Culture independent PCR: an alternative enzyme discovery strategy.
Degenerate primers were designed for use in a culture-independent PCR screening of DNA from composite fungal communities, inhabiting residues of corn stovers and leaves. According to similarity searches and alignments amplified clone sequences affiliated with glycosyl hydrolase family 7 and glycosyl hydrolase family 45 though significant sequence divergence was observed. Glycosyl hydrolases from families 7 and 45 play a crucial role in biomass conversion to fuel ethanol. Research in this renewable energy source has two objectives: (i) To contribute to development of a renewable alternative to world's limited crude fossil oil reserves and (ii) to reduce air pollution. Amplification with 18S rDNA-specific primers revealed species within the ascomycetous orders Sordariales and Hypocreales as well as basidiomycetous order Agaricales to be present in these communities. Our study documents the value of culture-independent PCR in microbial diversity studies and could add to development of a new enzyme screening technology.
Jacobsen
Jonas
J
Microbial Discovery, Novozymes A/S, Smørmosevej 25 1B1, DK-2880, Denmark.
Lydolph
Magnus
M
Lange
Lene
L
D000595:Amino Acid Sequence; D001203:Ascomycota / Q000201:enzymology* / Q000235:genetics; D001483:Base Sequence; D001487:Basidiomycota / Q000201:enzymology* / Q000235:genetics; D002480:Cellulase / Q000235:genetics / Q000302:isolation & purification*; D043366:Cellulose 1,4-beta-Cellobiosidase / Q000235:genetics / Q000302:isolation & purification*; D003001:Cloning, Molecular; D004271:DNA, Fungal / Q000737:chemistry / Q000235:genetics; D008969:Molecular Sequence Data; D010802:Phylogeny; D018515:Plant Leaves / Q000382:microbiology; D016133:Polymerase Chain Reaction / Q000379:methods*; D016415:Sequence Alignment; D003313:Zea mays / Q000382:microbiology
D004271:DNA, Fungal; D002480:Cellulase; D043366:Cellulose 1,4-beta-Cellobiosidase