Stock Number: 005708 Strain Name: B6.129-Apbb1tm1Quhu/J

Allele: Apbb1tm1Quhu Protocol Name: Apbb1tm1Quhu Method: Standard PCR Version: 2.1 Created: 08-August -2008 Updated: 24-September-2009

Expected Results: Mutant = 500 bp Heterozygote = 173 bp and 500 bp Wild type = 173 bp

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

Protocol Primers

Primer 5' Label Sequence 5' --> 3' 3' Label Primer Type oIMR0204 - CAC GGG TAG CCA ACG CTA TGT C - Mutant Reverse oIMR0221 - GCC CTG AAT GAA CTG CAG GAC G - Mutant Forward oIMR5163 - CTC ATC ACT GGC TGC ACT GT - Wild type oIMR5165 - ACG AAG AGG AGG AGG AGG AG - Wild type

Number of Reactions

Reaction/Components  Reaction Component Volume (µl) Final Concentration Total Volume (µl) ddH2O 2.52 - 2.52 10 X PCR Buffer II (ABI) 1.20 1.00 X 1.20 25 mM MgCl2 1.20 2.50 mM 1.20 2.5 mM dNTP 0.96 0.20 mM 0.96 20 uM oIMR0221 0.60 1.00 uM 0.60 20 uM oIMR0204 0.60 1.00 uM 0.60 20 uM oIMR5163 0.60 1.00 uM 0.60 20 uM oIMR5165 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 70 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 005708 B6.129-Apbb1tm1Quhu/J Genotyping resources and troubleshooting This page contains guidelines, frequently asked questions, and helpful external links to genotyping and PCR resources.