Stock No: 005304
Protocol 18706: End Point Analysis Assay - H2<b>rs253049200-Alt 1-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

Sequence

AAGCAGACACAGGAGAGAGGATGTTCTGTaaaacaagcatgtga

Aaggatgcg(A/-)tg(CGTG/-----)atgaaggattctttgctaaaGACACA

ATTATGGATCTGCAA

JAX Protocol

Protocol Primers

Primer 5' Label Sequence 5' → 3' 3' Label Primer Type Reaction Note
21943 Fluorophore-1 ATG CGA TGC GTG ATG AAG Quencher-1 MUT Probe
21944 Fluorophore-2 AAG GAT GCG TGA TGA AGG AT Quencher-2 WT Probe
22205 CAC AGG AGA GAG GAT GTT CTG Forward A
22206 AGG GTT GCA GAT CCA TAA TTG Reverse A

Reaction A

Component Final Concentration
Kapa Probe Fast QPCR 1.00 X
ddH2O
22205 0.40 uM
22206 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 -- repeat steps 2-3 for 40 cycles
JAX uses a very high speed Taq (~1000 bp/sec), use cycling times recommended for your reagents.

Strains Using This Protocol

Stock Number Strain Name
002845 C.B-H2b Tg(H2-Dd)D8Gja/LilJ
001952 C.B10-H2b/LilMcdJ
000438 C3.SW-H2b/SnJ
000666 C57BL/10SnJ
004353 C57BL/6-Tg(UBC-GFP)30Scha/J
000664 C57BL/6J
005304 C57BL/6NJ
007076 CByJ.B6-Tg(UBC-GFP)30Scha H2b/J
002591 NOD.B10Sn-H2b/J
039016 NOD.Cg-H2b Commd10Tg(Vav1-icre)A2Kio/JkraJ
039015 NOD.Cg-Myd88tm1.1Defr H2b/JkraJ
039017 NOD.Cg-Myd88tm1Defr H2b/JkraJ
039018 NOD.Cg-Myd88tm1Hlz H2b/JkraJ
13 strains use this protocol