Secondary metabolite production in filamentous fungi displayed.
We report the potential of differential display technology for the isolation of genes of biotechnological interest. We have assessed the usefulness of the technique for the cloning of genes involved in the production of secondary metabolites, many of which are of industrial use or interest. We have used the complex pathway for the biosynthesis of gibberellins, as well as bikaverin and carotenoids, present in the filamentous fungus Gibberella fujikuroi as a test system. From a total display of approximately 16000 PCR products for each RNA sample, 100 were derived from the derepressed but not the repressed condition. These products were analysed by Northern blotting and a subset of 16 such PCR products showed differential expression at the transcript level. A number of different mRNA species were observed on this basis which varied in their size. Hence, this approach appears suitable for the isolation of genes involved in the complex pathways often required for the synthesis of secondary metabolites in organisms which are genetically intractable. Moreover, the method has the advantage that it is quick, differential displays being obtained after 2 days and DNA clones in 6 days.
Appleyard
V C
VC
Plant Sciences Laboratory, School of Biological and Medical Sciences, University of St. Andrews, Scotland, UK.
Unkles
S E
SE
Legg
M
M
Kinghorn
J R
JR
D015152:Blotting, Northern; D018076:DNA, Complementary; D005285:Fermentation; D005800:Genes, Fungal; D005874:Gibberella / Q000235:genetics* / Q000254:growth & development / Q000378:metabolism*; D005875:Gibberellins / Q000032:analysis / Q000096:biosynthesis*; D007700:Kinetics; D016133:Polymerase Chain Reaction; D012331:RNA, Fungal / Q000096:biosynthesis / Q000302:isolation & purification
D018076:DNA, Complementary; D005875:Gibberellins; D012331:RNA, Fungal