Stock No: 016144
Protocol 29934: QPCR Assay - Human TARDBP
Version 3.0

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
10280 GTA CGG GGA TGT GAT GGA TG Transgene Forward A
10281 CGC AAT CTG ATC ATC TGC AA Transgene Reverse A
10282 Fluorophore-1 CCA AGC CAT TCA GGG CCT TTG C Quencher-1 Tg Probe
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
10280 0.40 uM
10281 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
010700 B6.Cg-Tg(Prnp-TARDBP*A315T)95Balo/J
017933 B6.Cg-Tg(Prnp-TARDBP*Q331K)103Dwc/J
017930 B6N.Cg-Tg(Prnp-TARDBP*Q331K)109Dwc/J
016608 C57BL/6-Tg(Prnp-TARDBP)3cPtrc/J
017604 C57BL/6-Tg(Prnp-TARDBP*M337V)4Ptrc/J
016144 STOCK Tg(Prnp-TARDBP)4Jlel/J
6 strains use this protocol