Protocol 21938: End Point Analysis Assay - Disc1<del>-EP
Version 3.0

Notes

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

Mutation is a 25 bp deletion

Sequence

GACCCTGACCTCTGCCAACC[aggctcccttccaggtggagccacc]TGAGACCCTCAGGAGGTACT

JAX Protocol

Protocol Primers

Primer 5' Label Sequence 5' → 3' 3' Label Primer Type Reaction Note
17573 GCT GTG ACC TGA TGG CAC T Forward A
17574 ATG GAC AAG GGA CAG CAA AG Reverse A
17575 Fluorophore-1 CTC CCT TCC AGG TGG AGC C Quencher-1 WT Probe
17576 Fluorophore-2 CTC TGC CAA CCT GAG ACC CTC Quencher-2 MUT Probe

Reaction A

Component Final Concentration
Kapa Probe Fast QPCR 1.00 X
ddH2O
17573 0.40 uM
17574 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.