Ocultar
Mostrar todos
Extracción de ARN total en plantas y hongos filamentosos
Resumen
La alta calidad de ARN total requerida como material de partida es un elemento común en la mayoría de las técnicas utilizadas en estudios moleculares tanto en plantas como en hongos fitopatógenos. En este manuscrito se profundiza en las características de cada uno de estos tejidos y la conveniencia del uso de determinados protocolos, los cuales van a estar, esencialmente, diferenciados por la aplicación de diferentes tipos de detergentes. También, se afrontan las posibles soluciones que se pueden dar para eliminar los altos contenidos de polisacáridos que dificultan la extracción de ARN total. Además, se hace referencia a protocolos de extracción de ARN efectivos para obtener buenos rendimientos en la interacción planta-hongo filamentosos. Como elemento común, tanto para plantas como para hongos filamentosos, se hace una descripción detallada de los pasos a seguir en la obtención de ARN total y se explica el porqué de cada uno de ellos, así como las diferentes variantes cuando se encuentran problemas relacionados con la estructura de los tejidos sujetos a investigación.
Palabras clave: detergentes, fenoles, polisacáridos
Referencias
Adams, RP, Pandey RN, Flournoy LE (1996) Inhibition of random amplified polymorphic DNAs (RAPDs) by plant polysaccharides. Plant Molecular Biology 14: 17-22
Ambion (2008) The basics: RNA isolation. En: http:// www.ambion.com/techlib/basics/ rnaisol/index.html#1 (consulta: abril 2008).
Barlow, JJ, Mathias A P, Williamson R, Gammack DB (1963) A simple method for the quantitative isolation of undegraded high molecular weight ribonucleic acid. Biochemical Biophysic Research Communications 13: 61-66
Carmona-Ribeiro, AM, Viera DB (2006) Cationic lipids and surfactants as antifungal agents:mode of action. Journal of Antimicrobial Chemotherapy 58: 760-767
Chang, SJ, Puryear, Caimey J (1993) A simple and efficient method for isolating RNA from pine trees. Plant Molecular Biology Reporter 11: 113-116
Chomczynski, P (1993) A reagent for the single-step simultaneous isolation of RNA, DNA and proteins from cell and tissue samples. Biotechniques 15: 532-537
Claros, MG y Canovas FM (1998) Rapid high quality RNA preparation from pine seedlings. Plant Molecular Biology Reporter 16: 9-18
Dawson, RM, Elliott CDC, Elliott WH, Jones KM (1986) Vitamins and coenzymes. En: Dawson RMC, Elliot DC y Jones KM (Eds) Data for Biochemical Research. Oxford Clarendon Press, UK.
Giambernardi, TA, Ulrich R, Klebe RJ (1998) Bovine serum albumin reverses inhibition of RT-PCR by melanin. Biotechniques 25: 564-566
Jelle, DK, Isabel RR, Erik VB, Arne H, Denis DK (2006) Efficient extraction of high-quality total RNA from various hop tissues. Biochemistry and Biotechnology 36: 355-362
Biotecnología Vegetal Vol. 11, No. 4, 2011
Jeon, J, Park SY, Chi MH, Choi J, Park J, Rho HS, Kim S, Goh J, Yoo S, Choi J, Park JY, Yi M, Yang S, Kwon MJ, Han SS, Kim BR, Khang CH, Park B, Lim SE, Jung K, Kong S, Karunakaran M, Oh HS, Kim H, Kim S, Park J, Kang S, Choi WB, Kang S, Lee YH (2007) Genome-wide functional analysis of pathogenicity genes in the rice blast fungus. Nature Genetics 39: 561-565
John, ME (1992) An efficient method for isolation of RNA and DNA from plants containing polyphenolics. Nucleic Acids Research 20: 2381
Kiefer, Werener EH, Dieter E (2000) A simple and efficient protocol for isolation of functional RNA from plant tissues rich in secondary metabolites. Plant Molecular Biology Reporter 18: 33-39
Kim, SH, Hamada T (2005) Rapid and reliable method of extracting DNA and RNA from sweetpotato, Ipomoea batatas (L). Lam. Biotechnology Letter 27: 1841-1847
Kwon-Chung, KJ, Rhodes JC (1986) Encapsulation and melanin formation as indicators of virulence in Cryptococcus neoformans. Infection and Immunology 51: 218-223
Lakhvir, L, Rashmita S, Rajesh K G, Priti S, Sanjay K (2001) RNA Isolation from high-phenolic tea leaves and apical buds. Plant Molecular Biology Reporter 19: 181a-181f
Liao, Z, Chen M, Guo L, Gong Y, Tang F, Sun X, Tang K (2004) Rapid Isololation of high quality total RNA from taxus and ginkgo. Preparative Biochemestry and Biotecnology 34: 209-214
Liu, JJ, Goh CJ, Loh CS, Liu P, Pua EC (1998) A method for isolation of total RNA from fruit tissues of banana. Plant Molecular Biology Reporter 16: 1-6
Logan, DA, Mukhtarb M, Parveenc Z (1998) Isolation of RNA from the filamentous fungus Mucor circinelloides. Journal of Microbiological Methods 33: 115-118
Loomis, WD (1974) Overcoming problems of phenolics and quinones in the isolation of plant enzymes and organelles. Methods in Enzymology 31: 528-545
Malnoy, M, Reynoird JP, Mourgues F, Cheverau E, Simoneau P (2001) A method for isolation total RNA from pear. Molecular Biology Reporter 19: 69-74
Manickavelu, A, Kambara K, Mishina K, Koba T (2007) An efficient method for purifying high quality RNA from wheat pistils. Colloids Surf B Biointerfaces 54: 254-258
Meisel, L, Fonseca B, Gonzalez S, Baezayates R, Cambiazo V, Campos R, Gonzalez M, Orelana
A, Retamales J, Silva H (2005) A rapid and efficient method for purifying high quality total RNA from peaches (prunus persica) for functional genomics analyses. Biological Research 38: 83-88
Mendoza-Rodríguez, M, Sánchez-Rodríguez A, Acosta-Suárez M, Roque B, Portal O, Jiménez E (2006) Construcción y secuenciación parcial de una biblioteca sustractiva en ‘Calcutta 4’ (Musa AA) en estadio temprano de infección con Mycosphaerella fijiensis Morelet. Biotecnología Vegetal 6: 213-117
Moriguchi, T, Hu CG, Honda C, Kita M, Tsuda T (2002) A simple protocol for RNA isolation from fruit trees containing high levels of polisaccharides and polyphenol compounds. Plant Molecular Biology Reporter 20: 69a-69g
NEED (2008) New European Interprice in Distribution. En: http://www.need.es/ list.aspx?c=122&hc=&md=2 (consulta: mayo 2009)
Portillo, M, Fenoll C, Escobar C (2006) Evaluation of different RNA extraction methods for small quantities of plant tissue. Physiologia Plantarum 128: 1-7
Rodríguez-García, CM, Islas-Flores I, Peraza-Echeverría L y Canto-Canché B (2006) Extraction of high-quality, melanin-free RNA from M. fijiensis for cDNA preparation. Molecular Biotechnology 34: 45-50
Rogers, SO y Bendish AJ (1988) Extraction of DNA from plant tissues. En: Gelvin, SB y Schilperoort RA (Eds) Plant molecular biology manual, pp. 1-10. Kluwer Academic Publishers, Dordrecht
Salzman, RA, Fujita T, Zhu-Salzman K, Hasegawa PM y Bressan RA (1999) An improved RNA isolation method for plant tissues containing high levels of phenolic compounds or carbohydrates. Plant Molecular Biology Reporter 17: 11-17
Sánchez-Rodríguez, A, Portal O, Rojas LE, Ocaña
B, Mendoza M, Acosta M, Jiménez E y Höfte M (2008) An efficient method for the extraction of high-quality fungal total RNA to study the Mycosphaerella fijiensis – Musa spp. interaction. Molecular Biotechnology 40: 299-305
Satyamoorthy, K, Van Belle LG, Elder PA y Herlyn M (2002) A versatile method for the removal of melanin from ribonucleic acids in melanocytic cells. Melanoma Research 12: 449-452
Biotecnología Vegetal Vol. 11, No. 4, 2011
Soanes, DM. Skinner W, Keon J, Hargreaves J y Talbot NJ (2002) Genomics of phytopathogenic fungi and the development of bioinformatic resources. Molecular Plant-Microbe Interactions 15(5): 421-427.
Sokolovsky, V, Kaldenhoff R, Ricci M y Russo VEA (1990) Fast and reliable mini-prep RNA extraction from Neurospora crassa. En: www.fgsc.net/fgn37/ sokol.html (consulta: febrero 2008).
Somma, M (2008) DNA extraction and purification, session 4th. En: The analysis of food samples for the presence of genetically modified organisms, pp. 1-18. Joint Research Centre, European Commission
Stergiopoulos, I, Lute-Harm Z y De Waard MA (2003) The ABC transporter MgAtr4 is a virulence factor of Mycosphaerella graminicola that affects colonization of substomatal cavities in wheat leaves. Molecular Plant Microbe Interactions 16: 689–698
Van den Burg, HA, Westerink N, Francoijs KJ, Roth R y Woestenenk E (2003) Natural disulfide bond-disrupted mutants of AVR4 of the tomato pathogen Cladosporium fulvum are sensitive to proteolysis, circumvent Cf-4-mediated resistance, but retain their chitin binding ability. Journal of Biological Chemistry 278: 27340-46
Velazco, R (2005) Marcadores moleculares y la extracción de ADN. Revista Biotecnología en el Sector Agropecuario y Agroindustrial 3: 14-18
Venkatachalam, P, Thanseem I y Thulaseedharan A (1999) A rapid and efficient method for isolation of RNA from bark tissues of Hevea brasiliensis. Current Science 77: 635-637
Wang, X, Tian W y Li Y (2008) Development of an efficient protocol of RNA isolation from recalcitrant tree tissues. Molecular Biotechnology 38: 57-64
Yamashita, Y, Sakurai T, Kuno N, Uchida K y Yokobayshi T (2005) RNA extraction reagent, and method for analyzing biological materials. Dickstein Shapiro Llp, Ac12q168fi. Washington DC, USA
Yang, J , Li CY, Li H, Liu L, Su Y, Li JB, Chang Q, Qu LJ, Wang YY y Zhu YY (2008) A Novel Gene Involved in Lesion Formation in Magnaporthe grisea. Journal of Phytopathology 156: 99-103
Zahiri, AR, Babu MR y Saville BJ (2005) Differential gene expression during teliospore germination in Ustilago maydis. Molecular Genetics and Genomics 273: 394-403
Copyright (c) 2016 Biotecnología Vegetal
Biotecnología Vegetal eISSN 2074-8647, RNPS: 2154. ISSN 1609-1841, RNPS: 0397 Editada por: Instituto de Biotecnología de las Plantas. Universidad Central Marta Abreu de Las Villas. Carretera a Camajuaní km 5.5, Santa Clara, Villa Clara, Cuba CP 54 830 Tel: 53 42200124, e-mail: info@ibp.co.cu
Biotecnología Vegetal está bajo una Licencia Creative Commons Atribución-NoComercial 4.0 Internacional.