Genotyping Protocols Database
Stock No: 002972
Protocol 26277: Standard PCR Assay - Sod1<tm1Leb>
Version 2.2

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

Melt Peaks
Wt = 83°C +/- 1
Mut = 92
°C +/- 1

Mutant (HPRT) = ~240 bp
Wild Type (Sod1) = ~123 bp

JAX Protocol

Protocol Primers

Primer 5' Label Sequence 5' → 3' 3' Label Primer Type Reaction Note
oIMR0781 TGT TCT CCT CTT CCT CAT CTC C Mutant A
oIMR0782 ACC CTT TCC AAA TCC TCA GC Mutant A
oIMR0878 TGA ACC AGT TGT GTT GTC AGG Wild type A
oIMR0888 TCC ATC ACT GGT CAC TAG CC Wild type A

Reaction A

Component Final Concentration
ddH2O
Kapa 2G HS buffer 1.30 X
MgCl2 2.60 mM
dNTP KAPA 0.26 mM
oIMR0781 0.50 uM
oIMR0782 0.50 uM
oIMR0878 0.50 uM
oIMR0888 0.50 uM
Glycerol 6.50 %
Dye 1.00 X
Kapa 2G HS taq polym 0.03 U/ul
DNA

Cycling

Step Temp °C Time Note
1 94.0 --
2 94.0 --
3 65.0 -- -0.5 C per cycle decrease
4 68.0 --
5 -- repeat steps 2-4 for 10 cycles (Touchdown)
6 94.0 --
7 60.0 --
8 72.0 --
9 -- repeat steps 6-8 for 28 cycles
10 72.0 --
11 10.0 -- hold
JAX uses a very high speed Taq (~1000 bp/sec), use cycling times recommended for your reagents.
JAX uses a 'touchdown' cycling protocol and therefore has not calculated the optimal annealing temperature for each set of primers.

Strains Using This Protocol

Stock Number Strain Name
003881 B6.129S7-Sod1tm1Leb/DnJ
002972 B6;129S-Sod1tm1Leb/J
2 strains use this protocol