Simple and rapid RNA extraction from freeze-dried tissue of brown algae and seagrasses
Abstract:An inexpensive and rapid RNA extraction protocol for brown algae and seagrasses is presented, based on homogenization in a simple CTAB buffer and selective precipitation of RNA with lithium chloride. The protocol avoids the use of toxic chaotropic agents and phenol; high concentrations of dithiothreitol are used to inhibit RNase activity and to prevent oxidative cross-linking of nucleic acids by phenolics. A relatively high throughput of about 100 samples in 24?h can be achieved for, for example, Northern analysis. Yields of 50–200?µg?g -1 fresh weight are comparable with those obtained for higher plants using commercially available kits. Tests of the extraction procedure demonstrate that high quality, intact RNA can be obtained from a variety of lyophilized brown algal tissues, even after prolonged storage at room temperature. Lyophilization is therefore suggested as an alternative to freezing tissue at -70°C to -80°C. The RNA obtained was used directly in several downstream applications to detect Fucus plastid-encoded transcripts by RNA-labelling, RT-PCR and Northern analysis.
Document Type: Research Article
Affiliations: CCMAR, CIMAR/Laboratório Associado, F.C.M.A., Universidade do Algarve, Gambelas, 8005–139 Faro, Portugal
Publication date: 2006-02-01