National Center of Excellence in Molecular Biology (CEMB)
Abstract
Background: RNA extraction from tree species like Eucalyptus is very tedious and difficult task. In this study a very short and efficient method of RNA extraction from Eucalyptus has been described.Methods: A very short and efficient protocol of two steps RNA extraction has been optimized for obtaining high quality and pure RNA from different tissue types of Eucalyptus tree. In first step whole nucleic acid was extracted from plant tissues and in second step RNA was purified from nucleic acid mixture. The newly optimized rapid CTAB RNA extraction method was compared with trizol extraction method for efficiency and quality of extracted RNA.Results: The newly optimized rapid CTAB RNA extraction method was found highly efficient and suitable over the trizol method. Three Eucalyptus dehydrin genes the dehydrin-10 (DHN-10), dehydrin-1 (DHN-1), and dehydrin-2 (DHN-2) were successfully amplified, TA cloned into pTZ57/RT vector, and transformed into Top10F’ strain of E.coli. These three Eucalyptus dehydrin genes have been reported for conferring abiotic stress tolerance to the Eucalyptus plant yet have not been reported to be cloned. These cloned genes would be further manipulated for developing abiotic stress tolerance in plants of interest.Conclusion: Rapid CTAB RNA extraction method is a brief and reproducible methodology for a hard job of RNA extraction from tree plants with high phenolic contents. Keywords: Dehydrin proteins, Eucalyptus abiotic stress tolerance proteins, cloning of dehydrin genes, DHN-1, DHN-2, DHN-1
