Protocol 20925: Probe Assay - Zfp92<em1J> 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

Mutant=  116 bp

Wild Type =93 bp

JAX Protocol

Protocol Primers

Primer 5' Label Sequence 5' → 3' 3' Label Primer Type Reaction Note
31315 CCT GAC TAT GGC AGC CAC TC Mutant Forward A
31316 Fluorophore-1 CAA AGC CCA AGG TAA GTG GT Quencher-1 MUT Probe
31317 TGT TCT TCT AAA TAG CTT GGA CTT T Wild type Forward A
31318 Fluorophore-2 AAC AAC CTG TCA GAC CCA TTT Quencher-2 WT Probe
31319 TGG GAA TCT ATG ATT TTG TGC Common A

Reaction A

Component Final Concentration
Kapa Probe Fast QPCR 1.00 X
ddH2O
31315 0.40 uM
31317 0.40 uM
31319 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.