Protocol 34424: Probe Assay - Ccsap<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

>chr8:123845651-123845767 117bp AGCTCCTGCTACGGAAACG CCAGCTATGGAAACAAATTACCAG

Mut= 123 bp

Wt= 117 bp

Fam=Mut

Hex=Wt

Sequence

Wt Sequence (deletion in lower case):

GAGTGACTTAGAAGCCAGTCTTCAGCAGCTGCCCAGTAAGGACAGCCTAGTCTGTCCGCAAAGGAAGCATCCAATAGAGAAGCCCTCAGGGGCCTGTGGAGGGAGCTCCTGCTACGGAAACGCTGCTCATTTGGATAGCGATGGTGActcctgtggatgggcatcacgggtgctttggaatagttttggtttaatgctggtaatttgtttccatagctggtttaagtaaaacctgtagagttaaacatgtgactgacttgatgaaacattatttttcaacttaagtcatatatatgtatatatgtatgtgtgtgtgtgtatatatatatatatatatatatatatgtatgtatgtgtatatatatatatgaacttgaactttaaagtttttccatctcatcttgtcccttttttttttccaaaagcaccaccaaaaaaggacacggaagagaaacccgaagaacacaagacaaaagagactgatggggcacccagtggccctggaccgcggcaacagcctagtgccttatgtgccagaggaagtaagaaagccaccagaagcccacagagatcgacgagcaagataaaagagaacaagcacccgtttgctctttatggctggggagagagacagatggacatgggaagccagaagacacacaacgtctgcgcgtcagcctctgtgcacgaggtgtgccttgaacggggccagactagccctgagctgccttgggggcatttctcctatggctacggtgcccagcgccactggaagtccttgggttatgtgggagtccatggaagccaaaagaaggggtctctggtcctagaaagatggagtggactttggttaaCTGGGGCTACAGCCAGGGCTAAGGTCTGATGACAGGAAGAGGTAGCTGTCATTGTGGTGTATGAGTTCTGTGGTCTGGGCCACAGGGATAATGGGAGCAGGAGTTATCCCCTGGGGAGGCATC

This mutation is a 707 bp deletion beginning at Chromosome 8 position 123,845,017 bp and ending after 123,845,723 bp (GRCm38/mm10).

JAX Protocol

Protocol Primers

Primer 5' Label Sequence 5' → 3' 3' Label Primer Type Reaction Note
43583 AGC TCC TGC TAC GGA AAC G Common A
43584 CCA GCT ATG GAA ACA AAT TAC CAG Wild type Reverse A
43585 CCC AGA CCA CAG AAC TCA TAC A Mutant Reverse A
43586 Fluorophore-1 CCT GTG GAT GGG CAT CAC Quencher-1 WT Probe
43587 Fluorophore-2 CTG GGG CTA CAG CCA GG Quencher-2 MUT Probe

Reaction A

Component Final Concentration
Kapa Probe Fast QPCR 1.00 X
ddH2O
43583 0.40 uM
43584 0.40 uM
43585 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.