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Biochemistry 28 , — Zengel J. Nucleic Acids Res. Coburn G. Petersen C.
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Life Sciences Cell Biology. Methods in Molecular Biology Free Preview. Buy eBook. Buy Hardcover. Buy Softcover. All RNA samples were intact as judged by the sharp and distinct cytosolic and plastid ribosomal bands on the agarose gel. Moreover, agarose gel electrophoresis showed a distinct individual band of intact genomic DNA as well as reliable restriction enzyme digestion patterns. The absence of smear on the gel confirms the spectrophotometric results, and provides further evidence that this protocol efficiently removed contaminants during DNA and RNA isolation from the different strains of E.
Similarly, the genomic DNA, free of interfering compounds, was efficiently used for PCR and therefore would be suitable for DNA sequencing, southern blot hybridization and whole genome methylation sequencing. Interestingly, a method  previously used to isolate RNA from a specific strain of E. Furthermore, although the effectiveness of the recently published protocol by Coelho et al. Despite the problematic metabolites present in the cell and associated with the cell wall, the DNA and RNA extracted were of excellent quality and applicable for downstream applications.
Together with the spectrophotometric and electrophoretic analyses these results provide evidence that the method successfully dealt with these interfering components. Moreover, by using this protocol it is possible to obtain high yields of nucleic acids from small quantities of biomass, and both yield and purity are strain-independent.
We further suggest that the protocol may have wider applicability to other algal species that have polyphenol- and polysaccharide-rich tissues. Summary of nucleic acids extraction from Ectocarpus siliculosus. Nucleic acids precipitation.
At this step it is possible to precipitate the nucleic acids by splitting the aqueous phase of one sample in multiple tubes usually two , and in a second step join the precipitated nucleic acids. The nucleic acids of one sample are combined in single tube. Nanodrop spectrophotometry measurements of REP Total RNA extracted from REP10—11, measured after DNase treatment and a purification step, are of high quality and free from appreciable levels of organic contaminants regardless of the biomass used in the extraction procedures. A 25 mg B 50 mg and C mg of starting biomass, respectively.
M: bp ladder.
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Strains collected from pristine sites exhibit a higher quantity of nucleic acids extracted compared to those from polluted sites. A differential decrease in the quantity of nucleic acids was recorded for all strains when the old method  was used compared with the new one. List of consumables, solutions and reagents, equipment as well as a guideline of nucleic acids extraction.
Saez for doctoral studies. The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript. National Center for Biotechnology Information , U. PLoS One. Published online May Brown , 2 and Maria Beatrice Bitonti 1. Claudio A. Murray T. Hoheisel, Editor. Author information Article notes Copyright and License information Disclaimer. Competing Interests: The authors have declared that no competing interests exist.
Received Nov 2; Accepted Apr 9. This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are properly credited. This article has been corrected.
See PLoS One. This article has been cited by other articles in PMC. Figure S2: Nucleic acids precipitation. Figure S3: The nucleic acids of one sample are combined in single tube. Table S2: Reagent used to remove contaminants. File S1: List of consumables, solutions and reagents, equipment as well as a guideline of nucleic acids extraction.
Abstract The brown seaweed Ectocarpus siliculosus is an emerging model species distributed worldwide in temperate coastal ecosystems. Introduction Brown algae are an ecologically and economically important group of marine photoautotrophs  —  that first appeared million years ago and evolved multicellularity independently of green and red algae and higher plants  , .
Open in a separate window. Figure 1. Summary of nucleic acids extraction from E. Repeat steps 4 and 5.
Table 1 Split and precipitate the aqueous phase of one sample in more tubes usually two. Table 2 Reagent used in the precipitation step. Aqueous Phase top layer e. Table 3 The nucleic acid of one sample precipitated in two different tubes is transferred in one tube after the resuspension in the appropriate volume of nuclease-free water. Add 0. Results 2. Figure 2.
Analysis of quality and integrity of extracted nucleic acids. Figure 3.