Stock No: 006679
Protocol 22141: QPCR Assay - Generic DsRed
Version 7.0

Notes

Taqman qPCR protocols are run on a real time PCR instrument. Use an appropriate
instrument specific Fluorophore/Quencher combination. The transgene genotype is
determined by comparing ΔCt values of each unknown sample against known
homozygous and hemizygous controls, using appropriate endogenous references.

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

Tg = 69 bp

IPC= 74 bp

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
oIMR4082 TGA AGG GCG AGA TCC ACA A Transgene A
oIMR4083 TGG ACT TGA ACT CCA CCA GGT A Transgene A
TmoIMR0067 Fluorophore-1 CTG AAG CTG AAG GAC GGC GGC C 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
oIMR4082 0.40 uM
oIMR4083 0.40 uM
Tg Probe 0.15 uM
IC Probe 0.15 uM
DNA

Cycling

Step Temp °C Time Note
1 95.0 --
2 95.0 --
3 60.0 -- repeat steps 2-3 for 40 cycles
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
006055 B6.Cg-Tg(CAG-Bgeo,-DsRed*MST)1Nagy/J
006051 B6.Cg-Tg(CAG-DsRed*MST)1Nagy/J
008705 B6.Cg-Tg(CAG-DsRed,-EGFP)5Gae/J
006872 B6.Cg-Tg(Ins1-DsRed*T4)32Hara/J
018410 B6.Cg-Tg(Pnkd*A7V*A9V,-DsRed)671Ljp/J
013082 B6.FVB-Tg(Per2-DsRed*T3)12Obr/Mmjax
006679 B6;129P2-Olfr151tm27Mom/MomJ
008605 B6;C3-Tg(CAG-DsRed,-EGFP)5Gae/J
029332 B6N.Cg-Tg(P2rx4-tdTomato)1Khakh/J
031160 C.FVB-Tg(Acta2-DsRed)1Rkl/J
024046 C57BL/6-Tg(Csf1r-HBEGF/mCherry)1Mnz/J
009655 C57BL/6J-Tg(Dcx-DsRed)14Qlu/J
005328 NOD/ShiLt-Tg(Cd4-DsRed)4Lt/J
005438 STOCK Tg(CAG-Bgeo,-DsRed*MST)1Nagy/J
005441 STOCK Tg(CAG-DsRed*MST)1Nagy/J
008241 STOCK Tg(Cspg4-DsRed.T1)1Akik/J
006866 STOCK Tg(Ins1-DsRed*T4)32Hara/J
17 strains use this protocol