Simple, Rapid and Cost-Effective Method for High Quality Nucleic Acids Extraction from Different Strains of Botryococcus braunii

Abstract

This study deals with an effective nucleic acids extraction method from various strains of Botryococcus braunii which possesses an extensive extracellular matrix. A method combining freeze/thaw and bead-beating with heterogeneous diameter of silica/zirconia beads was optimized to isolate DNA and RNA from microalgae, especially from B. braunii. Eukaryotic Microalgal Nucleic Acids Extraction (EMNE) method developed in this study showed at least 300 times higher DNA yield in all strains of B. braunii with high integrity and 50 times reduced working volume compared to commercially available DNA extraction kits. High quality RNA was also extracted using this method and more than two times the yield compared to existing methods. Real-time experiments confirmed the quality and quantity of the input DNA and RNA extracted using EMNE method. The method was also applied to other eukaryotic microalgae, such as diatoms, Chlamydomonas sp., Chlorella sp., and Scenedesmus sp. resulting in higher efficiencies. Cost-effectiveness analysis of DNA extraction by various methods revealed that EMNE method was superior to commercial kits and other reported methods by >15%. This method would immensely contribute to area of microalgal genomics.

A Novel Mechanism, Linked to Cell Density, Largely Controls Cell Division in Synechocystis

“…RNA was extracted according to Kim et al (2006), applying the freeze-thaw step described by Kim et al (2012) before disrupting the cells in a Tissuelyser (Qiagen). RNA concentration was quantified spectrophotometrically at 260 nm, and its integrity was checked by electrophoresis on a denaturing agarose gel (1.6% [m/v]; Goda and Minton, 1995).…”

Highly Efficient Transformation of the DiatomPhaeodactylum tricornutumby Multi-Pulse Electroporation

“…To examine the integration of the reporter genes in the diatom genome, genomic DNA was isolated from transformed cells that were subcultured 2-3 times on selective medium. The protocol described by Kim et al 17) was used to isolate genomic DNA. Primer pairs 0 -AGGTCCACATGGACTGGAAG-3 0 for actin were used in genomic PCR.…”

Growth characteristics and lipid content of three Korean isolates of Botryococcus braunii (Trebouxiophyceae)

“…All three strains grew best in BG-11 medium and worst in DY-III medium. Previous papers reported a critical effect of nitrate on the growth of B. braunii (Xu et al 2001, Zhila et al 2005a, 2005b, Choi et al 2011, Kim et al 2012. In fact, the concentration of nitrate was highest in BG-11, intermediate in Chu-13, and lowest in DY-III, in a ratio of 49:11:1.…”