Anne-Lise Brresen-Dale, Sigrid Lystad, and Anita Langerd, Department of Genetics, Institute for Cancer Research, The Norwegian Radium Hospital, N-0310 Oslo, Norway.
Temporal temperature gradient electrophoresis (TTGE) exploits the principles upon which denaturing gradient gel electrophoresis (DGGE) and constant denaturing gel electrophoresis (CDGE) are based. TTGE combines some of the advantages of these techniques, and eliminates some of the problems. The focusing of bands obtained by DGGE seems to be retained, and in TTGE there is no need for a chemical denaturing gradient which is the advantage in CDGE. The problem of determining the exact running time and denaturing conditions is eliminated, and since the denaturing conditions in TTGE span a wider range, several fragments with different melting behavior can be analyzed on the same gel. A new electrophoretic instrument from Bio-Rad Laboratories called the DCode universal mutation detection system has the ability to perform TTGE.
Improvements of DGGE by CDGE
DGGE, which is based on the discontinuous phenomenon of strand dissociation, allows the resolution of DNA fragments differing by as little as a single nucleotide substitution.1 In mutation analysis using parallel DGGE, field strength, temperature, and run time must be strictly controlled to achieve reproducible results. DGGE is complicated by the difficulties of choosing the exact running time and gel conditions to achieve the optimal separation. By running a DGGE gel too long, an achieved separation decreases, and may even be lost. Mutations present in a homo- or hemizygote state, where heteroduplexes are not formed, will be