Genotyping Protocols Database
Stock No: 013144
Protocol 31940: QPCR Assay - Generic LacZ QPCR Alternate 1
Version 8.0

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

JAX Protocol

Protocol Primers

Primer 5' Label Sequence 5' → 3' 3' Label Primer Type Reaction Note
37301 GTT TAT GCA GCA ACG AGA CG Transgene Reverse A
37302 Fluorophore-1 CAG GAT ATG TGG CGG ATG AGC Quencher-1 Tg Probe
mom0095 TGA CGG CAG TTA TCT GGA AG Transgene Forward A
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
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
37301 0.40 uM
mom0095 0.40 uM
oIMR1544 0.40 uM
oIMR3580 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
002982 B6.Cg-Tg(CAG-lacZ)32And/J
004623 STOCK Tg(TCF/Lef1-lacZ)34Efu/J
005317 B6.Cg-Tg(BAT-lacZ)3Picc/J
005707 STOCK Rag1tm1Mom Tg(TIE2-lacZ)182Sato/J
005728 STOCK Tg(tetO-Ipf1,lacZ)958.1Macd/J
007922 STOCK Gli2tm2.1Alj/J
007972 B6;CBA-Tg(Olfr151-taulacZ)4Mom/MomJ
007973 B6;CBA-Tg(Olfr16-taulacZ)1Mom/MomJ
007974 B6;CBA-Tg(Olfr160-taulacZ)V4-7Mom/MomJ
007975 B6;CBA-Tg(OR8A1-taulacZ)1Mom/MomJ
007976 B6;CBA-Tg(Olfr713-taulacZ)4Mom/MomJ
008344 B6;DBA-Tg(Fos-tTA,Fos-EGFP*)1Mmay Tg(tetO-lacZ,tTA*)1Mmay/J
013144 B6.129P2-Aim2Gt(CSG445)Byg/J
024050 B6.Cg-Tg(Hmgcr-lacZ)H253Sest/J
025101 B6.Cg-Avpr1atm1Dgen Tg(AVPR1A)1Bux/BuxJ
026216 B6;SJL-Tg(Tnnt2-lacZ)497Jl/J
027315 STOCK Vegfatm2.1Nagy/J
028635 B6.129(Cg)-Cntnap2tm2Pele/J
18 strains use this protocol