Stock No: 002231
Protocol 18486: End Point Analysis Assay - Pah<enu1>
Version 1.0

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

Mutant =C/C
Heterozygote = T/C
Wild type = T/T

Sequence

AGAGCCTGAGGAACGACATTGGTGCCACTg[T/C]cCATGAGCTTTCCCGAGACAAGGAAAAG

JAX Protocol

Protocol Primers

Primer 5' Label Sequence 5' → 3' 3' Label Primer Type Reaction Note
27129 ACC TAT CTG GAT AAG CGT AGC A Forward A
27130 GAC TCT CAC CTG TGT TCT TTT CC Reverse A
27131 Fluorophore-1 TGC CAC TGT CCA TGA GCT Quencher-1 WT Probe A
27132 Fluorophore-2 TGC CAC TGC CCA TGA GCT Quencher-2 MUT Probe A

Reaction A

Component Final Concentration
Kapa Probe Fast QPCR 1.00 X
ddH2O
27129 0.40 uM
27130 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 4.0 -- 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.