Protocol 32962: Probe Assay - Dnajc13<em1(IMPC)J>
Version 1.0

Notes

Taqman qPCR protocols are run on a real time PCR instrument. Use an appropriate instrument specific Fluorophore/Quencher combination.

 

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

>chr9:104238232-104238312 81bp GAAGAAGTTGTGTGTGTGAGAGC AAACAGTAACATGTGAAAGTCATCC

Mut= 87 bp

Wt= 81 bp

Fam=Mut

Hex=Wt

Sequence

Wt Sequence (deletion in lower case):

GAATTGGAGTTACACAGACAGTTGTGAGCTGCCACATGGGTGCTAGGAACTGAACACAGGTCCTCTGAAAGAGCACACAGTGCTCTTAAACCACTGAGCTATCTTTCTAGCCCCAAGGTCATACTTTTTATGCAAGAATATTAATAGTTCCCACTAAAAtaagtggcggatagattttaattttgataacattttaggacattaaatttgaaaatataaaaataaactatatttaatcaccattgaactattgatcagccaacatatttctatttatagaaaaatatgaaagtgtattcattttgtgttaatatcatatgttttcatttaaaaatttgttgcttaatactaaatgtaagcatgtctttccttgtaggtataaacgagtcttttcagttggaacccatgcagttactacctataatcccaataccttagaagtaacaaatcaggtaattttaatagaagttgttgagggtttatgtgaacctttaaaagtcaagattgcagcccatcttccctgctgaagtcagcagttcagaagagaaatgtcagcctgaagatgaatgtgtatatatgatgccttgtgttcagacctagaagtgaagaagttgtgtgtgtgagagcgagcgcgggcacatgctttctagactcTCGAGGGGATGACTTTCACATGTTACTGTTTTTAGCCATCTCTATAAAATTCGTCCCTTTCTAAAGTTGTTACTATTAATCAAGCATATTAGTTATGTCACAAGTGTTATTGTTCACAGACTTTCCAAAGAACTAGAATTACTTGTTTGTAAAGGCAA

This mutation is a 495 bp deletion beginning at Chromosome 9 position 104,238,263 bp and ending after 104,238,757 bp (GRCm38/mm10).

JAX Protocol

Protocol Primers

Primer 5' Label Sequence 5' → 3' 3' Label Primer Type Reaction Note
40349 GAA GAA GTT GTG TGT GTG AGA GC Wild type Forward A
40350 AAA CAG TAA CAT GTG AAA GTC ATC C Common A
40351 TTT CTA GCC CCA AGG TCA TAC Mutant Forward A
40352 Fluorophore-1 CGG GCA CAT GCT TTC TAG AC Quencher-1 WT Probe
40353 Fluorophore-2 ATA TTA ATA GTT CCC ACT AAA ATC GAG G Quencher-2 MUT Probe

Reaction A

Component Final Concentration
Kapa Probe Fast QPCR 1.00 X
ddH2O
40349 0.40 uM
40350 0.40 uM
40351 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.