Protocol 30424: Probe Assay - Dusp22<em1(IMPC)J> - Probe
Version 1.0

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

Mut= 110 bp

Wt= 113 bp

Sequence

Wt Sequence:

 

Mutant Sequence:

JAX Protocol

Protocol Primers

Primer 5' Label Sequence 5' → 3' 3' Label Primer Type Reaction Note
34583 CAA TGC CCA GAG CAG CTT AC Common A
34584 Fluorophore-1 TAA GTG CAG GGG GTT GAA AG Quencher-1 WT Probe
34585 CAA TTC TGG TAC CTG ACT GAC C Wild type Reverse A
34586 CAC CGT GAC AAC GAG CTG Mutant Reverse A
34587 Fluorophore-2 CTT ATG GTA CTT TAT GCT GTG TGC Quencher-2 MUT Probe

Reaction A

Component Final Concentration
Kapa Probe Fast QPCR 1.00 X
ddH2O
34583 0.40 uM
34585 0.40 uM
34586 0.40 uM
Wt Probe 0.15 uM
Mutant Probe 0.15 uM
DNA

Cycling

Step Temp °C Time Note
1 95.0 --
2 95.0 --
3 60.0 --
4 -- repeat steps 2-3 for 40 cycles
5 40.0 -- Forever
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.