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Notes
The genotyping protocol(s) presented here have been optimized for reagents and conditions used by The Jackson Laboratory (JAX).
To genotype animals, JAX recommends researchers validate the assay independently upon receipt of animals into their facility.
Reaction cycling temperature and times may require additional optimization based on the specific genotyping reagents used.
Expected Results
FAM = Mut
CY5 = IPC
JAX Protocol
Protocol Primers
| Primer | 5' Label | Sequence 5' → 3' | 3' Label | Primer Type | Reaction | Note |
|---|---|---|---|---|---|---|
| oIMR1544 | CAC GTG GGC TCC AGC ATT | Internal Positive Control Forward | A | |||
| oIMR3580 | TCA CCA GTC ATT TCT GCC TTT G | Internal Positive Control Reverse | A | |||
| oIMR5260 | TGG GTT CAA ACA AAG GTG CAA | Transgene Forward | A | |||
| oIMR7707 | GAT GAC GAT CAC TGT CGC TAT GAC | Transgene Reverse | A | |||
| TmoIMR0076 | Fluorophore-1 | CAT TGG ACT CAT GGT GGG CGG TG | Quencher-1 | Tg Probe | ||
| TmoIMR0105 | Fluorophore-2 | CCA ATG GTC GGG CAC TGC TCA A | Quencher-2 | IC Probe |
Reaction A
| Component | Final Concentration |
|---|---|
| Kapa Probe Fast QPCR | 1.00 X |
| ddH2O | |
| oIMR1544 | 0.40 uM |
| oIMR3580 | 0.40 uM |
| oIMR5260 | 0.40 uM |
| oIMR7707 | 0.40 uM |
| Wt Probe | 0.15 uM |
| Mutant Probe | 0.15 uM |
| DNA |
Cycling
| Step | Temp °C | Time | Note |
|---|---|---|---|
| 1 | 95.0 | -- | |
| 2 | 95.0 | -- | |
| 3 | 60.0 | -- | |
| 4 | -- | repeat steps 2-3 for 40 cycles | |
| 5 | 40.0 | -- | Forever |
JAX uses a very high speed Taq (~1000 bp/sec), use cycling times recommended for your reagents.
Strains Using This Protocol
| Stock Number | Strain Name |
|---|---|
| 006409 | 129S1.129(Cg)-Tg(APPSw)40Btla/Mmjax |
| 031988 | 129S4.Cg-Tg(APPSwe,tauP301L)1Lfa Psen1tm1Mpm/LfaJ |
| 006555 | A.129(B6)-Tg(APPSw)40Btla/Mmjax |
| 005300 | B6.129-Tg(APPSw)40Btla/Mmjax |
| 005301 | B6.129S2-Tg(APP)8.9Btla/J |
| 006406 | B6.129S4-Tg(APPSwLon)96Btla/Mmjax |
| 004661 | B6.Cg-Zbtb20Tg(PDGFB-APPSwInd)20Lms/1J |
| 006293 | B6.Cg-Zbtb20Tg(PDGFB-APPSwInd)20Lms/2Mmjax |
| 005866 | B6.Cg-Tg(APP695)3Dbo Tg(PSEN1dE9)S9Dbo/Mmjax |
| 029770 | B6.Cg-Tg(APPswe,PSEN1dE9)85Dbo Apptm1Dbo/Mmjax |
| 008730 | B6.Cg-Tg(APPSwFlLon,PSEN1*M146L*L286V)6799Vas/Mmjax |
| 004662 | B6.Cg-Tg(PDGFB-APP)5Lms/J |
| 006006 | B6.Cg-Tg(Prnp-APP)A-2Dbo/J |
| 006005 | B6.Cg-Tg(Prnp-App/APPswe)E1-2Dbo/Mmjax |
| 007051 | B6.Cg-Tg(tetO-APPSwInd)102Dbo/Mmjax |
| 007052 | B6.Cg-Tg(tetO-APPSwInd)107Dbo/Mmjax |
| 007049 | B6.Cg-Tg(tetO-APPSwInd)885Dbo/Mmjax |
| 004807 | B6;129-Tg(APPSwe,tauP301L)1Lfa Psen1tm1Mpm/Mmjax |
| 004462 | B6;C3-Tg(APPswe,PSEN1dE9)85Dbo/Mmjax |
| 006004 | B6C3-Tg(tetO-APPSwInd)885Dbo/Mmjax |
| 003375 | C3B6-Tg(APP695)3Dbo/Mmjax |
| 007027 | C57BL/6-Tg(Thy1-APPSwDutIowa)BWevn/Mmjax |
| 006472 | D2.129(B6)-Tg(APPSw)40Btla/Mmjax |
| 035561 | STOCK Tc(HSA21,CAG-EGFP)1Yakaz/J |
| 24 strains use this protocol | |