Stock Number: 008045 Strain Name: B6;129-Trp53tm2Holl/J

Allele: Trp53tm2Holl Protocol Name: Trp53tm2Holl Method: Standard PCR Version: 1.1 Created: 08-July -2008 Updated: 24-September-2009

Expected Results: HET = 362 bp and 450 bp MUT = 362 bp WT = 450 bp

Gel Image

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Gel Information: Separated by gel electrophoresis on a 1.5% agarose gel.

Protocol Primers

Primer 5' Label Sequence 5' --> 3' 3' Label Primer Type oIMR7777 - ACA GCG TGG TGG TAC CTT AT - Wild type Forward oIMR7778 - TAT ACT CAG AGC CGG CCT - Wild type Reverse oIMR7779 - CCT GGT CCT CTG ACT GCT CT - Mutant Forward oIMR7780 - GCC AGG CAT TGA AGT CTC AT - Mutant Reverse

Number of Reactions

Reaction/Components  Reaction Component Volume (µl) Final Concentration Total Volume (µl) ddH2O 3.36 - 3.36 10 X PCR Buffer II (ABI) 1.20 1.00 X 1.20 25 mM MgCl2 0.96 2.00 mM 0.96 2.5 mM dNTP 0.96 0.20 mM 0.96 20 uM oIMR7777 0.60 1.00 uM 0.60 20 uM oIMR7778 0.60 1.00 uM 0.60 20 uM oIMR7779 0.30 0.50 uM 0.30 20 uM oIMR7780 0.30 0.50 uM 0.30 5 mM DNA Loading Dye 1.66 0.69 mM 1.66 5 U/ul Taq DNA Polymerase 0.06 0.03 U/ul 0.06 DNA 2.00 - 2.00

Cycling  Step # Temp °C Time Note 1 94 3 min - 2 94 30 sec - 3 69 1 min - 4 72 1 min repeat steps 2-4 for 35 cycles 5 72 2 min - 6 10 - hold

This genotyping protocol is used for the following strains:   Stock Number Strain Name 008045 B6;129-Trp53tm2Holl/J Genotyping resources and troubleshooting This page contains guidelines, frequently asked questions, and helpful external links to genotyping and PCR resources.