Stock No: 027962
Protocol 22016: End Point Analysis Assay - Ngly1_exon8_R399del-EP
Version 2.0

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 = ---/---
Heterozygote = GGA/---
Wild type = GGA/GGA

Sequence

ctattttttaaaaaaattaaaggaacattgtgatttttgttaaatagttgtgttctgtaac
tgattccagGTAGTTGATGTCACCTGGCGATACTCTTGTAAACATGATGAGGTGATGTCCAG
GA[gga]CCAAGGTTAAAGAAGAGTTACTTCGAGAAACTATTAATGGGCTTAATAAGCAGgt
atgtagtggctgtcattttaaagaggacatttgaaattttacatatcaggaatagtccgtt
ttaactc

JAX Protocol

Protocol Primers

Primer 5' Label Sequence 5' → 3' 3' Label Primer Type Reaction Note
24512 TCA CCT GGC GAT ACT CTT GT Forward A
24513 AGC CCA TTA ATA GTT TCT CGA AG Reverse A
24514 Fluorophore-1 TGT CCA GGA GGA CCA AGG TTA Quencher-1 WT Probe
24515 Fluorophore-2 TGA TGT CCA GGA CCA AGG TTA AAG Quencher-2 MUT Probe

Reaction A

Component Final Concentration
Kapa Probe Fast QPCR 1.00 X
ddH2O
24512 0.40 uM
24513 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.