Stock Number: 008191 Strain Name: B6;129S2-Trp53tm1Tyj Nf1tm1Tyj/J

Allele: Nf1tm1Tyj Protocol Name: Nf1tm1Tyj Method: Standard PCR Version: 1.1 Created: 25-August -2008 Updated: 24-September-2009

Expected Results: Mutant = 350 bp Heterozygote = 203 bp and 350 bp Wild type = 203 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 oIMR8307 - GGT ATT GAA TTG AAG CAC - Wild type Forward oIMR8308 - TTC AAT ACC TGC CCA AGG - Common oIMR8309 - ATT CGC CAA TGA CAA GAC - Mutant Forward

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 oIMR8307 0.60 1.00 uM 0.60 20 uM oIMR8308 0.60 1.00 uM 0.60 20 uM oIMR8309 0.60 1.00 uM 0.60 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 60 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 008191 B6;129S2-Trp53tm1Tyj Nf1tm1Tyj/J 008192 B6.129S2-Nf1tm1Tyj/J Genotyping resources and troubleshooting This page contains guidelines, frequently asked questions, and helpful external links to genotyping and PCR resources.