Protocol 33541: Probe Assay - Xylt1<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

>chr7:117549217+117549332 116bp CCTGCCGTCTGCTATCAAG TGAACACAGCACAGCACATC

Mut= 116 bp

Wt= 116 bp

Fam=Mut

Hex=Wt

Sequence

Wt Sequence (deletion in lower case):

ATAGATTTACTTGCCCTTAGGCAGTGTGTGCATGTATAAAATGTATGCAATATATATAAAATATACACAATGTAGACAGCACTGGTCCATATCGTCTCCTTTATGCTTTGATCCTGAGGATTCCAGCTCTACTGTATTCCAACCATTATAGCAGCACCTCCTGCTGCCatttgaacagaaccccagagggaaattttccccagacaggagagtggaggaggggatcttggagaattgcagtcactgtcctggtcactgtgagcgtggagaggctgtcttgtgggagcagagtgggaagctagtactcagagaaaagcatttccacagtccgagccaatgagagagaagaatggcagtcttggctcgggcttctccacctgccaccctctcggggccagcctgcttaccagcatggctcagtgacaattttctgtttctctaatcctccaggatggctacttctctcaccgtcccaaagagaaagttcggaccgacagcaacaatgagaactcagtccccaaggactttgagaatgtggataacagcaacttcgcccccaggacccagaagcagaaacaccagccagagctggcaaagaagccccccagcaggcagaaagagcatttgcagagaaagctggatgcccaggacaaacgacagggccagtcagtcctggggaaaggccccaaggaggtgctgcctcctcgggagaaagccacaggcaacagtagccaagggaaggatctctcaagacacagccatgccaggaagagtggtggtggtgggtcccccgaaaccaagtctgaccaggcccccaagtgtgatatctctggcaaggaggccatctcagcactgacccgcgctaagtccaagcattgtcgccaggagattgcagaaacctactgtcgccacaagctggggctgctgatgccagagaaggtggctcgattctgtcccctagaaggtaggtctctttctcctgtccctggccccatctctctgccttctcaggccctctgaatatagccacactgcacctcaaaccctgaccacaagatctggtttgtctgggaactctgaactcctgccgtctgctatcaagaaagtttgacaaaatcatgacatgtgatagtatcatatgacctgttccctttagcagATATGATCTCGGTGTCTGCAGATGTGCTGTGCTGTGTTCATAGCTATCTGGAATTTACAGGCTACAGAAATTAGAAAAAAATTAGACTCTGTCAGACGTAGGCTTTTTACGTATGCAATATTGTACAGCCCCAACATTACATA

This mutation is a 991 bp deletion beginning at Chromosome 7 position 117,548,302 bp and ending after 117,549,292 bp (GRCm38/mm10).

JAX Protocol

Protocol Primers

Primer 5' Label Sequence 5' → 3' 3' Label Primer Type Reaction Note
41655 CCT GCC GTC TGC TAT CAA G Wild type Forward A
41656 TGA ACA CAG CAC AGC ACA TC Common A
41657 CGT CTC CTT TAT GCT TTG ATC C Mutant Forward A
41658 Fluorophore-1 CAT ATG ACC TGT TCC CTT TAG CA Quencher-1 WT Probe
41659 Fluorophore-2 CTG CCA TAT GAT CTC GGT GTC Quencher-2 MUT Probe

Reaction A

Component Final Concentration
Kapa Probe Fast QPCR 1.00 X
ddH2O
41655 0.40 uM
41656 0.40 uM
41657 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.