Rapid and effective DNA extraction method with bead grinding for a large amount of fungal DNA.
To identify a rapid method for extracting a large amount of DNA from fungi associated with food hygiene, extraction methods were compared using fungal pellets formed rapidly in liquid media. Combinations of physical and chemical methods or commercial kits were evaluated with 3 species of yeast, 10 species of ascomycetous molds, and 4 species of zygomycetous molds. Bead grinding was the physical method, followed by chemical methods involving sodium dodecyl sulfate (SDS), cetyl trimethyl ammonium bromide (CTAB), and benzyl chloride and two commercial kits. Quantity was calculated by UV absorbance at 260 nm, quality was determined by the ratio of UV absorbance at 260 and 280 nm, and gene amplifications and electrophoresis profiles of whole genomes were analyzed. Bead grinding with the SDS method was the most effective for DNA extraction for yeasts and ascomycetous molds, and bead grinding with the CTAB method was most effective with zygomycetous molds. For both groups of molds, bead grinding with the CTAB method was the best approach for DNA extraction. Because this combination also is relatively effective for yeasts, it can be used to extract a large amount of DNA from a wide range of fungi. The DNA extraction methods are useful for developing gene indexes to identify fungi with molecular techniques, such as DNA fingerprinting.
Division of Microbiology, National Institute of Health Sciences, Kamiyoga 1-18-1, Setagaya-ku, Tokyo 158-8501, Japan. mwatanabe@nihs.go.jp
M
M
Watanabe
K
K
Lee
K
K
Goto
S
S
Kumagai
Y
Y
Sugita-Konishi
Y
Y
Hara-Kudo
D015169:Colony Count, Microbial / Q000379:methods*; D003257:Consumer Product Safety; D004271:DNA, Fungal / Q000032:analysis / Q000302:isolation & purification*; D005506:Food Contamination / Q000032:analysis; D005516:Food Microbiology; D005658:Fungi / Q000235:genetics / Q000302:isolation & purification*; D005784:Gene Amplification; D006801:Humans; D013997:Time Factors; D015003:Yeasts / Q000235:genetics / Q000302:isolation & purification*
D004271:DNA, Fungal