The Jacks Lab

 

protocols

p53 R172H and R270H recombined point mutant alleles (germline)

and

Ser23Ala point mutant allele

Reaction Mix:

Tail DNA 1.0 ÁL
10x buffer 2.0ul
DT020200.1 (40uM) 0.5ÁL
DT011200.3 (40uM) 0.5ÁL
dH20 15 ÁL
Taq (1/25) 1.0ul

PCR Primers:

DT020200.1 5' AGC CTG CCT AGC TTC CTC AGG 3'
DT011200.3 5' CTT GGA GAC ATA GCC ACA CTG 3'

PCR Conditions:

1. 94C 3:00
2. 94C 1:00
3. 60C 2:00
4. 72C 2:00
5. Goto 2 x30 cycles
6. 72C 5:00
7. 4C 0:00
END

10X PCR Buffer:

500 mM KCl
100mM Tris pH 8.3
15 mM MgCl2
1 mg/ml BSA
2 mM dNTPs
dH20

(store at -20C)

Expected Bands:

Wild Type = ~290 bp
1 Lox = ~330 bp