Strain Protocols

Strain Information
Strain Code: 01XBX
Strain Name: B6.129S-Trp53tm1Snj/Nci
Gene Symbol: Tp53
Protocol Information
Allele: Tp53<tm1Snj>/Nci
Protocol Number: 1
Introductory Comment: The genotyping protocol utilizes two alleleic specific amplifications: one is for mutant allele and the other is for wild type allele. The genotyping results from both PCR is combined to determine the genotypes. Mutant specific PCR: primers Y707 and Y709. Wild type specific PCR: primers Y707 and Y708. The primer concentration used in PCR is 0.5 uM each.

Primers

Y707 : 5'-CCT CGA GCT CCC TCT GAG-3'
Y708 : 5'-TGG CCT CCA ACT TGC TAT GT-3'
Y709 : 5'-CCT CGA ACT CCC TCT GGC-3'

Product Sizes

Primer Combination Product Size
Y707 / Y709 266 bp    Mutant
Y707 / Y708 266 bp    Wild type

Master Mix

Reagent Final Concentration
10X Buffer 1X
25mM MgCl2
2.5mM dNTP Mix 0.2mM
100 µMY707 0.5 µM
100 µMY708 0.5 µM
100 µMY709 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 1:00
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.