Protocol 32424: Probe Assay - Prrc2c<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

>chr1:162722804-162722959 156bp TGCCTACTAACTCCAAGAATAGAGTC AAGCAGAATGCTGGGTTCC

Mut= 150 bp

Wt= 156 bp

Fam=Mut

Hex=Wt

Sequence

Wt Sequence (deletion in lower case):

 

TGTGTAACCCTAAAGGGCTTAGAACTTAACAGTGGATCAGGCCAGCCTCCAACATGAAGCTATTTTCCCACTTTCACCTTCCAAGTTGTGTAGTTTGAACTTTCTTCTCTTAACATTTTAAGTTTTTTACTTAAAATAGATTGCCTGAAAACATTTCAAAACTAGCATTTCCCCTTCCCATATTCccagtatatagtgttccatatttagaagttaaggggttctggtgggtgggtttgttttgttttttgagacaaattttgctggggtcccaagatggcttaatattttgagtggattgtttactttttaccttcaattaatgttgtaggttataaactgagtttatagagctctactttaacattggggactagtcagagattgttaactcctgttcatgtatattatttgttgaataaaatgtataatagagtgatgaacagtagatgttctaaggaaattgatgttactgatcttttagttgcagctcgacatgggttacaaagtcttggaaaagttggtatttcacggcgaatgcccccacctgctaaccttccaagtctcaaagcagagaacaaaggcaatgatcctaatgtgaatattgtgcccaaagatgggacaggatgggcatcgaaacaagagcaacatgaagaagaaaagtgagtcagaatctgttaagaataaaacacttctctcattgtaggaaactgccaccgctgtgcactgttcagccaatagaaacactctatgacacttataattcagagagttcttcctcagatgaaaacacaagtcagcttactctttgatgcctactaactccaagaatagagtcttggacttctcttaacactattacttagcaaaaacatgaaaaatgaacttaacattctaagaacaaatctacaagccaaccaatatttactgcaaactattatcTGAGTGTGGAACCCAGCATTCTGCTTTAATAAGTCCTTGATAAATGAAGAGTTAAAAAGATAATATAGATATTATAACTCTTTTGTGGTGTTATATTTAAAGTGTGAATAGACACATTTAGTGTAAATATATAATAACTGTGTATACACACTTTTAGTCGTTAGGCTCTTTGAGTTCAGTAGCATTAGCCATAAGCAC

This mutation is a 757 bp deletion beginning at Chromosome 1 position 162,722,830 bp and ending after 162,723,586 bp (GRCm38/mm10).

JAX Protocol

Protocol Primers

Primer 5' Label Sequence 5' → 3' 3' Label Primer Type Reaction Note
39080 TGC CTA CTA ACT CCA AGA ATA GAG TC Wild type Forward A
39081 AAG CAG AAT GCT GGG TTC C Common A
39082 ATT TTC CCA CTT TCA CCT TCC Mutant Forward A
39083 Fluorophore-1 AGC CAA CCA ATA TTT ACT GCA AAC Quencher-1 WT Probe
39084 Fluorophore-2 TTC CCC TTC CCA TAT TCT GAG Quencher-2 MUT Probe

Reaction A

Component Final Concentration
Kapa Probe Fast QPCR 1.00 X
ddH2O
39080 0.40 uM
39081 0.40 uM
39082 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.