Open AccessMolecular approach to the characterisation of fungal communities: methods for DNA extraction, PCR amplification and DGGE analysis of painted art objects
Author(s) -
Möhlenhoff Petra,
Müller Lars,
Gorbushina Anna A,
Petersen Karin
Publication year - 2001
Publication title -
fems microbiology letters
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.899
H-Index - 151
eISSN - 1574-6968
pISSN - 0378-1097
DOI - 10.1111/j.1574-6968.2001.tb10516.x
Subject(s) - temperature gradient gel electrophoresis , polymerase chain reaction , dna extraction , biology , dna , primer (cosmetics) , ribosomal dna , 16s ribosomal rna , gel electrophoresis , extraction (chemistry) , microbiology and biotechnology , bacteria , chromatography , chemistry , gene , genetics , phylogenetics , organic chemistry
A protocol for efficient extraction of fungal DNA from micromycetes colonising painted art objects was developed. Polymerase chain reaction (PCR) inhibitors were successfully removed by a combined application of a Chelex‐100 adsorption resin and a Geneclean Kit for Ancient DNA. Universal fungal primers for PCR amplification of 28S rDNA (U1 and U2) were tested for their applicability in denaturing gradient gel electrophoresis (DGGE) analysis of fungal communities. Artificially produced mortar samples inoculated with fungal pure cultures isolated from mural paintings were used as model objects for DNA extractions and DGGE analysis. Good resolution in DGGE was achieved using 260‐bp rDNA fragments amplified with U1/DGGE and U2 primers directly from model communities.