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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.
Tg = 69 bp
IPC= 74 bp
Strain 414192 - results should match Tg(Ins2-HBEGF)6832Ugfm.
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 |
Component | Final Concentration |
---|---|
ddH2O | |
Kapa Probe Fast QPCR | 1.00 X |
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 |
Step | Temp °C | Time | Note |
---|---|---|---|
1 | 95.0 | -- | |
2 | 95.0 | -- | |
3 | 60.0 | -- | repeat steps 2-3 for 40 cycles |
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-Or8a1tm27Mom/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 |
037280 | C57BL/6-Tg(Irgm1-DsRed2)1Nci/Mmjax |
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 |
025693 | STOCK Tg(CAG-mCherry/Pxn)#Mgin/KleyJ |
008241 | STOCK Tg(Cspg4-DsRed.T1)1Akik/J |
006866 | STOCK Tg(Ins1-DsRed*T4)32Hara/J |
19 strains use this protocol |