Stock Number: 006952 Strain Name: B6.Cg-Akt2tm1.1Mbb Ldlrtm1Her/J

Allele: Akt2tm1.1Mbb Protocol Name: Akt2tm1.1Mbb Method: Standard PCR Version: 1.1 Created: 03-July -2008 Updated: 09-November -2009

Expected Results: Mutant = 277 bp Heterozygote = 110 bp and 277 bp Wild type = 110 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 oIMR6745 - TAC ACT TCA TTC TCA GTA TTG TTT TGC - Mutant Reverse oIMR6746 - ACC AAC CCC CTT TCA GCA CTT G - Common oIMR6747 - TGC ACA ATC TGT CTT CAT GCC AC - Wild type 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 oIMR6745 0.60 1.00 uM 0.60 20 uM oIMR6746 0.60 1.00 uM 0.60 20 uM oIMR6747 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 64 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 006952 B6.Cg-Akt2tm1.1Mbb Ldlrtm1Her/J 006966 B6.Cg-Akt2tm1.1Mbb/J Genotyping resources and troubleshooting This page contains guidelines, frequently asked questions, and helpful external links to genotyping and PCR resources.