Stock No: 007069
Protocol 22364: Standard PCR Assay - Apoe<tm1Unc>
Version 1.3

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 = ~245 bp
Heterozygote = ~155 bp and ~245 bp
Wild type = ~155 bp
Separated by gel electrophoresis on a 1.5% agarose gel.

JAX Protocol

Protocol Primers

Primer 5' Label Sequence 5' → 3' 3' Label Primer Type Reaction Note
oIMR0180 GCC TAG CCG AGG GAG AGC CG Common A
oIMR0181 TGT GAC TTG GGA GCT CTG CAG C Wild type Reverse A
oIMR0182 GCC GCC CCG ACT GCA TCT Mutant Reverse A

Reaction A

Component Final Concentration
ddH2O
Kapa 2G HS buffer 1.30 X
MgCl2 2.60 mM
dNTP KAPA 0.26 mM
oIMR0180 0.50 uM
oIMR0181 0.50 uM
oIMR0182 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
002246 B6.129-Apoetm1Unc Ldlrtm1Her/J
002052 B6.129P2-Apoetm1Unc/J
004631 B6.Cg-Apoetm1Unc Cdh18Tg(GFAP-APOE_i4)1Hol/J
007224 B6.Cg-Apoetm1Unc (D12Mit182-D12Mit2)/Pgn
008525 B6.Cg-Tg(Cyp21a1-Apoe)619Fet/J
004632 B6.Cg-Tg(GFAP-APOE_i2)14Hol Apoetm1Unc/J
004633 B6.Cg-Tg(GFAP-APOE_i3)37Hol Apoetm1Unc/J
002878 B6;129-Apobtm1Sgy Apoetm1Unc/J
002879 B6;129-Apobtm2Sgy Apoetm1Unc/J
002245 B6;129-Apoetm1Unc Ldlrtm1Her/J
004362 B6;129-Scarb1tm1Kri Apoetm1Unc/J
007067 D2.129P2(B6)-Apoetm1Unc/J
007069 STOCK Apoetm1Unc/J
13 strains use this protocol