Stock No: 027080
Protocol 22051: End Point Analysis Assay - Tbk1<em#3(G217R)Lutzy>
Version 2.0

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 = A/A
Heterozygote = G/A
Wild type = G/G

Sequence

cttcagCATCCGGACATGTATGAAAGGGCAGTGCTAAGAAAGGACCATCAGAAGAAGTACGG
GGCTACCGTTGATCTGTGGAGTGTTGGAGTGACATTCTACCATGCAGCCACG(g/a)GGTCGCTG
CCGTTTAGACCCTTCGAGGGGCCTCGGAGGAACAAAGAAGTAATgtaagcacct

JAX Protocol

Protocol Primers

Primer 5' Label Sequence 5' → 3' 3' Label Primer Type Reaction Note
24482 TGT GGA GTG TTG GAG TGA CA Forward A
24483 GGT GCT TAC ATT ACT TCT TTG TTC C Reverse A
24484 Fluorophore-1 AGC CAC GGG GTC GCT Quencher-1 WT Probe
24488 Fluorophore-2 CAG CCA CGA GGT CGC TG Quencher-2 MUT Probe

Reaction A

Component Final Concentration
Kapa Probe Fast QPCR 1.00 X
ddH2O
24482 0.40 uM
24483 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.