A modified CTAB method for the extraction of high-quality RNA from mono-and dicotyledonous plants rich in secondary metabolites.
| Autor: | Kiss T; Food and Wine Research Institute, Eszterházy Károly Catholic University, Eger, Hungary. kiss2.tibor@uni-eszterhazy.hu., Karácsony Z; Food and Wine Research Institute, Eszterházy Károly Catholic University, Eger, Hungary., Gomba-Tóth A; Food and Wine Research Institute, Eszterházy Károly Catholic University, Eger, Hungary., Szabadi KL; HUN-REN Centre for Ecological Research, Institute of Ecology and Botany, Vácrátót, Hungary.; Doctoral School of Biological Sciences, Hungarian University of Agriculture and Life Sciences, Gödöllő, Hungary., Spitzmüller Z; Food and Wine Research Institute, Eszterházy Károly Catholic University, Eger, Hungary., Hegyi-Kaló J; Food and Wine Research Institute, Eszterházy Károly Catholic University, Eger, Hungary., Cels T; Food and Wine Research Institute, Eszterházy Károly Catholic University, Eger, Hungary., Otto M; Food and Wine Research Institute, Eszterházy Károly Catholic University, Eger, Hungary., Golen R; Food and Wine Research Institute, Eszterházy Károly Catholic University, Eger, Hungary., Hegyi ÁI; Food and Wine Research Institute, Eszterházy Károly Catholic University, Eger, Hungary.; Doctoral School of Environmental Sciences, Hungarian University of Agriculture and Life Sciences, Gödöllő, Hungary., Geml J; HUN-REN-EKKE Lendület Environmental Microbiome Research Group, Hungarian Research Network and Eszterházy Károly Catholic University, Eger, Hungary., Váczy KZ; Food and Wine Research Institute, Eszterházy Károly Catholic University, Eger, Hungary. |
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| Jazyk: | angličtina |
| Zdroj: | Plant methods [Plant Methods] 2024 May 04; Vol. 20 (1), pp. 62. Date of Electronic Publication: 2024 May 04. |
| DOI: | 10.1186/s13007-024-01198-z |
| Abstrakt: | Background: High-quality RNA extraction from woody plants is difficult because of the presence of polysaccharides and polyphenolics that bind or co-precipitate with the RNA. The CTAB (cetyl trimethylammonium bromide) based method is widely used for the isolation of nucleic acids from polysaccharide-rich plants. Despite the widespread use of the CTAB method, it is necessary to adapt it to particular plant species, tissues and organs. Here we described a simple and generalized method for RNA isolation from mature leaf tissues of several economically important woody (17) and herbaceous plants (2) rich in secondary metabolites. High yields were achieved from small amount (up to 50 mg) of plant material. Two main modifications were applied to the basic protocol: an increase in β-mercaptoethanol concentration (to 10%v/v) and the use of an effective DNase treatment. As opposed to similar studies, we tried to describe a more detailed protocol for isolating RNA, including the exact quantity and concentration of the reagents were used. Results: Our modified CTAB method is proved to be efficient in extracting the total RNA from a broad range of woody and herbaceous species. The RNA yield was ranged from 2.37 to 91.33 µg/µl. The A Conclusions: Our RNA isolation method, with fine-tuned and detailed instructions, can produce high quality RNA from a small amount of starting plant material that is suitable for use in downstream transcriptional analyses. The use of an increased concentration of the reducing agent β-mercaptoethanol in the extraction buffer, as well as the application of DNaseI-treatment resulted in a method suitable for a wide range of plants without the need of further optimalization, especially in Rhus typhina (Staghorn sumac), for which molecular-genetic studies have not yet been sufficiently explored. (© 2024. The Author(s).) |
| Databáze: | MEDLINE |
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