Protocol 46804: Probe Assay - Generic APP human genomic or cDNA -Alt1
Version 7.0

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