Protocol 42698: Probe Assay - Gt(ROSA)26Sor<em#Mvw> (generic)
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

Mutant=  122 bp

Wild Type = 71 bp

>chr6:113075985-113076055 71bp GATCTGCAACTCCAGTCTTTCT ACACCAGGTTAGCCTTTAAGC

JAX Protocol

Protocol Primers

Primer 5' Label Sequence 5' → 3' 3' Label Primer Type Reaction Note
61347 GAT CTG CAA CTC CAG TCT TTC T Forward A
61348 ACA CCA GGT TAG CCT TTA AGC Reverse A
61349 Fluorophore-1 AGA AGA TGG GCG GGA GTC Quencher-1 WT Probe
61350 Fluorophore-2 CAG TGG TGT ACG GTA CAA ACC GG Quencher-2 MUT Probe

Reaction A

Component Final Concentration
Kapa Probe Fast QPCR 1.00 X
ddH2O
61347 0.40 uM
61348 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

Stock Number Strain Name
037406 129S1/SvImJ-Gt(ROSA)26Sorem12Mvw/Mmjax
037407 A/J-Gt(ROSA)26Sorem18Mvw/Mmjax
028573 C57BL/6J-Gt(ROSA)26Sorem2Mvw/Mmjax
037409 DBA/2J-Gt(ROSA)26Sorem14Mvw/Mmjax
037410 FVB/NJ-Gt(ROSA)26Sorem20Mvw/Mmjax
037412 NZO/HlLtJ-Gt(ROSA)26Sorem31Mvw/Mmjax
037413 PWK/PhJ-Gt(ROSA)26Sorem19Mvw/Mmjax
7 strains use this protocol