein classes had vital roles in response to osmotic stress at 24 HAI. The results indicated that protein abundance under osmotic stress depends on the severity and amount of stress and imbibition time.
Pages 372-380 | Full Text PDF
Molecular modeling and in-silico characterization of Glycine max Inositol (1, 3, 4) tris 5/6 kinase-1 (Gmitpk1) - a potential candidate gene for developing low phytate transgenics
Veda Krishnan, Priyanka Jain, Vinutha T, Alkesh Hada, Manickavasagam M, Ganapathi A, Raj D Rai, Archana Sachdev*
Division of Biochemistry, Indian Agricultural Research Institute (IARI), New Delhi, India
National Research Centre on Plant Biotechnology (NRCPB), New Delhi, India
Department of Biotechnology and Genetic Engineering, Bharathidasan University, Tiruchirappalli, India
Inositol (1, 3, 4) tris 5/6 kinase (Itpk) is a key player in lipid-independent pathway of phytate biosynthesis. In this study, the full length coding sequence of Gmitpk1 was cloned and blasted to retrieve the available inositol phosphate kinases (ipks) from the public domain. Sequence analysis of the selected 18 plant kinases revealed a consensus ATP-grasp domain. Secondary structure predictions showed high alpha helix content (80.02%) which justified the structural flexibility as well as the versatility of these kinases. Homology modelling of the Gmitpk1 performed using the template crystal structure of inositol tetrakisphosphate-1-kinase (2q7d.1.A) from Homo sapiens revealed the presence of N and C-terminal domains with a mixed a/฿ topology having an active site located in the deep cleft between the domains. The model was further refined using intrinsic dynamic tools like ProSA, Verify3D, WEBnm and ElNemo. This study has enabled the elucidation of the 3Dstructure of Gmitpk1and the data has been submitted to protein model data base (PMD) - PM0079572 (first report). Ligand binding residues and energy computations have revealed Mg2+, ATP and ADP as the most likely ligands for Gmitpk1. The study throws light on some novel insights into the structural features of Gmitpk1, a potential candidate for developing low phytate transgenic soybean.
Pages 381-391 | Full Text PDF
Selection of spring bread wheat genotypes for resistance to cereal cyst nematode (Heterodera avenae Woll.) based on field performance and molecular markers
Khaled A. Moustafa, Abdullah A. Al-Doss, Mohamed I. Motawei*, Solaiman Al-Otayk, Ahmed A.M. Dawabah, Ahmed L. Abdel-Mawgood, Suloiman M. Al-Rehiayani, Ahmad S. Al- Hazmi
College of Food and Agricultural Science, King Saud University, Saudi Arabia
Agricultural Research Center, Field Crops Research Institute, Egypt
Plant Production and Protection Department, Al-Qassim University, Saudi Arabia
The cereal cyst nematode (CCN), Heterodera avenae Woll., is a devastating root nematode parasite of wheat (Triticum aestivum L.). This study aimed to screen wheat germplasm for resistance to CCN. The performance of 17 genetically diverse wheat genotypes (local and international material) were evaluated for two years (2009 and 2010) in a H. avenae-naturally-infested field at the Hial region, north Saudi Arabia. Results show that the tested wheat genotypes were significantly different in field performance and resistance to CCN. The grain yield ranged from 4.58 tons/ha for cv. Yecora Rojo (the susceptible) to 8.2 tons/ha for the genotype 15-SAWYT-31. Ten local genotypes were designated as resistant. The local cv. KSU 119 was the most resistant genotype (no. cysts/plant = 0.7) among all the genotypes tested. In addition, microsatellite markers linked to Cre1 and Cre3 genes were used. The dendogram generated using SSR data divided wheat genotypes into two main clusters. Ten out of 17 wheat genotypes (LNM-72, LNM-99, LNM-126, LNM-136, KSU118, L11-8, L11-17, L11-21, KSU 119, and AUS-30851) had both Cre genes and were found in the same sub-cluster. All these genotypes, except AUS-30851, LNM-72 and L11-17, were found to be the resistant to CCN. Therefore, Cre3, Cre1 and other Cre resistance genes are now used in our marker-assisted selection (MAS) programs to identify CCN-resistant wheat genotypes.
Pages 392-397 | Full Text PDF
Molecular and functional characterization of the chalcone synthase gene (DcCHS1) promoter in response to hormones
Jia-Yuan Wang, Hao-Fu Dai, Hui-Liang Li, Dong Guo, Wen-Li Mei*, Shi-Qing Peng*
Key Laboratory of Biology and Genetic Resources of Tropical Crops, Ministry of Agriculture, Institute of Tropical Bioscience and Biotechnology, Chinese Academy of Tropical Agricultural Sciences, Haikou 571101, China
Dragons blood, which is widely used in traditional medicines, is a red resin that forms in the stem of Dracaenas plants after they are wounded. Flavonoids are the main compounds in dragon's blood. Research on dragons blood has focused mainly on its chemical composition and pharmacological activity, whereas the molecular mechanism of dragons blood formation is still unknown. Chalcone synthase (CHS) is the first key enzyme in biosynthesis of flavonoids. However, little information is available on regulation of CHS gene expression in Dracaenas plants. In this study, a 1404-bp promoter region of DcCHS1 was cloned from Dracaena cambodiana. A typical TATA-box was located at -24 bp to -20 bp upstream from the transcription start site (TSS) of DcCHS1. Several typical regulatory elements were found in the DcCHS1 promoter region, including cis-acting elements involved in light responsive, ABRE for responding to abscisic acid, TCA-element for responding to salicylic acid, TGA-element for auxin, and TC-rich repeat elements involved in defense and stress responsiveness. Promoter deletion analysis revealed that the DcCHS1 promoter induces the specific expression of the GUS gene, and 149 nucleotides upstream of the TSS were sufficient for the expression function. Transient gene expression assay revealed that the expression of DcCHS1 promoter was positively regulated by methyl jasmonate and 6-benzylaminopurine.
Pages 398-404 | Full Text PDF
Modulated gene expression during the cold acclimation process in tolerant and sensitive clones of cultivar Leccino (Olea europaea L.)
Rodolfo Bernardi*, Giorgio Bartolini, Raffaella Petruccelli, Mariangela Salvini, Mauro Durante
Department of Agriculture, Food and Environment, University of Pisa, Via del Borghetto, 80, 56124 Pisa, Italy
Trees and Timber Institute, National Research Council of Italy, Via Madonna del Piano 10, 50019 Sesto Fiorentino (Florence), Italy
Scuola Normale Superiore, Piazza dei Cavalieri, 7, 56126 Pisa, Italy
Biotechnology Centre - UEM, Maputo, Mozambique
The plants are constantly exposed to environmental changes that need to be addressed with appropriate modifications of gene expression. The genetic approach to the study of the response to biotic and abiotic stresses is of great importance in plants, for the identification of the genes involved and their activation mechanisms. In our work, we have characterized the expression of some genes induced during the acclimation under low temperatures in plants of Olea europaea L., by isolating DNA sequences differentially expressed in a cold tolerant clone of Leccino cultivar by using the suppression subtractive hybridisation (SSH). The sequences obtained were analysed by sequencing. Some sequences of the libraries and from orthologous genes expressed in various abiotic/biotic stress treatments, were labelled and used as probes for slot blot hybridization with total RNAs extracted from cold sensitive and tolerant Leccino clones treated with decreasing temperatures down to -10 ฐC, thus mimicking a process of cold acclimation. Six genes were found to be of interest for the adaptation to cold stress, whose expression is differentially modulated in the two clones. The most important result was in a bimodal response during the acclimation phases, the causes of which have been discussed.
Pages 405-411 | Full Text PDF
Identification of acting domains mediating the protein interactions between flowering repressors flowering locus C (FLC) and short vegetative phase (SVP) from birch (Betula platyphylla Suk.)
Tangchun Zheng, Lina Zang, Lijuan Dai, Yue Zhao, Chuanping Yang*, Guan-Zheng Qu*
State Key Laboratory of Tree Genetics and Breeding, Northeast Forestry University, Harbin 150040, China
College of Life Science, Northeast Forestry University, Harbin 150040, China
Flowering, transition from vegetative to reproductive phase in plants, is regulated by both endogenous and environmental signals. Flowering locus C (FLC) and short vegetative phase (SVP) are the key genes in delaying flowering. For study on the mechanism of interaction between SVP and FLC in birch, the test was performed by yeast two-hybrid. The target genes (BpFLC and BpSVP) and truncated genes of BpFLC1~6 and BpSVP1~6 were cloned from subcloning vectors, digested with Nde I and EcoR I, then ligased into yeast expression vector pGBKT7 and pGADT7. The test of transformed strains in different conditions showed there were no self-activated abilities and no toxic effects in yeast bait vectors of pGBKT7-BpFLC and pGADT7-BpSVP. The process of yeast two-hybrid indicated there was a direct interaction between BpFLC and BpSVP. Further study showed Y2HGold [BD-BpFLC2~5 ื AD-BpSVP] could grow on selective agar plates TDO, QTO/A, or QDO/X/A with blue stains indicating that BpFLC2~5 truncated forms and BpSVP protein could act with each other to form heterodimers. In addition, the result of yeast two-hybrid also showed that BpFLC and BpSVP2~5 truncated forms could combine with each other to activate downstream reporter genes. The interactions between BpFLC3 and BpSVP3 strongly showed that the K domain of BpFLC and the K domain of BpSVP were the key structure domains and mediated the protein interactions between BpFLC and BpSVP. These results greatly increase our understanding of the mechanisms extending the vegetative phase and enhance genetic studies of birch traits and other woody plants.
Pages 412-419 | Full Text PDF | Supplementary Data xls
Genome-wide analysis and functional identification of the annexin gene family in maize (Zea mays L.)
Zhongbao Zhang, Xianglong Li, Meng Han, Zhongyi Wu*
Beijing Agro-Biotechnology Research Center, Beijing Academy of Agriculture and Forestry Sciences, Beijing 100097, China
Beijing Key Laboratory of Agricultural Genetic Resources and Biotechnology, Beijing 100097, China
College of Agriculture, Yangtze University, Hubei Collaborative Innovation Center for Grain Industry, Jingzhou 434023, Hubei, China
Annexins have previously been identified and characterized in Arabidopsis and rice. They compose a multigene family in plants. In this study, we indicated the isolation and characterization of maize annexin genes across a whole genome using bioinformatics, microarray and real-time PCR methods. A total of 12 members of this family were identified in the maize genome. The 12 maize annexins were distributed on eight maize chromosomes. Multiple alignment and motif display results revealed that most maize annexin proteins contained 1-4 annexin repeats. A phylogenetic analysis indicated the maize annexin gene family could be divided into four subfamilies. In brief, putative cis-elements involved in abiotic stress response, phytohormones, pollen-specific elements and seed development were observed in the promoters of maize annexin genes. Microarray data showed that the maize annexin genes had tissue-specific expression patterns in the maize developmental steps. The QRT-PCR analysis result indicated that all 12 genes were induced in the seedling leaves by PEG and NaCl.
Pages 420-428 | Full Text PDF | Supplementary Data PDF
Expression profiling and down-regulation of three histone lysine methyltransferase genes (PvATXR3h, PvASHH2h, and PvTRX1h) in the common bean
Aar๓n Barraza, Francisco Luna-Martํnez, Joselyn Cristina Chแvez-Fuentes, Ra๚l มlvarez-Venegas*
Centro de Investigaci๓n y de Estudios Avanzados del IPN, Unidad Irapuato, Guanajuato, M้xico
Universidad Polit้cnica de Guanajuato, Universidad Norte s/n, Juan Alonso Cortazar, Guanajuato, M้xico
Unidad Profesional Interdisciplinaria de Ingenierํa del IPN, Campus Guanajuato, Guanajuato, M้xico
Histone methylation is a conserved epigenetic mechanism in eukaryotes. Most of the histone lysine methyltransferases (HKMTases) conferring such modifications are proteins with a conserved SET domain responsible for enzymatic activity. Genetic studies in Arabidopsis thaliana have revealed that proteins from the Trithorax group (TrxG) are critical in activating transcription by methylating lysine 4 and lysine 36 of histone H3. Two TrxG proteins, ATXR3 and ATX1 (also called SET DOMAIN GROUP 2 and 27, respectively) are necessary for global genome-wide H3K4me3 deposition in Arabidopsis, whilst ASHH2 (also called SDG8) is a multi-functional enzyme with H3K4 and H3K36 methylation activity. Using phylogenetic analysis, we have identified the common bean (Phaseolus vulgaris L.) gene orthologs to Arabidopsis ATXR3, ASHH2, and ATX1 genes, which we have designated PvATXR3h, PvASHH2h, and PvTRX1h, respectively. Analysis of these genes with qRT-PCR reveals that all three are broadly expressed during plant and nodule development. Through a reverse genetics approach, we created common bean composite plants to knock-down PvATXR3h, PvTRX1h, and PvASHH2h expression. From analysis of the transgenic root phenotype, we conclude that transgenic root growth and development in the common bean was hindered by PvASHH2h gene downregulation.
Pages 429-440 | Full Text PDF
Analysis of ESTs from the date palm (Phoenix dactylifera L.) cv. Sukary, an elite variety
Sulieman A. Alfaifi, Muhammad A. Khan, Hussein M. Migdadi, Jernej Jakse, Megahed H. Ammar, Ehab H. El-Harty, Mohammad I. Althamrah, Muhammad Afzal, Muhammad M. Javed, Salem S. Alghamdi
Legume Research Group, Plant Production Department, Faculty of Food and Agricultural Sciences, King Saud University, P.O .Box 2460, Riyadh 11451, Saudi Arabia
Agronomy Department, Biotechnical Faculty, University of Ljubljana, Jamnikarjeva 101, SI-1000 Ljubljana, Slovenia
Expressed sequence tags (ESTs) were generated from a normalized cDNA library of the date palm cv. Sukary to uncover the transcriptomic profile of this well-known high-quality variety in Saudi Arabia. The RNA was isolated from leaves and developed fruits at different stages from a single Sukary female tree. Total RNA was pooled and the normalized cDNA were cloned into a pAL17.3 cloning vector. The positives clones were sequenced using BigDyeฎ Terminator chemistry. A total of 6943 high quality ESTs were generated, out of which 6671 ESTs were submitted to (LIBEST_028537). The generated ESTs were assembled into 6362 unigenes, consisting of 494 (14.4%) contigs and 5868 (84.53%) singletons. The functional annotation showed that the majority of the ESTs were associated with binding (44%), catalytic (40%), transporter (5%) and structural molecular activities (5%). The Blastx results showed that 73% of the unigenes have significant similarities to known plant genes and 27% were novel. The latter could be of particular interest in date palm genetics studies. Further analysis showed that some ESTs were categorized as stress/defense and fruit development related genes. These newly generated ESTs could significantly enhance date palm EST databases that are in the public domain and are available to scientists and researchers across the globe.
Pages 441-448 | Full Text PDF | Supplementary Data PDF | Supplementary Data xls
Alleviation of salinity stress in faba bean (Vicia faba L.) plants by inoculation with plant growth promoting rhizobacteria (PGPR)
Ehab M.R. Metwali, Tamer S. Abdelmoneim, Mostafa A. Bakheit, Naif M.S. Kadasa
Genomic and Biotechnology Division, Biological Science Department Faculty of Science, University of Jeddah, Saudi Arabia
Microbiology Division, Biological Science Department Faculty of Science, University of Jeddah, Saudi Arabia
Department of Agricultural Botany, Faculty of Agriculture, Suez Canal University, 41522 Ismailia, Egypt
Department of Legume Research, Field Crop Research Institute, Agriculture Research Center, Cairo, Egypt
The effects of three bacterial isolates (Pseudomonas putida, P. fluorescens and Bacillus subtilis) of plant growth promoting rhizobacteria were investigated to improve the growth of six cultivars from Vicia faba L. under two levels of salinity stress (S1=4000 and S2=8000 ppm). Different morphological and biochemical parameters were studies. The highest values of final germination percent were observed in cultivar Wadi 1 and Line 1 (96%) in the presence of P. fluorescens. Application of P. putida decreased the mean germination time in the presence or absent salinity stress. The salinity treatments induced a significant decreased in all of plant growth parameters, plant chlorophyll and plant soluable proteins, while plant proline was increased, comparing to control. The plants treated with P. fluorescens showed significant increase in growth traits such as plant length (10.66%), plant shoot fresh weight (9.52%) and plant leaf area (61.86%). This increasing trend was followed by application of B. subtilis then P. putida. The phylogenetic diversity relationship and the level of genetic diversity among the cultivars were assessed using 9 arbitrary primers. The results indicated that 6 out of the 9 primers [(GATA)4GC, (AGAC)4GC, GAC(GATA)4, (AC)8YC, CGC(GATA)4 and (AG)8YC] can generate clear multiplex banding profiles. Among the 48 polymorphism bands, only 14 were found to be useful as positive or negative markers related to salt stress. The cluster analysis classified the cultivars into two main groups. The inter-simple sequence repeat can be used to identify alleles associated with the salt stress in faba bean germplasm.
Pages 449-460 | Full Text PDF
Antioxidant enzymes metabolism and cellular differentiation during the developmental stages of somatic embryogenesis in Torilis japonica (Houtt.) DC
Abinaya Manivannan, Sonali Jana, Prabhakaran Soundararajan, Chung Ho Ko, Byoung Ryong Jeong*
Division of Applied Life Science (BK21 Plus), Graduate School, Gyeongsang National University, Jinju, Korea 660-701
Institute of Agriculture and Life Science, Gyeongsang National University, Jinju, Korea 660-701
Research Institute of Life Science, Gyeongsang National University, Jinju, Korea 660-701
In the present study, an efficient protocol for high frequency somatic embryogenesis from stem explants of Torilis japonica was established. Explants were cultured in Murashige and Skoog (MS) medium containing 0-2.0 mgทL-1 2,4- dichlorophenoxy acetic acid (2,4-D) with or without the combination of indole-3-butyric acid (IBA). The embryogenic callus induction and somatic embryogenesis frequency were measured at four week intervals. The highest embryogenic callus induction (100ฑ0.0%) was achieved in 2.0 mgทL-1 2,4-D and 1.0 mgทL-1 IBA. Maximum somatic embryo induction (100ฑ0.0%) was noted on the MS medium augmented with 2.0 mgทL-1 2, 4-D, 1.0 mgทL-1 IBA, and 1.0 mgทL-1 gibberellic acid (GA3) with an average of 75.8ฑ3.7 somatic embryos per explant. Scanning electron microscopic investigation and histological analysis demonstrated the systemic development of somatic embryos. Comparatively, the embryogenic callus induction and plantlet conversion stages possessed more total protein and hydrogen peroxide content than other stages. As a consequence, the activities of superoxide dismutase (SOD), guaiacol peroxidase (GPX), ascorbate peroxidase (APX), and catalase (CAT) were also increased during the induction of somatic embryos and bipolar shoot-root axis differentiation stages. Clonal fidelity assessment by random amplified polymorphic DNA (RAPD) fingerprinting and inter sequence specific repeats (ISSR) markers displayed the monomorphic banding pattern across the micropropagated plantlets. Thus, the RAPD and ISSR markers validated the genetic homogeneity or the true-to-type nature of the in vitro plants.
Pages 461-471 | Full Text PDF
Anthocyanin contents and composition of VlmybA1-2 and VlmybA2 genes in Vitis labrusca hybrid grape cultivars and cross seedlings
Eun Su Kim, Eun Ha Chang, Youn Young Hur, Tae Wan Kim, Sung Min Jung*
Fruit Research Division, National Institute of Horticultural and Herbal Science, Suwon 440-706, Korea
Department of Plant and Environmental Science, Hankyoung National University, Ansung 456-749, Korea
The grape Vitis labrusca and hybrids from V. vinifera ? V. labrusca have different haplotypes of Myb gene combinations from the V.vinifera grape. The Myb gene PCR results show that the white-skinned V.labrusca hybrid cultivar Cheongsoo has a Myb allele contained in two pairs of haplotypes A (VvmybA1a) like the V.vinifera grape. However, other red-skinned V.labrusca hybrid grape cultivars have haplotype A2 (containing VlmybA2), haplotype A1-2 (containing VlmybA1-2), or both (Hap A1-2 and A2). The V.labrusca hybrid cultivars containing the haplotype A2 show higher anthocyanin contents. However, the anthocyanin content of the haplotype A2 was significantly higher than the haplotype A1-2 in the seedlings from the cross between the Campbell Early grape (V. labrusca) ?the Neo Muscat grape (V. vinifera). These results show that the VlmybA2 gene is closely related to anthocyanin contents and grape skin color in the V.labrusca hybrid grape cultivars and their cross seedlings.
Pages 472-478 | Full Text PDF