Stock No: 018443
Protocol 37649: Standard PCR Assay - Arl6<tm2Vcs>-Alternate2
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 = 155 bp
Heterozygote = 155 bp and 286 bp
Wild type = 286 bp

JAX Protocol

Protocol Primers

Primer 5' Label Sequence 5' → 3' 3' Label Primer Type Reaction Note
51857 CAT GGC AGA TTT CTT CCT TCA Wild type Forward A
51858 CTT GGG AGG TAA GAT AAC AGA TCA Common A
51860 AGG GGA TCA ATT CAT AAC TTC G Mutant Forward A

Reaction A

Component Final Concentration
ddH2O
Kapa 2G HS buffer 1.30 X
MgCl2 2.60 mM
dNTP KAPA 0.26 mM
51857 0.50 uM
51858 0.50 uM
51860 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
021216 129(B6)-Arl6tm2Vcs/J
021215 B6.129-Arl6tm2Vcs/J
018443 B6;129-Arl6tm2Vcs/J
021217 C3.129(B6)-Arl6tm2Vcs/J
4 strains use this protocol