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Mutant = ~295 bp
Heterozygote = 185 bp and ~295 bp
Wild type = 185 bp
Primer | 5' Label | Sequence 5' → 3' | 3' Label | Primer Type | Reaction | Note |
---|---|---|---|---|---|---|
15442 | ATG GGC GAG TGC TAC TTC AC | Wild type Forward | B | |||
15801 | GCC CGC GTT CGC TCC AGG | Common | A, B | |||
oIMR1100 | GCT ATC AGG ACA TAG CGT TGG | Mutant Forward | A | neo F |
Component | Final Concentration |
---|---|
ddH2O | |
Kapa 2G HS buffer | 1.30 X |
MgCl2 | 2.60 mM |
dNTPS-kapa | 0.26 mM |
15801 | 0.50 uM |
oIMR1100 | 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 |
15442 | 0.50 uM |
15801 | 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 |
---|---|
038341 | B6.Cg-H2-Ab1b-tm1Gru Tg(Itgax-H2-Ab1)1Tml/J |
038339 | B6.Cg-H2-Ab1b-tm1Gru Tg(KRT14-H2-Ab1)1Glm/TmlJ |
002956 | B6NTac.Cg-H2-Ab1b-tm1Gru Tg(Ab1TL)1Gru/J |
004256 | NOD.129S2(B6)-H2-Ab1b-tm1Doi/DvsJ |
006021 | NOD.Cg-H2-Ab1b-tm1Gru Tg(CD2-CD4,HLA-DQA1,HLA-DQB1)1Ell/EllJ |
006023 | NOD.Cg-H2-Ab1b-tm1Gru Tg(CD4,HLA-DQA1,HLA-DQB1)N8Ell/EllJ |
026561 | NOD.Cg-Prkdcscid H2-Ab1b-tm1Doi Il2rgtm1Wjl Tg(HLA-DQA1,HLA-DQB1)1Dv/SzJ |
021885 | NOD.Cg-Prkdcscid H2-Ab1b-tm1Doi Il2rgtm1Wjl/SzJ |
006022 | NOD.Cg-Rag1tm1Mom H2-Ab1b-tm1Gru Tg(CD2-CD4,HLA-DQA1,HLA-DQB1)1Ell/EllJ |
006024 | NOD.Cg-Rag1tm1Mom H2-Ab1b-tm1Gru Tg(CD4,HLA-DQA1,HLA-DQB1)N8Ell/EllJ |
030331 | NOD.Cg-Tg(HLA-DRA*0101,HLA-DRB1*0101)1Dmz Prkdcscid H2-Ab1b-tm1Doi Il2rgtm1Wjl/SzJ |
11 strains use this protocol |