Stock No: 015821
Protocol 41430: Probe Assay - Htr2c<tm1Jke>-Probe Alternate 1
Version 2.0

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

>chrX:147074037+147074167 131bp TGGACTCTTCGAGTTCCCACT CAGGAGGATGGGAGGTGAGT

Mut= 136 bp

Wt= 131 bp

Fam=Mut

Hex=Wt

X-Linked

Sequence

Wt Sequence:

aggccctgtctatttactgtaggtggactcttcgagttcccacttcccaatgttgagcattttggctaaggtcaccccaattgagtcctaagagtctcttgactcttgggtctctagaattttctagagagcccactcacctcccatcctcctgaggcagtgtagttccattctttctcttggccctctgggcttctctcct

Mutant Sequence:

GAAGGCAGGTCAGAGACCCCACTGGACAAACAGTGGCTGGACTCTGCACCATAACACACAATCAACAGGGGagtgagctccggatccataactTCGTATAGCATACATTATACGAAGTTATTCTTGACTCTTGGGTCTCTAGAATTTTCTAGAGAGCCCACTCACCTCCCATCCTCCTGAGGCAGTGTAGTTCCATTCTTTCTCTTGGCCCTCTGGGCTTCTCTCCTGTCCCCCCATACCTGATCCTGTTTCCCTTTTCCCCTTCCCATCCCCTCTCCTATCCAGGTCCTTCTCTTCCG

JAX Protocol

Protocol Primers

Primer 5' Label Sequence 5' → 3' 3' Label Primer Type Reaction Note
57761 TGG ACT CTT CGA GTT CCC ACT Wild type Forward A
57762 CAG GAG GAT GGG AGG TGA GT Common A
57763 CTG CAC CAT AAC ACA CAA TCA Mutant Forward A
58328 Fluorophore-1 CAT TTT GGC TAA GGT CAC CCC A Quencher-1 WT Probe
58329 Fluorophore-2 AGT GAG CTC CGG ATC CAT AAC TTC Quencher-2 MUT Probe

Reaction A

Component Final Concentration
Kapa Probe Fast QPCR 1.00 X
ddH2O
57761 0.40 uM
57762 0.40 uM
57763 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.