The Jacks Lab

 

protocols

Nf2 PCR

Reaction Mix (store at -20C):

100 pmol stock NF2WT 20 ul
100 pmol stock NF2X2 20 ul
100 pmol stock neo18 20 ul
10 X buffer (see below) 500 ul
milliQ H2O 4430 ul

For each reaction

Tail DNA 2 ul
RX Mix 17 ul
Taq (1/25) 1.0ul

PCR Primers:

Nf2WT 5' GTC TGG GAA GTC TGT GGA GG 3'
Nf2X2 5’ GGG GCT TCG GGA AAC CTG G 3’
Neo18 5’ CTA TCA GGA CAT AGC GTT GG 3’

PCR Conditions:

1. 94C 1:00
2. 94C 0:30
3. 61C 1:45
4. 72C 1:45
5. Go to 2 x29 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:

WT = 306 bp
Mut = 574 bp