Strain Protocols

Strain Information
Strain Code: 01XAA
Strain Name: B6.Cg-Apctm2Rak/Nci
Gene Symbol: Apc
Protocol Information
Allele: Apc<tm2Rak>
Protocol Number: 1
Introductory Comment: This strain carries a conditional allele of Apc with LoxP sites flanking Exon 14. The strategy of this genotyping protocol is to detect the LoxP1 site upstream Exon 14. This strain is maintained homozygously, and it should be noted if the genotyping results reveal two products, as this means the strain was mistakenly outcrossed.

Primers

ApcF2 : 5'-gag aaa ccc tgt ctc gaa aaa a-3'
ApcR2 : 5'-agt gct gtt tct atg agt caa c-3'
Diagram showing the primer relataionships for Strain 01XAA.  This strain carries a conditional allele of Apc with LoxP sites flanking Exon 14. The strategy of this genotyping protocol is to detect the LoxP1 site upstream Exon 14.  This strain is maintained homozygously, and it should be noted if the genotyping results reveal two products, as this means the strain was mistakenly outcrossed.

Product Sizes

Primer Combination Product Size
ApcF2 / ApcR2 320 bp    WT Allele
ApcF2 / ApcR2 430 bp    LoxP

Master Mix

Reagent Final Concentration
10X Buffer 1X
25mM MgCl2 2.0mM
2.5mM dNTP Mix 0.2mM
100 µMApcF2 0.5 µM
100 µMApcR2 0.5 µM
5U/┬Ál Taq Pol 0.025 U/µl
Use 50 - 100 ng of DNA per reaction

Cycling Conditions

(Time in minutes:seconds)
Temperature Time Label
94ºC 3:00
94ºC 0:30
60ºC 0:30 35 Cycles
72ºC 0:30
72ºC 3:00
4ºC HOLD

Disclaimer: PCR conditions were optimized using QIAGEN-purified mouse tail DNA and a PTC-200 thermocycler (MJ Research). Optimal conditions may vary with DNA quality or equipment used.