This protocol is used to distinguish between KRas LA1 and KRas LA2 as the PCR protocol cannot.
5'internal xba/sal probe
Digest with Hind 111(#798 983) and Asp 718(#1175 050) both from Boehringer Mannheim.
Wash 3 times at 68C (max) each for 20-30mins in
2ml 0.5M EDTA
50ml 20% SDS
5' Mut 8.9kb
Activation of Kras in lungs should be shown by loss of intensity of the 5' mutant bands.
WT DNA (10kb only)
Lat 44 DNA (10kb, 8.9kb, and 6.1kb bands)
Act DNA (10kb and 8.9kb bands, the recombination was allowed to occur in vitro)