Protocol 34665: End Point Analysis Assay - Snx1<em1Adiuj>
Version 1.0

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

Mutant = C, T, A

Wild type = A, G, G

Sequence

gctccatccagaccttaagtgagcctctctttcctcttcctagTGGGAATCTCGGGTGACTCAGTATGAA(a/c)G(g/t)(g/a)ACTTTGAAAGGATTTCCACAGTTGTCCGAAAGGAAGTGACACGATTTGAGgtgagctagaaaccatgtgtt

JAX Protocol

Protocol Primers

Primer 5' Label Sequence 5' → 3' 3' Label Primer Type Reaction Note
44177 CTC TTC CTA GTG GGA ATC TCG Forward A
44178 TTC CTT TCG GAC AAC TGT GG Reverse A
44179 Fluorophore-1 CTC AGT ATG AAA GGG ACT TTG AAA G Quencher-1 WT Probe
44180 Fluorophore-2 CTC AGT ATG AAC GTA ACT TTG AAA GG Quencher-2 MUT Probe

Reaction A

Component Final Concentration
Kapa Probe Fast QPCR 1.00 X
ddH2O
44177 0.40 uM
44178 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.