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Mutant = 479 bp
Heterozygote = 322 bp and 479 bp
Wild type = 322 bp
Primer | 5' Label | Sequence 5' → 3' | 3' Label | Primer Type | Reaction | Note |
---|---|---|---|---|---|---|
oIMR0295 | TTG TAA ATC AGA AGG AAG TGG | Wild type Reverse | A | |||
oIMR0296 | AGA GTT ACT CTT GAT GTA GAT CTC C | Common | A, B | |||
oIMR2060 | CAC GAG ACT AGT GAG ACG TG | Mutant Reverse | B |
Component | Final Concentration |
---|---|
ddH2O | |
Kapa 2G HS buffer | 1.30 X |
MgCl2 | 2.60 mM |
dNTPS-kapa | 0.26 mM |
oIMR0295 | 0.50 uM |
oIMR0296 | 0.50 uM |
Glycerol | 6.50 % |
Dye | 1.00 X |
Kapa 2G HS taq polym | 0.03 U/ul |
DNA |
Step | Temp °C | Time | Note |
---|---|---|---|
1 | 94.0 | -- | |
2 | 94.0 | -- | |
3 | 65.0 | -- | -0.5 C per cycle decrease |
4 | 68.0 | -- | |
5 | -- | repeat steps 2-4 for 10 cycles (Touchdown) | |
6 | 94.0 | -- | |
7 | 60.0 | -- | |
8 | 72.0 | -- | |
9 | -- | repeat steps 6-8 for 28 cycles | |
10 | 72.0 | -- | |
11 | 10.0 | -- | hold |
Component | Final Concentration |
---|---|
ddH2O | |
Kapa 2G HS buffer | 1.30 X |
MgCl2 | 2.60 mM |
dNTPS-kapa | 0.26 mM |
oIMR0296 | 0.50 uM |
oIMR2060 | 0.50 uM |
Glycerol | 6.50 % |
Dye | 1.00 X |
Kapa 2G HS taq polym | 0.03 U/ul |
DNA |
Step | Temp °C | Time | Note |
---|---|---|---|
1 | 94.0 | -- | |
2 | 94.0 | -- | |
3 | 65.0 | -- | -0.5 C per cycle decrease |
4 | 68.0 | -- | |
5 | -- | repeat steps 2-4 for 10 cycles (Touchdown) | |
6 | 94.0 | -- | |
7 | 60.0 | -- | |
8 | 72.0 | -- | |
9 | -- | repeat steps 6-8 for 28 cycles | |
10 | 72.0 | -- | |
11 | 10.0 | -- | hold |
Stock Number | Strain Name |
---|---|
008437 | B6.129S2-Seletm1Hyn Selptm1Hyn/J |
008441 | B6.129S2-Selltm3Hyn Selptm1Hyn/J |
008432 | B6.129S2-Selptm1Hyn/J |
003807 | B6;129S-Seletm1Hyn Selltm1Hyn Selptm1Hyn/J |
003805 | B6;129S-Selltm3Hyn Selptm1Hyn/J |
002916 | B6;129S2-Seletm1Hyn Selptm1Hyn/J |
002217 | B6;129S2-Selptm1Hyn/J |
008438 | C.129S2(B6)-Seletm1Hyn Selptm1Hyn/J |
008442 | C.129S2(B6)-Selltm3Hyn Selptm1Hyn/J |
008433 | C.129S2(B6)-Selptm1Hyn/J |
10 strains use this protocol |