Protocol 32013: Probe Assay - Cep95<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

>chr11:106799813+106799895 83bp TGACACTCAAGCTCCTGCAC TGATCACAGCACAGTTTCACC

Mut= 83 bp

Wt= 83 bp

Fam=Mut

Hex=Wt

Sequence

Wt Sequence: (deleted region in lower case)

 

GTGGAGCGGTAAGTGAGCCTTGCTTTAACAGTGGGGCTGGATAGGGCTGGCAGAGCCTTGGTGGGTAAGCATTTGCCTCATAAGCTTGATAACCTGAATTCCATTCCAAGAACCCACAGAGAAGGAGAGAAAGAACTCCCCAAAGTTGTCCTCTGACCTCCACACTTGGGTTGTGACACTCAAGCTCCTGCACAAGTATGCCTCTCCCCCCCAAAGTgtgcaaacagggggaaatggtgaaactgtgctgtgatcacctgcctgccctgctgtgagagccccccttgtctcttcttctccaagggactcatggcaggtggtggttggcttgctcatgtcattatcatgttggctcaggacagacagggtcttaacttagcctttatgtttgtctttttaaacaggtgaaactgagcagtattctaaagattctcatggagaagaagctggggaggacctagaacgtaccgaggaggcgaagtggagaaacgcttcttttatgaggtagtgtgcagtgcctgtgaatttcatcagacaagttatgcttcccactctgtgagattaagtccagcatactctaatgtccctccaaaagctgatgtgaacattttatgcctttctGAGCTCTTGACTTTCCTTTCCTATCTCTGCTTTCACAACTTTTTTCTTTTTCTTTTTTTTTTTTTTCTTTTTTTGGTTTTTCGAGACAGGGTTTCTCTGTATAGCCCTGGCTATCCTGGAACTCACTTTGTAGACCAGGCTGGCCTCGAACTCAGAAATCCGCCTGCCTCTGCCTCCCAAGTGCTGGGAGTAAAGGCGTGCGCCACCACGCCCGGCTCACAACTTTTTTCATTCTAGACTTTTGTTGAGATGGTTC

This is a 404 bp deletion beginning at Chromosome 11 positive strand position 106,799,857 bp and ending after 106,800,260 bp (GRCm38/mm10).

JAX Protocol

Protocol Primers

Primer 5' Label Sequence 5' → 3' 3' Label Primer Type Reaction Note
38172 TGA CAC TCA AGC TCC TGC AC Common A
38173 TGA TCA CAG CAC AGT TTC ACC Wild type Reverse A
38174 GTT GTG AAA GCA GAG ATA GGA AAG Mutant Reverse A
38175 Fluorophore-1 TGC AAA CAG GGG GAA ATG Quencher-1 WT Probe
38176 Fluorophore-2 CCC CAA AGT GAG CTC TTG AC Quencher-2 MUT Probe

Reaction A

Component Final Concentration
Kapa Probe Fast QPCR 1.00 X
ddH2O
38172 0.40 uM
38173 0.40 uM
38174 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.