Stock No: 002258
Protocol 33838: Probe Assay - Lta<tm1Dch> Probe
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

>chr17:35204123-35204242 120bp CCTACCAACCATCCCACTTG GCAGTGAGTTCTGCTTGCTG

Mutant=  96 bp

Wild Type = 120 bp

Sequence

Wt Sequence: cctaccaaccatcccacttgtccagtgcctgctccctcagggatggggacctctgatcttgatagccccccaatgtcttgtgcctcttcccagGGTACCCCAGCAAGCAGAACTcaCTGC

 

Mutant Sequence: aaaagcgcctcccctacccggtagaattaattcgatatcaagcttatcgataccgtctcgacctcgactaGTACCCCAGCAAGCAGAACTCACTGCTCTGGAGAGCAAGCACGGATCGTGCCTTTCTCCGACATGGCTTCTCTTTGAGCAA

JAX Protocol

Protocol Primers

Primer 5' Label Sequence 5' → 3' 3' Label Primer Type Reaction Note
21722 AAA AGC GCC TCC CCT ACC Mutant Forward A
41941 CCT ACC AAC CAT CCC ACT TG Wild type Forward A
42307 GCA GTG AGT TCT GCT TGC TG Common A
42308 Fluorophore-1 CTT ATC GAT ACC GTC TCG ACC T Quencher-1 MUT Probe
42309 Fluorophore-2 TGC CTG CTC CCT CAG G Quencher-2 WT Probe

Reaction A

Component Final Concentration
Kapa Probe Fast QPCR 1.00 X
ddH2O
21722 0.40 uM
41941 0.40 uM
42307 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.