بررسی مقایسه‏ ای ترنسکریپتوم برگ برنج تحت شرایط کنترل و تنش خشکی با استفاده از داده ‏های EST

نوع مقاله: علمی پژوهشی

نویسندگان

1 کارشناس بخش زیست شناسی سامانه ها، پژوهشکده بیوتکنولوژی کشاورزی ایران، سازمان تحقیقات، آموزش و ترویج کشاورزی

2 استادیار بخش زیست شناسی سیستم‌ها، پژوهشکده بیوتکنولوژی کشاورزی ایران، سازمان تحقیقات، آموزش و ترویج کشاورزی

3 استادیار، پژوهشکده بیوتکنولوژی کشاورزی ایران، سازمان تحقیقات، آموزش و ترویج کشاورزی، کرج

4 دانشجوی دکترای کامپیوتر، دانشکده کامپیوتر، دانشگاه امیرکبیر، تهران

چکیده

برنج غذای اصلی بیش از نیمی از جمعیت جهان به ویژه کشورهای در حال توسعه است و خشکی مهمترین عامل کاهش عملکرد برنج در آسیا می‌باشد. این تحقیق به منظور شناسایی ژن‌های پاسخ دهنده به تنش خشکی با کمک تجزیه و تحلیل اطلاعات EST دو کتابخانه برگ برنج انجام شد. داده‌های کتابخانه‌های EST در شرایط شاهد و تنش خشکی، از پایگاه ‏داده NCBI دریافت و با استفاده از نرم‏افزار EGassembler ویرایش، دسته‏بندی و هم‏گذاری شدند. توالی‏های کانتیگ و سینگلتون به دست آمده به عنوان الگو برای جستجوی بلاست ‏x در بانک توالی پروتیین برنج و انتساب گروه عملکردی با استفاده از نرم‏ افزار CLC Genomic Workbench و AgriGO به کار برده شدند. پروتئین‌های شناسایی شده در کتابخانه‌های شاهد و خشکی به ترتیب در70 و 82 گروه کارکردی مختلف قرار گرفتند. برای شناسایی تفاوت‌های معنی‌دار بین گروه‌های کارکردی در کتابخانه‌های شاهد و تنش خشکی از نرم‏افزار IDEG6 استفاده شد. بررسی هستی‌شناسی ژن‌ها، تفاوت معنی‏‌دار در 20 گروه عملکردهای مولکولی، 35 گروه فرایندهای زیستی و 12 گروه اجزای درون سلول را آشکار ساخت. به منظور تعیین بیان افتراقی ژن‏ها بین دو کتابخانه، 4012 EST دارای کد Unigene با استفاده از الگوریتم پیاده‏ سازی شده در نرم‏افزار MATLAB انتخاب و با استفاده از نرم ‌افزار IDEG6 تفاوت معنی‌دار بین 42 ژن تحت تنش خشکی نسبت به شاهد، مشاهده شد (31 ژن افزایش و 11 ژن کاهش بیان). ژن‏های افزایش بیان یافته، در پاسخ به تنش‌های محیطی و اکسیداتیو، برقراری هموستازی، پروتئولیز و گلیکولیز نقش داشته و ژن‌های مربوط به فتوسنتز کاهش بیان نشان دادند.

کلیدواژه‌ها

موضوعات


عنوان مقاله [English]

Comparative analysis of rice leaves transcriptome under control and drought stress conditions using EST data

نویسندگان [English]

  • Manzar Heidari 1
  • Zahra-Sadat Shobbar 2
  • Parisa Koobaz 3
  • Mohammad javad Heydari 4
1 Research assistant / Agricultural Biotechnology Research Institute Of Iran, Karaj
2 Assistant Professor, Department of Systems Biology, Agricultural Biotechnology Research Institute of Iran (ABRII), Education and Extension Organization (AREEO), Karaj, Iran.
3 Assistant Professor, Department of Systems Biology, Agricultural Biotechnology Research Institute of Iran (ABRII), Education and Extension Organization (AREEO), Karaj, Iran.
4 Ph.D. Student of Computer, Department of Computer and Information Technology, Amirkabir University of Technology, Tehran, Iran.
چکیده [English]

Rice is the staple food for more than half the world's population, especially in developing countries. Drought is the most yield-limiting factor for rice production in Asia. The current study was conducted to identify the drought stress responsive genes through EST data analysis of two rice leaves libraries. EST libraries data under normal and drought stress conditions were downloaded from NCBI databank. Preprocessing, clustering and assembly of the EST sequences were done using EGassembler software. Generated contig and singleton sequences were used as template for BLASTx analysis against rice protein database and functional category assignment using CLC Protein Workbench software and AgriGO. The identified proteins in the normal and drought libraries were allocated to 70 and 82 functional categories, respectively. IDEG6 were used to identify significant differences between functional categories in control and drought stress libraries. Gene ontology analysis, revealed significant differences in 20 groups of molecular function, 35 groups of biological processes and 12 groups of the intracellular components. In order to find the significant differential expression between the two libraries, 4012 ESTs with unigene accession numbers were implemented through applying an algorithm by MATLAB software and were analyzed by IDEG6 software, where 42 genes were found to be differentially expressed between drought and normal conditions (31 up-regulated and 11 down-regulated genes). The up-regulated genes were involved in environmental and oxidative stress response, homestasis, proteolysis and glycolysis, while photosynthesis related genes were down-regulated.

کلیدواژه‌ها [English]

  • rice
  • Expressed Sequence Tag (EST)
  • Differentially expressed genes
  • Drought
Ali Q, Ahsan M, Tahir M, Elahi M, farooq J and Waseem M, (2011) Gene expression and functional genomics approach for abiotic stress tolerance in different crop species. IJAVMS 5: 221-248.

Allen DJ, Ort DR (2001) Impact of chilling temperatures on photosynthesis in warm climate plants. Trends Plant Sci. 6(1): 36-41.7.

 

1. Ribulose-l,5-bisphosphate carboxylase-oxygenase

2. Gene encoding the small subunit of Rubisco

Amini Z, Hadad R, Moradi F (2009) The Effect of Water Deficit Stress on  Antioxidant Enzymes During Generative Growth Stages in Barley (Hordeum vulgare L.). JWSS-Isfahan University of Technology. 12(46): 65-74.

Asada K (1992) Ascorbate peroxidase–a hydrogen peroxide‐scavenging enzyme in plants. Physiol. Plant. 85(2): 235-241.

Bartholomew DM, Bartley GE, Scolnik PA (1991) Abscisic acid control of rbcS and cab transcription in tomato leaves. Plant Physiol. 96(1): 291-296.

Batlang U, Baisakh N, Ambavaram MM, Pereira A (2013) Phenotypic and physiological evaluation for drought and salinity stress responses in rice. Methods Mol. Biol. 956: 209-225

Brkljacic JM, Samardzic JT, Timotijevic GS, Maksimovic VR (2004) Expression analysis of buckwheat (Fagopyrum esculentum Moench) metallothionein-like gene (MT3) under different stress. J. Plant Physiol. 161(6): 741-746.

Buchanan BB, Balmer Y (2005) Redox regulation: A Broadening Horizon. Annu. Rev. Plant Biol. 56: 187-220.

Cameron KD, Teece MA, Smart LB (2006) Increased accumulation of cuticular wax and expression of lipid transfer protein in response to periodic drying events in leaves of tree tobacco. Plant Physiol. 140(1): 176-183.

Caverzan A, Passaia G, Rosa SB, Ribeiro CW, Lazzarotto F, Margis-Pinheiro M. (2012) Plant responses to stresses: Role of ascorbate peroxidase in the antioxidant protection. Genet. Mol. Biol. 35(4): 1011-1019.

Cheah B. H, Nadarajah K, Divate M. D, Wickneswari R. (2015). Identification of four functionally important microRNA families with contrasting differential expression profiles between drought-tolerant and susceptible rice leaf at vegetative stage. BMC Genomics. 16(1): 692.

Cobbett C, Goldsbrough P (2002) Phytochelatins and metallothioneins: roles in heavy metal detoxification and homeostasis. Annu. Rev. Plant Biol. 53: 159-182.

Degenkolbe T, Do PT, Zuther E, Repsilber D, Walther D, Hincha DK, Köhl KI (2009) Expression profiling of rice cultivars differing in their tolerance to long-term drought stress. Plant Mol. Biol. 69: 133-153.

Dong Q, Kroiss L, Oakley FD, Wang BB, Brendel V (2005) Comparative EST analysis in plant systems. Methods Enzymol. 395: 400-416.

Fleury D, Jefferies S, Kuchel H, Langridge P (2010) Genetic and genomic tools to improve drought tolerance in wheat. J. Exp. Bot. 4: erq152.

Gautam N, Verma PK, Verma S, Tripathi RD, Trivedi PK, et al. (2012) Genome-wide identification of rice class I metallothionein gene: tissue expression patterns and induction in response to heavy metal stress. Funct Integr Genomics 12: 635-647

Gorantla M, Babu PR, Lachagari VR, Reddy AM, Wusirika R, Bennetzen JL, Reddy AR (2007) Identification of stress-responsive genes in an indica rice (Oryza sativa L.) using ESTs generated from drought-stressed seedlings. J. Exp. Bot. 58: 253-265.

Gorantla M, Babu PR, Lachagari VR., Feltus FA, Paterson AH, Reddy AR (2005) Functional genomics of drought stress response in rice: transcript mapping of annotated unigenes of an Indica rice (Oryza sativa L. cv. Nagina 22). Curr. Sci-Bangalore. 89(3): 496.

Hajheidari M, Eivazi A, Buchanan BB, Wong JH, Islam M, Hosseini Salekdeh Gh (2007) Proteomics Uncovers a Role for Redox in Drought Tolerance in Wheat. ‎J. Proteome Res. 6: 1451-1460.

Hazen SP, Pathan M. S, Sanchez A, Baxter I, Dunn M, Estes B, Nguyen HT (2005). Expression profiling of rice segregating for drought tolerance QTLs using a rice genome array. Funct. Integr. Genomics. 5(2): 104-116.

Heidari P, Maleki zanjani B, Heidary Sh (2012) A study of gene expression and functional genomics of wheat, rice, cotton and festuca plants under drought stress by analyzing expressed sequence tags (EST). Genetic Novin 2: 129-140. 

Hosseini Salekdeh Gh, Siopongco J, Wade LJ, Ghareyazie B, Bennett J (2002) Proteomic analysis of rice leaves during drought stress and recovery. Proteomics 2(9): 1131-1145.

Ji K, Wang Y, Sun W, Lou Q, Mei H, Shen S, Chen H (2012). Drought-responsive mechanisms in rice genotypes with contrasting drought tolerance during reproductive stage. J. Plant Physiol. 169: 336-344

Kader JC (1996) Lipid-transfer proteins in plants. Annu. Rev. Plant biol. 47(1): 627-654.

Karpinski S, Escobar C, Karpinska B, Creissen G, Mullineaux PM (1997) Photosynthetic electron transport regulates the expression of cytosolic ascorbate peroxidase genes in Arabidopsis during excess light stress. Plant Cell. 9: 627-640.

Kobrehel K, Wong JH, Balogh A, Kiss F, Yee BC, Buchanan BB (1992) Specific reduction of wheat storage proteins by thioredoxin h. Plant Physiol. 99: 919-924.

Kumari S, Roy S, Singh P, Singla-Pareek S, Pareek A (2013) Cyclophilins: proteins in search of function. Plant Signal Behav. 8(1): e22734.

Lozano RM, Wong JH, Yee BC, Peters A, Kobrehel K, Buchanan BB (1996) New evidence for a role for thioredoxin h in germination and seedling development. Planta. 200: 100-106.

Ma DP, Tan H, Si Y, Creech RG, Jenkins JN (1995) Differential expression of a lipid transfer protein gene in cotton fiber. Biochim. Biophys. Acta. 1257(1): 81-84.

Maclean JL, Hettle GP (Eds) (2002) Rice almanac: Source book for the most important economic activity on earth. IRRI (free PDF download) CABI.

Manickavelu A, Kawaura K, Oishi K, Shin T, Kohara Y, Yahiaou N ... & Yano K (2012) Comprehensive functional analyses of expressed sequence tags in common wheat (Triticum aestivum). DNA Res. 19(2): 165-177.

Marx C, Wong JH, Buchanan BB (2003) Thioredoxin and germinating barley: targets and protein redox changes. Planta. 216: 454-460.

Masoudi-Nejad A, Tonomura K, Kawashima S, Moriya Y, Suzuki M, Itoh M, Kanehisa M, Endo T, Goto S (2006) EGassembler: online bioinformatics service for large-scale processing, clustering and assembling ESTs and genomic DNA fragments. Nucleic Acids Res. 34: 459-462.

Mohammadi PP, Moieni A, Komatsu S (2012) Comparative proteome analysis of drought-sensitive and drought-tolerant rapeseed roots and their hybrid F1 line under drought stress. Amino Acids. 43(5): 2137-2152.

Nagaraj SH, Gasser R.B, Ranganathan S (2007) A hitchhiker's guide to expressed sequence tag (EST) analysis. Brief. Bioinform. 8: 6-21.

Navabpour S, Morris K, Allen R, Harrison E, Mackerness S, Buchanan-Wollaston V (2003) Expression of senescence-enhanced genes in response to oxidative stress. J. Exp. Bot. 54: 2285-2292.

Ogata O, Suzukim H (2011) Plant expressed sequence tags databases: practical uses and the improvement of their searches using network module analysis. Plant Biotech. 28: 351-360.

Orosz F, Olah J, Ovadi J (2006) Triosephosphate isomerase deficiency: facts and doubts. IUBMB Life. 58(12): 703-715.

Ramamoorthy R, Jiang SY, Kumar N, Venkatesh PN, Ramachandran S (2008) A comprehensive transcriptional profiling of the WRKY gene family in rice under various abiotic and phytohormone treatments. Plant Cell Physiol. 49: 865-879.

Rosa SB, Caverzan A, Teixeira FK, Lazzarotto F, Silveira JA, Ferreira-Silva SL, ... & Margis-Pinheiro M (2010). Cytosolic APx knockdown indicates an ambiguous redox responses in rice. Phytochemistry. 71(5): 548-558.

Serrato AJ, Cejudo FJ (2003) Type-h thioredoxins accumulate in the nucleus of developing wheat seed tissues suffering oxidative stress. Planta. 217(3): 392-399.

Shaar-Moshe L, Hübner S, Peleg Z. (2015). Identification of conserved drought-adaptive genes using a cross-species meta-analysis approach. BMC Plant Biol. 15(1): 1.

Spreitzer RJ (2003) Role of the small subunit in ribulose-1, 5-bisphosphate carboxylase/oxygenase. Arch. Biochem. Biophys. 414(2): 141-149.

Stewart GA (2000) The molecular biology of allergens (Chapter 72). In: Busse W.W, Holgate S.T (ed) Asthma and Rhinitis, Blackwell Science Ltd USA, 1113-1114.

Szira F, Börner A, Neumann K, Nezhad K. Z, Galiba G, Bálint AF (2011) Could EST-based markers be used for the marker-assisted selection of drought tolerant barley (Hordeum vulgare) lines?. Euphytica. 178(3): 373-391.

Talame V, Ozturk NZ, Bohnert HJ, Tuberosa R. (2007) Barley transcript profiles under dehydration shock and drought stress treatments: a comparative analysis. J. Exp. Bot. 58(2): 229-240.

Williams J, Bulman MP, Neill SJ (1994) Wilt induced ABA biosynthesis, gene expression and down regulation of rbcS mRNA levels in Arabidopsis thaliana. Physiol. Plant. 91(2): 177-182.

Xoconostle-Cazares B, Ramirez-Ortega FA, Flores-Elenes L, Ruiz-Medrano R (2010) Drought tolerance in crop plants. Am. J. Plant Physiol. 5(5): 1-16.

Xu Y, Harris-Haller LW, McCollum JC, Hardin SH, Hall TC (1993) Nuclear gene encoding cytosolic triosephosphate isomerase from rice (Oryza sativa L.). J. Plant physiol. 102(2): 697.

Yang Z, Wu Y, Li Y, Ling HQ, Chu C (2009) OsMT1a, a type 1 metallothionein, plays the pivotal role in zinc homeostasis and drought tolerance in rice. Plant Mol. Biol. 70(1-2): 19-229.

Zhang J, Kirkham MB (1994) Drought-Stress-Induced Changes in Activities of Superoxide Dismutase, Catalase, and Peroxidase in Wheat Species. Plant Cell Physiol. 35(5): 785-791.

Zhou GK, Xu YF, Liu JY (2005) Characterization of a rice class II metallothionein gene: tissue expression patterns and induction in response to abiotic factors. J. Plant Physiol. 162: 686-696.

Zhou J, Wang X, Jiao Y, Qin Y, Liu X, He K, … & Deng XW (2007) Global genome expression analysis of rice in response to drought and high-salinity stresses in shoot, flag leaf, and panicle. Plant Mol. Biol. 63(5): 591-608.