Protocol 36496: Probe Assay - Slc6a7<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

>chr18:61009177-61009277 101bp CTTTGAATTCCCAGCCTGATAC TTCCACCTACCATAATAGGCAAC

Mut = 108 bp

Wt = 101 bp

Fam = Mut

Hex = Wt

Sequence

Wt Sequence (deletions in lower case):

 TTCTGATGCCTTCCTCAGGGAGCCCACATTCTCTGAAATCTTTTGCTTCCTGTGCCAAGGGTACCCTGGCTTCTTCCCTCTGTCCCCCTTGGAGCCTCTTGTCCTGAACACCTTCCAGACTGGCCACCTGTCCTCCGGTAAGCTCCACACTGTAGGCAAAGACTGAGCCATTAGCATTAGGAGTTCTGATGCCCCGCCTGGTTCTCCTGGCCAAGCAATCAAAACCCTCAGGTCGTTCATGACCCCTCCTGGAGCCTCTGCTTTCTTCACACTTGGAATAGAGTGTGGGTGAGTAGTTGGCTCAGGTGTAGAATGTTTACTGCTATAATCACTAACACGTGGTGACAACACACAGTGACTGCCACAAAGATCTTCACTGGCAGTGATATTAAAGTGGgtcatccttggcctatttctgctagccaagatggagacgggcagcttcctaaggcctgagccagctgccggcccctaaggatctgcaagctaggccctcttgagtccaggtgaaatcaccggatcatctgaacctgatgcgagatttcccactcaaccttgctcaacccacggtctcagtttggcctcaccctcctccctctctgctttttttcttctctctctctctcttactggcagcccatcaccccagacctgctgatgactcccagtgaccagggcgatgtggacctggatgtagacttcgcagcagacaggggcaactggacgggcaagctggacttcctgctgtcctgcatcggctactgcgtgggcttgggaaatgtctggcgctttccctaccgagcctacaccaatggaggcggtatgggcctgggaacctcttgggagtagctggggagacagaccaaggcttgggggctataggatggcctttggagtgaccttaggttagagggaggtggcttactctgcctgtcccttttcaccgcctaagcccctcctcctgctttgaattcccagcctgatacccaggtttcctccctaagacctaaagctttcctagaacctatagtggGCAGACATTGTTGCCTATTATGGTAGGTGGAAAGGGTAGTACCCACTGTCACTATTCATTGGCTTTGCAGTGTATCCTGTCCCCTCCCCCACCTCTCCCAGCGCCCTGCTCTACCCACAGTTCCCTTTCTAGAATCTTCCTTGTTCCCTCCTCTGCTGATTCTTTCCATTCAGACTGAAAAACGCAAGATCT

This mutation is a 636 bp deletion beginning at Chromosome 18 position 61,009,209 bp and ending after 61,009,844 bp (GRCm38/mm10).

JAX Protocol

Protocol Primers

Primer 5' Label Sequence 5' → 3' 3' Label Primer Type Reaction Note
48905 TTC CAC CTA CCA TAA TAG GCA AC Common A
48906 CTT TGA ATT CCC AGC CTG ATA C Wild type Forward A
48907 GCT ATA ATC ACT AAC ACG TGG TGA C Mutant Forward A
48908 Fluorophore-1 CAG GTT TCC TCC CTA AGA CCT AA Quencher-1 WT Probe
48909 Fluorophore-2 CAG TGA TAT TAA AGT GGG CAG ACA Quencher-2 MUT Probe

Reaction A

Component Final Concentration
Kapa Probe Fast QPCR 1.00 X
ddH2O
48905 0.40 uM
48906 0.40 uM
48907 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.