Protocol 30067: QPCR Assay - Tg(Pax8-rtTA2S*M2)1Koes

B6.Cg-Tg(Pax8-rtTA2S*M2)1Koes/J

Stock No: 007176

Version 4.0

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

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
oIMR7387		CTG GAG AAC GCA CTG TAC GC		Transgene Forward	A
oIMR7388		CCA ATA CGC AGC CCA GTG T		Transgene Reverse	A
TmoIMR0107	Fluorophore-1	CCA ATG GTC GGG CAC TGC TCA A	Quencher-1	IC Probe
TmoIMR0110	Fluorophore-2	CTG TCC GCC GTG GGC CAC TT	Quencher-2	Tg Probe

Reaction A

Component	Final Concentration
ddH2O
Kapa Probe Fast QPCR	1.00 X
oIMR1544	0.40 uM
oIMR3580	0.40 uM
oIMR7387	0.40 uM
oIMR7388	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

This is the only strain that uses this protocol.