Stock Number: 004912 Strain Name: B6.129P2-Akt1tm1Mbb/J

Allele: Akt1tm1Mbb Protocol Name: Akt1tm1Mbb Method: Standard PCR Version: 1.1 Created: 07-April -2004 Updated: 16-November -2009

Expected Results: Mutant = 259 bp Heterozygote = 143 bp and 259 bp Wild type = 143 bp

Gel Image

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Gel Information: Separated by gel electrophoresis on a 3.0% agarose gel.

Protocol Primers

Primer 5' Label Sequence 5' --> 3' 3' Label Primer Type oIMR3181 - AGC TCT TCT TCC ACC TGT CTC - Wild type oIMR3182 - GCT CCA TAA GCA CAC CTT CAG G - Common oIMR3994 - GTG GAT GTG GAA TGT GTG CGA G - Common

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 oIMR3181 0.60 1.00 uM 0.60 20 uM oIMR3182 0.60 1.00 uM 0.60 20 uM oIMR3994 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 65 30 sec - 4 72 30 sec repeat steps 2-4 for 35 cycles 8 72 2 min - 9 10 - hold

This genotyping protocol is used for the following strains:   Stock Number Strain Name 004912 B6.129P2-Akt1tm1Mbb/J Genotyping resources and troubleshooting This page contains guidelines, frequently asked questions, and helpful external links to genotyping and PCR resources.