A SIMPLE AND RAPID TECHNIQUE FOR THE ISOLATION OF DNA FROM MICROALGAE¹

Abstract

A simple method for the purification of PCRquality DNA from microalgae is presented. This method uses the detergent dodecyltrimethylammonium bromide coupled with cell breakage by agitation in the presence of glass beads and chloroform. A final purification step involves a commercial cartridge system. The procedure requires only about 1-2 mL of algal culture and can be completed in about 20 min. DNA suitable for PCR has been obtained from several algal lineages using this method, including numerous green algae and stramenopiles.

A taxonomic study ofEudorina unicocca(Volvocaceae, Chlorophyceae) and related species, based on morphology and molecular phylogeny

“…The protocol of Fawley & Fawley (2004) was used to prepare the total DNA of seven strains (Table 1), with some modifications (see Nakada & Nozaki, 2007). The method for sequencing the chloroplast gene (rbcL) for the large subunit of RuBisCO of two strains of Y. unicocca (Hasu-1 and NIES-870), E. unicocca UTEX 1221 and two aplanosporic strains (TKI-C-2 and 990601-IE-5) was essentially identical to that described previously (Nozaki et al, 1995(Nozaki et al, , 1997(Nozaki et al, , 2000(Nozaki et al, , 2002.…”

New Design Strategy for Development of Specific Primer Sets for PCR-Based Detection ofChlorophyceaeandBacillariophyceaein Environmental Samples

“…Finally, the specificity of the primers was investigated using standard PCR amplification performed on a variety of algal and nonalgal cultured strains (Table 2). DNA extraction was slightly modified from the method of Fawley and Fawley (6). Universal eukaryotic 18S rDNA primers were used to test whether the extracted DNA was amplifiable (12).…”

Toxigenic algae and associated phycotoxins in two coastal embayments in the Ebro Delta (NW Mediterranean)

“…by light microscopy and presence of the toxin domoic acid (DA). These samples were then processed (following Andree et al, 2011;Fawley and Fawley, 2004) by targeting the species Pseudo-nitzschia arenysensis (=Pseudo-nitzschia delicatissima, strain Ra3), Pseudo-nitzschia brasiliana, Pseudo-nitzschia calliantha, Pseudo-nitzschia delicatissima (strain Ra2), Pseudo-nitzschia fraudulenta, Pseudo-nitzschia galaxiae, Pseudo-nitzschia multistriata, and Pseudo-nitzschia pungens. For DNA extraction, cell pellets were thawed and suspended in 200 mL lysis buffer (1 M NaCl, 70 mM Tris, 30 mM EDTA, pH 8.0), then each was transferred to a 2 mL cryo-tube containing approximately 50 mg of 0.5 mm-diameter zirconium glass beads (BioSpec, Bartlesville, Oklahoma, USA).…”

“…Then 100 mL of aqueous supernatant was transferred to a new tube. Extraction of the genomic DNA continued from this supernatant using a GeneClean Kit (MP Biomedicals, LLC, LLC, Santa Ana, California, USA), following the procedure of Fawley and Fawley (2004). A hemispecific assay, containing one genus-specific 5.8S primer and one species-specific ITS-1 or ITS-2 primer, was used for identification of taxa (Andree et al, 2011).…”