Comparison of RNA extraction methods from biofilm samples of <it>Staphylococcus epidermidis</it>

• Main Authors: França Angela, Melo Luís DR, Cerca Nuno
• Format: Article
• Language: English
• Published: BMC 2011-12-01
• Series: BMC Research Notes
• Online Access: http://www.biomedcentral.com/1756-0500/4/572

<p>Abstract</p> <p>Background</p> <p>Microbial biofilms are communities of bacteria adhered to a surface and surrounded by an extracellular polymeric matrix. Biofilms have been associated with increased antibiotic resistance and tolerance to the immune system. <it>Staphylococcus epidermidis </it>is the major bacterial species found in biofilm-related infections on indwelling medical devices. Obtaining high quality mRNA from biofilms is crucial to validate the transcriptional measurements associated with the switching to the biofilm mode of growth. Therefore, we selected three commercially available RNA extraction kits with distinct characteristics, including those using silica membrane or organic extraction methods, and enzymatic or mechanical cell lysis, and evaluated the RNA quality obtained from two distinct <it>S. epidermidis </it>bacterial biofilms.</p> <p>Results</p> <p>RNA extracted using the different kits was evaluated for quantity, purity, integrity, and functionally. All kits were able to extract intact and functional total RNA from the biofilms generated from each <it>S. epidermidis </it>strain. The results demonstrated that the kit based on mechanical lysis and organic extraction (FastRNA^®Pro Blue) was the only one that was able to isolate pure and large quantities of RNA. Normalized expression of the <it>icaA </it>virulence gene showed that RNA extracted with PureLink™ had a significant lower concentration of <it>icaA </it>mRNA transcripts than the other kits tested.</p> <p>Conclusions</p> <p>When working with complex samples, such as biofilms, that contain a high content extracellular polysaccharide and proteins, special care should be taken when selecting the appropriate RNA extraction system, in order to obtain accurate, reproducible, and biologically significant results. Among the RNA extraction kits tested, FastRNA^®Pro Blue was the best option for both <it>S. epidermidis </it>biofilms used.</p>