A SIMPLE AND EFFICIENT METHOD FOR HIGH-QUALITY TOTAL RNA ISOLATION FROM OLEAGINOUS MICROALGAE
JUAN C. CASTRO *
Unidad Especializada de Biotecnología, Centro de Investigaciones de Recursos Naturales de la Amazonía (CIRNA), Universidad Nacional de la Amazonía Peruana (UNAP), Psje. Los Paujiles S/N, Iquitos, Postal Code: 16024, Perú
HICLER N. RODRÍGUEZ
Unidad Especializada de Biotecnología, Centro de Investigaciones de Recursos Naturales de la Amazonía (CIRNA), Universidad Nacional de la Amazonía Peruana (UNAP), Psje. Los Paujiles S/N, Iquitos, Postal Code: 16024, Perú
J. DYLAN MADDOX
The Field Museum of Natural History, Pritzker Laboratory for Molecular Systematics and Evolution, 1400 S. Lake Shore Drive, Chicago, IL, Postal Code: 60605, USA and Environmental Sciences, American Public University System, Charles Town, WV, Postal Code: 25414, USA
BRUCE JIU
Laboratorio de Biotecnología y Bioenergética, Universidad Científica del Perú (UCP), Av. Abelardo Quiñones Km 2.5, Iquitos, Postal Code: 6024, Perú
JOE B. PETTERMAN
Laboratorio de Biotecnología y Bioenergética, Universidad Científica del Perú (UCP), Av. Abelardo Quiñones Km 2.5, Iquitos, Postal Code: 6024, Perú
JORGE L. MARAPARA
Unidad Especializada de Biotecnología, Centro de Investigaciones de Recursos Naturales de la Amazonía (CIRNA), Universidad Nacional de la Amazonía Peruana (UNAP), Psje. Los Paujiles S/N, Iquitos, Postal Code: 16024, Perú
MARIANELA COBOS *
Laboratorio de Biotecnología y Bioenergética, Universidad Científica del Perú (UCP), Av. Abelardo Quiñones Km 2.5, Iquitos, Postal Code: 6024, Perú
*Author to whom correspondence should be addressed.
Abstract
Microalgae are a very diverse group of microorganisms with great potential for biotechnological applications. To best exploit these organisms is necessary to expand our knowledge at its molecular levels. To realize these studies, however, it is essential to have efficient and reproducible methods for total RNA isolation. The objective of this research was to develop a method for high-quality total RNA isolation from three oleaginous microalgae species promissories for biotechnological applications. Three strains Ankistrodesmus sp., Chlorella sp., and Scenedesmus sp. were cultured on CHU-10 medium. Then, were harvested by centrifugation and total RNA was isolated with three commercial kits, one previously published method, and our improved CTAB-LiCl method. Quality and quantity of total RNA isolated were assayed with standard spectrophotometric and electrophoretic techniques. The highest quality (A260/A280: 1.95-2.02, A260/A230: 1.12-1.84) and yield (16.40-93.63 mg/g fresh weight) of total RNA from the three microalgae strains were obtained with our improved CTAB-LiCl method. In addition, the time required to complete the isolation process and the cost were relatively lowest. In conclusion, the improved CTAB-LiCl method, in contrast to frequently used commercial kits, and a previously published method, is effective and reliable to produce total RNA of high quality and quantity that is suitable for cDNA synthesis and polymerase chain reaction.
Keywords: Ankistrodesmus, Chlorella, oleaginous microalgae, RNA isolation, Scenedesmus