Protocol 33584: End Point Analysis Assay - Nexmif<em6(ins1)Lutzy>
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 = G/G
Heterozygote = -/G
Wild type = -/-

X-linked

Sequence

ATGATGAGTCAACCCTGGGCAGTGATGTCTGCTCCCTCAAAATTCGATATGAGTCCT[g]TTCAGGACAATGTTCGAGACAAGACCACTCTTCTGATGCAGGAAGATGCCCAATTCAATTTT

JAX Protocol

Protocol Primers

Primer 5' Label Sequence 5' → 3' 3' Label Primer Type Reaction Note
41755 CAG TGA TGT CTG CTC CCT CA Forward A
41756 TCC TGC ATC AGA AGA GTG GT Reverse A
41757 Fluorophore-1 CGA TAT GAG TCC TTT CAG GAC AAT Quencher-1 WT Probe
41758 Fluorophore-2 ATA TGA GTC CTG TTC AGG ACA ATG Quencher-2 MUT Probe

Reaction A

Component Final Concentration
Kapa Probe Fast QPCR 1.00 X
ddH2O
41755 0.40 uM
41756 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.