Stock No: 002167
Protocol 21936: End Point Analysis Assay - Cst6<ichq>End Point
Version 3.0

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/T, T/T, -/-
Heterozygote = C/T, C/T, G/-
Wild type = C/C, C/C, G/G

Sequence

 

CTGTAAAAGCCGCGCAGGATGGAAGTCCAGACACTGAATCCGCGGCTATGGAGCGTCCTCAC
TTCCCGCTGGCGATGGG(c/t)CTTGGC(c/t)T[g]CTCGCATTCTGCCTCCTGACCCTGTCACCTG
ACGCCCGCGCCGAGCTGCGGAGCCGCAGGACCGGAGAACGCCAGAACTTGTCACCCACCG
ACCCGCGGGTGCAAAAGGCCGCACAGGCAGCT
 
[-] indicates a deletion in the mutant sequence

 

JAX Protocol

Protocol Primers

Primer 5' Label Sequence 5' → 3' 3' Label Primer Type Reaction Note
14996 TAT GGA GCG TCC TCA CTT CC Forward A
14997 GAC AAG TTC TGG CGT TCT CC Reverse A
14998 Fluorophore-1 GGG CCT TGG CCT GCT C Quencher-1 WT Probe
14999 Fluorophore-2 ATG GGT CTT GGC TTC TCG Quencher-2 MUT Probe

Reaction A

Component Final Concentration
Kapa Probe Fast QPCR 1.00 X
ddH2O
14996 0.40 uM
14997 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.