Stock No: 036270
Protocol 40931: Probe Assay - Naa10<em3(indel)Lutzy>
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

>chrX:73919816-73919905 90bp CAGTGGTCGGACTTGTACCC CTGGTCCATCAGCTTCTGAG

Mutant=  92 bp

Wild Type = 90 bp

X-Linked

Sequence

Wt Sequence:

tgagtataactactgagggtttggtagaggtagactggagataattgaaggagccacagtggtcggacttgtacccaatcttcttctctcagGCTGTGAAGCGTTCCCACCGGCGCCTTGGCCTGGCTCAGAAGCTGATGGACCAGGCCTCTCGAGCCATGATAGAGAACTTCAATGCCAAATACGTCTCCCTGCATGTCAG

Mutant Sequence:

aactactgagggtttggtagaggtagactggagataattgaaggagccacagtggtcggacttgtacccaatcttcttctctcagGCTGTGAAGCGTTCCCACCGGCGagCCTTGGCCTGGCTCAGAAGCTGATGGACCAGGCCTCTCGAGCCATGATAGAGAACTTCAATGCCAAATACGTCTCCCTGCATGTCAGGAAGAGgtggaagccaagttgggggaaag

JAX Protocol

Protocol Primers

Primer 5' Label Sequence 5' → 3' 3' Label Primer Type Reaction Note
57102 CAG TGG TCG GAC TTG TAC CC Forward A
57103 CTG GTC CAT CAG CTT CTG AG Reverse A
57106 Fluorophore-1 CAC CGG CGC CTT GGC Quencher-1 WT Probe
57107 Fluorophore-2 ACC GGC GAG CCT TGG C Quencher-2 MUT Probe

Reaction A

Component Final Concentration
Kapa Probe Fast QPCR 1.00 X
ddH2O
57102 0.40 uM
57103 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.