Stock No: 026627
Protocol 21985: End Point Analysis Assay - Il12a<tm1Jm> Plps-EP
Version 3.0

Notes

Converting to 12ul, email genotyping@jax.org with jpeg

Wild Type: GCCCCCT

Mutant:     GCCCCCCT

Mutation is an inserted C

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

Wild Type: GCCCCCT

Mutant:     GCCCCCCT

 

Sequence

GGGAACAGATACCAGCTAT(GCCCCCT/GCCCCCCT)GGAGATCCATTGGGGTTCAGAGGGTACTGCTCTG

JAX Protocol

Protocol Primers

Primer 5' Label Sequence 5' → 3' 3' Label Primer Type Reaction Note
17517 GGT GGG AAG AAC ATC AAA GG Forward A
17518 GAA CCC CAC AGA GCA GTA CC Reverse A
17519 Fluorophore-1 GCT ATG CCC CCT GGA GA Quencher-1 WT Probe
17520 Fluorophore-2 GCT ATG CCC CCC TGG AGA Quencher-2 MUT Probe

Reaction A

Component Final Concentration
Kapa Probe Fast QPCR 1.00 X
ddH2O
17517 0.40 uM
17518 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.