Correct heteroduplex formation for mutation detection analysis.
BACKGROUND
The majority of mutation detection methods for unknown mutations are polymerase chain reaction (PCR)-based methods dependent on the formation of heteroduplexes between wild-type and mutant strands of DNA.


RESULTS
This report discusses the difficulties associated with forming heteroduplexes with a large DNA fragment and the implications for subsequent mutation detection by the chemical cleavage of mismatch technique and other methods reliant on heteroduplex formation. It was found that the size and sequence context of the fragment being investigated inhibited correct heteroduplex formation. The problem was overcome by dividing the sequence into two overlapping fragments.


CONCLUSIONS
Early identification of this problem in other fragments will help with the rapid optimization of PCR-based mutation detection methods.
Smith
M J
MJ
Department of Pathology, The University of Melbourne, Melbourne.
Humphrey
K E
KE
Cappai
R
R
Beyreuther
K
K
Masters
C L
CL
Cotton
R G
RG
D020137:Base Pair Mismatch / Q000235:genetics; D004252:DNA Mutational Analysis / Q000379:methods*; D020180:Heteroduplex Analysis / Q000379:methods*; D006801:Humans; D008565:Membrane Proteins / Q000032:analysis / Q000235:genetics; D053764:Presenilin-1; D011786:Quality Control; D012015:Reference Standards
D008565:Membrane Proteins; C508853:PSEN1 protein, human; D053764:Presenilin-1