Stock No: 002848
Protocol 28049: Standard PCR Assay - Fcgr2b<tm1Ttk>alternate1
Version 4.2

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

Mutant = ~253 bp
Heterozygote = ~253 bp and 439 bp
Wild type = 439 bp

JAX Protocol

Protocol Primers

Primer 5' Label Sequence 5' → 3' 3' Label Primer Type Reaction Note
12379 ATC GCC TTC TAT CGC CTT CT Mutant Forward A
14095 TGC TTT ACC CTG CTG AGG AT Wild type Forward A
14096 GGT CTG CTC CAT TTG ACA CC Common A

Reaction A

Component Final Concentration
ddH2O
Kapa 2G HS buffer 1.30 X
MgCl2 2.60 mM
dNTP KAPA 0.26 mM
12379 0.50 uM
14095 0.50 uM
14096 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
003817 B6;129S-Fcgr2btm1Ttk Clcn1adr-mto5J/J
002848 B6;129S-Fcgr2btm1Ttk/J
2 strains use this protocol