The use of biomagnetic separation to recover DNA suitable for PCR from Claviceps species.
DNA analysis of agriculturally important fungi using polymerase chain reaction (PCR)-based methods is becoming routine in research and for diagnostic purposes. Rapid, small-scale DNA isolation methods that take advantage of the sensitivity, speed and automation potential of PCR technology are needed for timely analysis of important plant pathogens. DNA isolated from Claviceps africana (causal agent of ergot of sorghum) using several standard DNA extraction protocols was found to be unamplifiable using PCR. The standard methods apparently failed to separate DNA from substances inhibitory to the Taq polymerase enzyme. We obtained DNA amenable to PCR analysis using a novel method involving magnetic beads and high salt extraction buffer. The biomagnetic purification method allowed us to obtain reliable PCR amplification of the internal transcribed spacer (ITS) regions of rDNA of Claviceps africana, making genetic comparisons possible.
Scott
D L
DL
Morehouse College, Department of Biology, Atlanta, GA, USA.
Clark
C W
CW
Tooley
P W
PW
Carras
M M
MM
Maas
J L
JL
D002967:Claviceps / Q000254:growth & development / Q000302:isolation & purification*; D004271:DNA, Fungal / Q000032:analysis / Q000302:isolation & purification*; D008280:Magnetics; D010935:Plant Diseases / Q000382:microbiology*; D016133:Polymerase Chain Reaction / Q000379:methods*
D004271:DNA, Fungal