Genotyping Protocols Database
Protocol 28934: Standard PCR Assay - Apoe<tm1.1(APOE*4)Adiuj>
Version 1.2

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 = 148 bp
Heterozygote = 148 bp and 224 bp
Wild type = 224 bp

Gel on 3% due to nonspecific bands close to wt.

JAX Protocol

Protocol Primers

Primer 5' Label Sequence 5' → 3' 3' Label Primer Type Reaction Note
24463 AAT TTT TCC CTC CGC AGA CT Common A
24464 ACA GCT GCT CAG GGC TAT TG Wild type Reverse A
24465 AGG AGG TTG AGG TGA GGA TG 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
24463 0.50 uM
24464 0.50 uM
24465 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
027894 B6(SJL)-Apoetm1.1(APOE*4)Adiuj/J
028709 B6(SJL)-Apoetm1.1(APOE*4)Adiuj Trem2em1Adiuj/J
029017 B6.Cg-Apoeem3(APOE*)Adiuj/J
029018 B6.Cg-Apoeem2(APOE*)Adiuj/J
030283 B6.Cg-Apoetm1.1(APOE*4)Adiuj Abca7em2Adiuj Trem2em1Adiuj/J
030304 B6.Cg-Apoetm1.1(APOE*4)Adiuj Il1rapem1Adiuj Trem2em1Adiuj/J
030320 B6.Cg-Apoetm1.1(APOE*4)Adiuj Abca7em1Adiuj Trem2em1Adiuj/J
030670 B6.Cg-Apoetm1.1(APOE*4)Adiuj Appem1Adiuj Trem2em1Adiuj/J
030673 B6.Cg-Apoetm1.1(APOE*4)Adiuj Ceacam1em1Adiuj Trem2em1Adiuj/J
030674 B6.Cg-Apoetm1.1(APOE*4)Adiuj Plcg2em3Adiuj Trem2em1Adiuj/J
030771 B6;SJL-Apoetm1.1(APOE*4)Adiuj Plcg2em1Msasn Trem2em1Adiuj/J
030922 B6.Cg-Mthfrem1Adiuj Apoetm1.1(APOE*4)Adiuj Trem2em1Adiuj/J
031668 B6(SJL)-Cr2tm1(CR2,CR1)How Apoetm1.1(APOE*4)Adiuj Trem2em1Adiuj/J
031722 B6.Cg-Apoetm1.1(APOE*4)Adiuj Appem2Adiuj Trem2em1Adiuj/J
031938 B6.Cg-Kif21bem1Adiuj Apoetm1.1(APOE*4)Adiuj Trem2em1Adiuj/J
031940 B6.Cg-Apoetm1.1(APOE*4)Adiuj Sorl1em1Adiuj Trem2em1Adiuj/J
031942 B6.Cg-Apoetm1.1(APOE*4)Adiuj Snx1em1Adiuj Trem2em1Adiuj/J
031944 B6.Cg-Apoetm1.1(APOE*4)Adiuj Clasp2em1Adiuj Trem2em1Adiuj/J
031948 B6;SJL-Slc6a17em1Adiuj Apoetm1.1(APOE*4)Adiuj Trem2em1Adiuj/J
031950 B6;SJL-Apoetm1.1(APOE*4)Adiuj Mtmr4em1Adiuj Trem2em1Adiuj/J
031952 B6;SJL-Apoetm1.1(APOE*4)Adiuj Shc2em1Adiuj Trem2em1Adiuj/J
21 strains use this protocol