Stock No: 001162
Protocol 18500: End Point Analysis Assay - H2-Ab1<bm12> EP
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 =(t)TCCTGGAGC(a)AA(a)
Heterozygote = (a/t)TCCTGGAGC(g/a)AA(c/a)
Wild type = (a)TCCTGGAGC(g)AA(c)

Sequence

SEQ

JAX Protocol

Protocol Primers

Primer 5' Label Sequence 5' → 3' 3' Label Primer Type Reaction Note
27153 AGA CGC CGA GTA CTG GAA CA Forward A
27154 GGC CCC TCG TAG TTG TGT CT Reverse A
27155 Fluorophore-1 AGA TCC TGG AGC GAA CGC Quencher-1 WT Probe A
27156 Fluorophore-2 AGT TCC TGG AGC AAA AGC Quencher-2 MUT Probe A

Reaction A

Component Final Concentration
Kapa Probe Fast QPCR 1.00 X
ddH2O
27153 0.40 uM
27154 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.