Protocol 36279: Probe Assay - Exosc1<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

>chr19:41924549-41924709 161bp TAAGTTCTTGCTCCCAAGTCC ACATCTTTAGCACTGGGTGTTAC

Mut= 160 bp

Wt= 161 bp

Fam=Mut

Hex=Wt

Sequence

MUT Sequence:

aaccccctcctctgacctctgtagacagcagacacacatgtggtacacacacatacatgtaggcaaaacactcatcgcatcagataactctaaaaataaaagcaaaaccaagcaaagacccatggtcccctacaattgtaacacccagtgctaaagatgt

This mutation is a 3625 bp deletion beginning at Chromosome 19 position 41,924,589 bp and ending after 41,928,213 bp (GRCm38/mm10).

JAX Protocol

Protocol Primers

Primer 5' Label Sequence 5' → 3' 3' Label Primer Type Reaction Note
48293 TAA GTT CTT GCT CCC AAG TCC Wild type Forward A
48294 ACA TCT TTA GCA CTG GGT GTT AC Common A
48295 AAC CCC CTC CTC TGA CCT CT Mutant Forward A
48296 Fluorophore-1 ACC ATG GCT ATC TGG GAA GC Quencher-1 WT Probe
48297 Fluorophore-2 ACC AAG CAA AGA CCC ATG GT Quencher-2 MUT Probe

Reaction A

Component Final Concentration
Kapa Probe Fast QPCR 1.00 X
ddH2O
48293 0.40 uM
48294 0.40 uM
48295 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.