Stock No: 034974
Protocol 39694: End Point Analysis Assay - Kcnq2<em8(indel)Lutzy>-alternate2
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 = -/A
Wild type = -/-

  
>chr2:181113022-181113101 80bp TGGAAGCTCTTGGGATCG ATGGTAAACAGCAGGCTCCA

Mutation is an A insertion

The WT and Mut probes (probes 54075 and 54076) anneals over the nucleotide sequence containing mouse genomic variation rs27642402.

Sequence

WT sequence showing [insertion]:

GTATGGACCGGAGGGGTGGCACCTGGAAGCTCTTGGGATCGGTAGTCTACGCTCACA[a]GCAAGGTGAGCAGCACTGACCCAGCCTGGAGCCTGCTGTTTACCATGACTTCAAGGCTCCAGGCTGCTCTA

JAX Protocol

Protocol Primers

Primer 5' Label Sequence 5' → 3' 3' Label Primer Type Reaction Note
54075 Fluorophore-1 ACG CTC ACA GCA AGG TGA Quencher-1 WT Probe
54076 Fluorophore-2 ACG CTC ACA AGC AAG GTG A Quencher-2 MUT Probe
54414 TGG AAG CTC TTG GGA TCG A
54415 ATG GTA AAC AGC AGG CTC CA A

Reaction A

Component Final Concentration
Kapa Probe Fast QPCR 1.00 X
ddH2O
54414 0.40 uM
54415 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.