PLANT OMICS

Transcriptome analysis of Arabidopsis to predict stress length-specific regulatory networks involved in drought response

Sedigheh Akhtartavan*, Majid Talebi

Department of Agricultural Biotechnology, Payame Noor University, Tehran, Iran
Department of Biotechnology, College of Agriculture, Isfahan University of Technology, Isfahan, Iran


Abstract
Drought stress is one of the major environmental factors impairing crops productivity worldwide. Plants use various regulatory genes to reprogram genome activities to cope with such stresses. Among regulatory genes, transcription factors (TFs) function as terminal transducers and directly regulate the expression of wide spectrum of downstream genes. Multiplicity of the TF families and the complex interactions between TFs and cis-elements on the promoters of target genes as well as cross-talk between TFs in response to stress indicate the complexity of signaling networks involved in plant stress responses. This study aimed to use computational and statistical approaches to analyze a microarray dataset from Arabidopsis which covering different time periods of drought stress. After identifying and functional grouping of differentially expressed gens (DEGs), genes encoded TFs were determined and networked based on gene set enrichment analysis (GSE). Hierarchical regulatory network in each condition was assigned. After that, networks were used to conduct network topology analysis. Results indicated an obvious orientation in genome activity toward response to different cues; energy homeostasis and photosynthesis stability was occurred under drought stress. Also, 3787, 2931 and 5115 genes were differentially expressed under the early, moderate and prolonged drought stress, respectively, among them, 169, 140 and 261 TF were identified. Analysis of constructed regulatory networks of each drought condition revealed that plant recruits different but somewhat overlapping strategies to cope with stress in a long period of time. In each drought period, specific or common signaling pathways are activated using several numbers of transcription factors. It seems that among all identified TFs, ARR5, ARR6, ABF3, MYB29, MYB76 and SIGs genes are good candidate to manipulate plant stress tolerance.

Pages 1-9 | Full Text PDF| Supplementary Data| doi: 10.21475/poj.11.01.18.pne735
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Comparative transcriptome analysis of two Gerbera hybrida cultivars revealed cold tolerance mechanism

Shenchong Li, Qinli Shan, Rongpei Yu, Abu Nasar Siddique, Xia Li, Chunmei Yang, Jiwei Ruan, Mingyang Li*

College of Horticulture and Landscape Architecture, Southwest University, Chongqing 400715, China
Flower Research Institute, Yunnan Academy of Agricultural Sciences, Kunming 650205, China
National Engineering Research Center for Ornamental Horticulture, Kunming 650205, China
Kunming Institute of Botany, Chinese Academy of Sciences, Kunming 650201, China


Abstract
Temperature is one of central factors that shape the distribution and growth pattern of Gerbera hybrida. The cultivars of this species have vast variation in the morphology and stress adaptation. In the current study, comparative transcriptome analysis of two haploid cultivars with discriminative phenotype, i.e., Gerbera hybrida Hongjixing and Gerbera hybrida Qiuri, was used to uncover differentially expressed functional unigenes using an Illumina Hiseq2000 sequencing platform. Through high-throughput cDNA sequencing (RNA-seq), we generated 99160 UniGenes from 19.09G primary reads with an average length of 978 bp and an N50 equaling 1620. It was filtered by Trimmomatic and assembled by Trinity. A total of 58127 transcripts were functional annotated by mapping to five protein databases by BLASTX, with topmost differently expressed unigenes (DEGs) directed to transcriptional regulation, secondary metabolism and stress response, etc. Cold treatment (-2℃) of haploid seedlings in intelligent cooled incubator had adverse effects on both cultivars, which was more serious in Qiuri which was exhibited by the lower chlorophyll fluorescence after cold treatment, compared to Hongjixing. This indicates the better cold tolerance of Hongjixing. The qRT-PCR result indicated that besides common pathways, novel pathways like latex biosynthesis potentially contributes to the better cold tolerance of Hongjixing. In view of the ubiquity of cold temperature, this study provided new insights into the mechanism dissection and guidance for cold stress condition.

Pages 10-19 | Full Text PDF| Supplementary Data| doi: 10.21475/poj.11.01.18.pne874
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An efficient heat-shock protocol for transformation of Agrobacterium rhizogenes without spontaneous generation to antibiotic resistance

Juan Manuel Jiménez-Antaño, Josefina Pérez-Vargas, Armando Ariza-Castolo, Octavio Gómez Guzmán, Graciano Calva-Calva*

Biotecnología y Bioingeniería, Centro de Investigación y de Estudios Avanzados del Instituto Politécnico Nacional. Avenida Instituto Politécnico Nacional 2508, Colonia San Pedro Zacatenco, México Distrito Federal. CP 07360, Mexico
Tecnológico de Estudios Superiores de Ecatepec. División Ingeniería Bioquímica, Posgrado en Ingeniería Bioquímica. Av. Tecnológico S/N. Colonia Valle de Anáhuac, Ecatepec de Morelos, Estado de México, CP 55210
Química, Centro de Investigación y de Estudios Avanzados del Instituto Politécnico Nacional. Avenida Instituto Politécnico Nacional 2508, Colonia San Pedro Zacatenco, México Distrito Federal. CP 07360.


Abstract
Antibiotic resistance occurs when the use of antibiotics to treat bacterial infections or to select for transformed bacteria from in vitro cultures becomes ineffective. Although this phenomenon is primarily observed in medicine, it also affects the success of scientific research when bacterial-based genetic transformation experiments are performed. During the agrotransformation of plant cells, tissue, and organs, the acquisition of antibiotic resistance by Agrobacterium spp. has been widely observed, making it difficult to select for the transformed bacteria. The objective of this study was to develop a heat-shock protocol for the transformation of Agrobacterium rhizogenes without the spontaneous generation of antibiotic resistance, to increase its sensitivity and specificity to produce transgenic hairy roots. After the bacterium was transformed, it was cultured in liquid culture medium and plated on solid medium to isolate colonies. The genetic transformation of the bacteria and the plant tissue was verified by PCR and by β-glucuronidase assays. The reproducibility of the method was assessed among the A. rhizogenes strains LBA 9402, A4 and 15834 using streptomycin, kanamycin and ampicillin for selection.

Pages 20-29 | Full Text PDF| doi: 10.21475/poj.11.01.18.pne918
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CrMYC1 transcription factor overexpression promotes the production of low abundance terpenoid indole alkaloids in Catharanthus roseus

Sima Sazegari, Ali Niazi, Farajollah Shahriari-Ahmadi*, Nasrin Moshtaghi, Younes Ghasemi

Department of Plant Breeding and Biotechnology, Ferdowsi University of Mashhad, Iran
Institute of Biotechnology, Collage of Agriculture, Shiraz University, Iran
Department of Pharmaceutical Biotechnology, School of Pharmacy, Shiraz University of Medical Sciences, Iran


Abstract
Catharanthus roseus is known as the only source for the low-abundance anticancer agents namely vinblastine and vincristine. Fine tuning of accumulation of such secondary metabolites is highly governed by the regulatory genes. Among these genes, Catharanthus roseus MYC1 (CrMYC1) is known as one of the key transcription factors regulating the biosynthesis of terpenoid indole alkaloid metabolites in C. roseus. In this study, CrMYC1 coding sequence (AF283506) was isolated and cloned in PBI121 plant binary vector. Then, CRMYC1 was transiently overexpressed in C. roseus leaves using agroinfiltration method. In addition to molecular analysis for confirming CrMYC1 overexpression, the profile of some chief terpenoid indole alkaloids in control and transgenic plants was evaluated by HPLC to elucidate the role of CrMYC1 in an increased in the anticancer components. The results indicated that overexpression of CrMYC1 transcription factor can increase most important terpenoid indole alkaloids including vinblastine, vincristine, and catharanthine in C. roseus. HPLC analysis of catharanthine and vinblastine contents showed about 3 and 2.5 fold increases, respectively, while the increase in vinecristine was not significant compared to that of the control. Therefore, CrMYC1 is introduced as an efficient candidate for manipulating TIA pathway in C. roseus and increasing at least the most valuable terpenoid indole alkaloids in this plant.

Pages 30-36 | Full Text PDF| doi: 10.21475/poj.11.01.18.pne1020
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Isolation and characterization of Rep PepYLCV encoding gene from West Sumatra

Bastian Nova, Elly Syafriani, Lily Syukriani, Alfi Asben, Jamsari Jamsari*

Biotechnology Department, Magister Program, Andalas University, Padang, West Sumatera, Indonesia
Plant Breeding Department, Faculty of Agriculture, Andalas University, Padang, West Sumatera, Indonesia
Department of Food Technology, Faculty of Agriculture, Andalas University, Padang, West Sumatera, Indonesia


Abstract
Replicase (Rep) protein of Geminivirus is known as one of the important components not only for its successful replication in their host but also known to interact with various host plant proteins. However, it is still unclear if those interactions are associated with symptoms level. This research aims to explore the possibility of Rep as pathogenic determinant by in silico approach. Here we report the comparison of three Rep sequences isolated from Pesisir Selatan and Tanah Datar districts in West Sumatra Indonesia. The PCR-based cloning approach was used in this study to isolate the gene sequences from all isolates. Pathogenic determinant was predicted from phenotype and genotype analysis. Phenotype data showed symptoms appearance after 8 dpi for PSSWS14 and 20 dpi for PSSWS3. Furthermore, genotype showed that the nonconserved region in N-terminal of Rep makes different in its putative binding site. It is prospective to be related to the symptoms appearance rates. We predict the differences in N-terminal of Rep affecting the symptoms appearance rates of Geminivirus infection.

Pages 37-41 | Full Text PDF| doi: 10.21475/poj.11.01.18.pne1055
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Transcriptome and gene expression analysis revealed mechanisms for producing high oleoresin yields from Simao pine (Pinus kesiya var. langbianensis)

Wang Yi, Yuan Xiaolong, Hua Mei, Li Jiang, Wang Juan*

Key Laboratory of Forest Plant Cultivation and Utilization, Yunnan Academy of Forestry, Kunming Yunnan 650204, China

Abstract
Simao pine (Pinus kesiya Royle ex Gordon var. langbianensis (A. Chev.) Gaussen) is an important oleoresin source in Yunnan Province, China. However, the molecular mechanisms of high oleoresin yield from Simao pine remain unclear. In this study, RNA-Seq was used to investigate the transcriptome of the species and compare the gene expression profiles of wounded bark from high and low oleoresin-yielding individuals, and the expression levels of genes in the methyl-erythritol 4-phosphate (MEP) pathway were detected by qPCR. A total of 1.41 billion reads were obtained and assembled into 68,881 unigenes from samples of the wounded bark of Simao pine. The KEGG analysis of differentially expressed genes showed that MEP pathway genes were obviously differentially expressed, while most genes in the mevalonate (MVA) pathway were not. The qPCR analysis showed that the MEP pathway gene expression of wounded bark from high oleoresin-yielding genotypes was higher than that of low oleoresin-yielding genotypes. The gene expression differences of 1-deoxy-d-xylulose-5-phosphate synthase (DXS), 4-hydroxy-3-methylbut-2-enyl diphosphate reductase (HDR), and geranylgeranyl pyrophosphate synthase (GGPPs) between high and low oleoresin-yielding genotypes were significant. This implies that DXS, HDR, and GGPPs play important roles in high oleoresin production in Simao pine.

Pages 42-49 | Full Text PDF| doi: 10.21475/poj.11.01.18.pne1085
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Role of sulphate (SO42-) in improvement the growth of rocket plants (Eruca sativa L.) under selenate (SeO42-) levels

Awatif S. Ali*, Atiat M. Hassan, Nabil E. Saber, Eman G Elhosary

Department of Botany, Faculty of Science, Kafr El-Sheikh University, Egypt
Department of Botany and Microbiology, Faculty of Science, Alexandria University, Egypt
Department of Botany and Microbiology, Faculty of Science, Damanhour University, Egypt


Abstract
Selenium is an essential microelement for plant growth and productivity, but high selenium levels impose toxicity in plants. A pot experiment was conducted to investigate the role of sulphate concentrations for ameliorate the adverse effects of high selenate levels on growth of 30-day old rocket plants (cv Egyptian baladi). High Se levels preferentially decreased the biomasses, succulent values in leaves and roots as well as leaf area of rocket plants. This was associated with a suppression of photosynthetic pigments and foliar nutrient elements (K, P, S, Mg and N as NO3-) with an increase of Se content. Supplementation the high Se-containing medium with double SO42- concentration in 1/10 Hoagland solution markedly antagonized Se uptake and its assimilation reflecting the shift- off to some extent- the inhibitory effect of high Se on growth through induction of minerals uptake and biosynthesis of photosynthetic pigments. In addition; changing the protein profile revealed variations in protein patterns and a decrease in selenoprotein biosynthesis by increasing sulphate concentration. However, supplementation of plants with low selenate (2 ppm) level combined with high sulphate (2 ppm) concentration resulted in an optimal growth.

Pages 50-57 | Full Text PDF| 10.21475/poj.11.01.18.pne1112
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Dynamic roles of microRNAs in nutrient acquisition and plant adaptation under nutrient stress: A review

Raheel Shahzad*, Putri Widyanti Harlina, Mohammed Ayaad, Mohamed Ewas, Elsayed Nishawy, Shah Fahad, Hizar Subthain, Mohamed H. Amar

National Key Laboratory of Crop Genetic Improvement and National Center of Plant Gene Research, Huazhong Agricultural University, Wuhan 430070, China
Department of Food Science and Technology, Huazhong Agricultural University, Wuhan 430070, China
Department of Plant Research, Radioisotopes Application Division, Nuclear Research Center of Atomic Energy Authority, Inshas, Egypt
Egyptian Deserts Gene Bank (EDGB), Genetic Resources Department, Desert Research Center (DRC), Cairo, Egypt
Department of Plant Science and Technology, Huazhong Agricultural University, Wuhan 430070, China


†Raheel Shahzad and Putri Widyanti Harlina contributed equally to this work.

Abstract
A constant supply of soil nutrients is critical for the normal growth and development of plants. However, most environments are unstable and this variability depends on numerous factors that include availability of water content, pH, redox potential, an abundance of organic matter as well as microorganisms in soils. To overcome these hurdles and to maintain nutritional homeostasis, plants have evolved sophisticated systems for the continuous provision of soil nutrients necessary for their uninterrupted growth. In this pressing scenario, plant microRNAs (miRNAs) have emerged as a central regulator of nutrients uptake and transport during limited nutrient conditions. Numerous studies establish the intrinsic involvement of miRNAs and their immediate targets facilitating the core mechanisms related to nutrient homeostasis. In this review, we focus on global overview of miRNAs and their dynamic roles involved in keeping nutritional balance within the plants mediated via post-transcriptional regulation by transcript cleavage or translational inhibition of their target mRNAs. In addition, we have also focused on some of the forefront plant adaptations mediated by miRNAs during nutrient deficiency, such as root architecture modifications, transport channel modulation, long distance signaling and subsequent nutrient mobilization through phloem. Moreover, plant strategy to bring out such alterations is a highly perplexing mechanism that requires changes at large scale which involves coordinated regulation of miRNAs and plant hormones at multiple levels. Deciphering the underlying miRNAs-based mechanisms for streamlining nutrient uptake and transport would be a giant step towards solving this conundrum.

Pages 58-79 | Full Text PDF| doi: 10.21475/poj.11.01.18.pne1014


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