Protocol 36280: Probe Assay - Dnajc2<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

>chr5:21781122-21781223 102bp GGGACCTTGAGTGTTCTCAG AACCCAAAACAAACGAAACC

Mut= 97 bp

Wt= 102 bp

Fam=Mut

Hex=Wt

Sequence

Wt Sequence (deletion in lower case):

 ATCCACGACTGGCGCCTAACTTACATGACTGCACAATGACTTACATTCTTTTTCTTGTTGGTTGGTTGGTTGGTTTTGGTTTTGTTTTGTTCTTTTGATATGGGGTTTCTTGGTGTAGCCCTAGCTGTCCTGGAACTTGGATCTGTAGACAAGACTAGCCTTGAACTCATAGAAATATTTGGGCCTCTGCCTCCCAAGTGCTGGGACTTAAGGCATGCTGCATCACCACTCGCCTACTCATTTACACAGAGCCATTCTGTACAGAAGCTCTGGATTCTGCTACGGATGCTGCACTTCCGTTTGCTTGCTTGCTTGTGCTGCTAGAGATGGAACTCGGGCTCATTTCATGCTAAGCAGGTGCTctgtggctccagcctgaagcctcgctttggaaaaattttaatgatattaaaacatgagggttggatgtactctgttcagatttaactcccttttgtgtgtttatcagaatcaagatcattatgcggttcttggacttggccacgtcagatacacagctacacagagacagatcaaagcagctcgtaagtacttacttagtatgggaggtataactccaaactcgggggtgcagcaccctgaagttcatgagaccccatcttagaaagggaagggattagggggaccttgagtgttctcagagttcaagtcctgcaggaagagatcagaaaccctgtaagaatCATTGCCCATCAAGGAAACGGGTTTCGTTTGTTTTGGGTTTGGATGGTGCTGGGAATCAAACCTGAGGCCTTGAGTCCTGGCCAGCTCTGCCTGCCACTGAGCTTCATGCCTAGTCCTCTGCTCTGAGGATTTCTTCCAGGTAAGAGAGAAACAGGGTCTTGCTGTTCAGACCTTAGATCCATGTCCGCCTTCCAAGTGCTGGGATTGTAGACGGTTAGGTATGATCACCATGTTCTCGTCTAATTAAAAAATTAGGTCTCTTTTAGTTTTGCTTCCTAAAAAAGATTTTAATTTTGAAAAATTAGATGTGTGTATCTGTGTGGGTATGTGCACGTGAGTGTAGGTGTGTATCTGTGTGGGT

This mutation is a 3625 bp deletion beginning at Chromosome 19 position 41,924,589 bp and ending after 41,928,213 bp (GRCm38a 342 bp deletion beginning at Chromosome 5 position 21,781,162 bp and ending after 21,781,503 bp (GRCm38/mm10). mm10).

JAX Protocol

Protocol Primers

Primer 5' Label Sequence 5' → 3' 3' Label Primer Type Reaction Note
48298 GGG ACC TTG AGT GTT CTC AG Wild type Forward A
48299 AAC CCA AAA CAA ACG AAA CC Common A
48300 TGC TTG CTT GTG CTG CTA GA Mutant Forward A
48301 Fluorophore-1 AGT CCT GCA GGA AGA GAT CAG A Quencher-1 WT Probe
48302 Fluorophore-2 CTA AGC AGG TGC TCA TTG CC Quencher-2 MUT Probe

Reaction A

Component Final Concentration
Kapa Probe Fast QPCR 1.00 X
ddH2O
48298 0.40 uM
48299 0.40 uM
48300 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.