Tempel Lab Protocols

FGFR3 GENOTYPE

You will Need

Mouse Tail Digests

3 Primers:
JC 14
JC 18
JC 41

PCR Reagents:
Sterile water
10x buffer w/MgCl2
10mM Nucleotide mix
FastStart taq
Primer Sequences

JC 14 GGG CTC CTT ATT GGA CTC GC
JC 18 TGC TAA AGC GCA TGC TCC AGA CTG C
JC 41 AGG TAT AGT TGC CAC GAT CGG AGG G

Procedure
The PCR reaction can be run with all three primers together.

PCR Rxn
Sterile Water

37.6 ul
10x PCR buffer w/MgCl2 5ul
10mM Nucleotide mix 1 ul
JC 14 primer (20 uM) 2 ul
JC 18 primer (20 uM) 1 ul
JC 41 primer (20 uM) 1 ul
FastStart TAQ 0.4ul
Tail Lysate 2 ul
Total Volume 50 ul
PCR Protocol

94 deg C 4:30
94 deg C
 0:30
55 deg C
 1:00 x30
72 deg C
 3:00
72 deg C 7:00
4 deg C Hold

Gel
2% Agarose + 3 ul EtBr (10mg/ml)

Bands
WildType: 322 bp
Mutant: 200 bp