Quantitative transcriptional analysis of the genes encoding glycoside hydrolase family 7 cellulase isozymes in the basidiomycete Phanerochaete chrysosporium.
Cellulolytic fungi generally secrete a cellulase mixture consisting mainly of glycoside hydrolase family 7 cellulases (Cel7s) during degradation of crystalline cellulose. Although several Cel7s have been investigated so far, the marked similarity in their amino acid and nucleotide sequences makes independent quantitative analysis difficult. Here, we present a real-time PCR method for the detection and quantification of Cel7 genes (cel7A-F/G) in the basidiomycete Phanerochaete chrysosporium using PCR primer sets designed based on the 3' untranslated region sequences. It was confirmed by agarose gel electrophoresis, sequencing, and dissociation curve analysis of the PCR products that each cel7 transcript was specifically amplified by the corresponding primers. We applied this real-time reverse-transcription PCR method using the presented primer sets to evaluate quantitatively the expression changes of cel7 genes in P. chrysosporium under conditions of carbon catabolite derepression.
Suzuki
Hitoshi
H
Department of Biomaterials Sciences, Graduate School of Agricultural and Life Sciences, The University of Tokyo, 1-1-1 Yayoi, Tokyo, Japan.
Igarashi
Kiyohiko
K
Samejima
Masahiro
M
D002480:Cellulase / Q000096:biosynthesis* / Q000235:genetics; D017931:DNA Primers / Q000235:genetics; D020869:Gene Expression Profiling; D015966:Gene Expression Regulation, Fungal; D007527:Isoenzymes / Q000096:biosynthesis / Q000235:genetics; D020075:Phanerochaete / Q000201:enzymology* / Q000235:genetics; D020133:Reverse Transcriptase Polymerase Chain Reaction / Q000379:methods
D017931:DNA Primers; D007527:Isoenzymes; D002480:Cellulase