Stock No: 032998
Protocol 34902: Probe Assay - Il1b<em2(del286)Mmjax>-Probe
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=  112 bp

Wild Type = 94 bp 

>chr2:129195347-129195440 94bp AGGTGAGACTCTGAGCTTGG GAGCAAGGCAGTCCCATATC

Sequence

Wt Sequence:

ATTTAGGTGTCAGAAGTCAACAGAGAGCAACCAGAGCACCCActtacctggcccagaatctgcttctagactgttacttacgtagcactgtagaacttcccagaagtctttgggaagctttaagagtcatggtaggtgtggttttatgaacaatggaagtctgtctgctcagtattgataaggagctatcacttgaccacatctgttttcattacagtgatgagaatgacctgttctttgaagttgacggaccccaaaagatgaaggtgagactctgagcttggctcctaacctgtggagggactgtggcatgggaaaaaaaaacccaAACGGGAAGGGATATGGGACTGCCTTGCTCCCAGGCTATTTTGTGATGGGA

Mutant Sequence: TGCTAGTCTAGAATTTAGGTGTCAGAAGTCAACAGAGAGCAACCAGAGCACCCaaACGGGAAGGGATATGGGACTGCCTTGCTCCCAGGCTATTTTGTGATGGGATTATTTTGCCTCTTTC

JAX Protocol

Protocol Primers

Primer 5' Label Sequence 5' → 3' 3' Label Primer Type Reaction Note
44749 AGG TGA GAC TCT GAG CTT GG Wild type Forward A
44750 GAG CAA GGC AGT CCC ATA TC Common A
44751 Fluorophore-1 ACC TGT GGA GGG ACT GTG G Quencher-1 WT Probe
44752 GGC AGA GAG CTC AGA CTT TAG C Mutant Forward A
44753 Fluorophore-2 AGA GCA CCC AAA CGG GAA Quencher-2 MUT Probe

Reaction A

Component Final Concentration
Kapa Probe Fast QPCR 1.00 X
ddH2O
44749 0.40 uM
44750 0.40 uM
44752 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.