Stock Number: 006336 Strain Name: B6.129-Selplgtm1Rpmc/J

Allele: Selplgtm1Rpmc Protocol Name: Selplgtm1Rpmc Method: Separated PCR Version: 2.1 Created: 30-October -2008 Updated: 24-September-2009

Expected Results: Mutant = ~365 bp Heterozygote = 202 bp and ~365 bp Wild type = 202 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 oIMR7915 - CGG TTG GCA TTA TTT GTG TG - Common oIMR7916 - CCA AGA TGG TCA GCA GCA C - Wild type Reverse oIMR8162 - TGG ATG TGG AAT GTG TGC GAG - Mutant Reverse

Number of Reactions

Reaction/Components  Reaction Component Volume (µl) Final Concentration Total Volume (µl) ddH2O 3.72 - 3.72 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 oIMR7915 0.60 1.00 uM 0.60 20 uM oIMR8162 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 61 1 min - 4 72 1 min repeat steps 2-4 for 35 cycles 5 72 2 min - 6 10 - hold

Reaction/Components  Reaction Component Volume (µl) Final Concentration Total Volume (µl) ddH2O 3.72 - 3.72 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 oIMR7915 0.60 1.00 uM 0.60 20 uM oIMR7916 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 This genotyping protocol is used for the following strains:   Stock Number Strain Name 006336 B6.129-Selplgtm1Rpmc/J Genotyping resources and troubleshooting This page contains guidelines, frequently asked questions, and helpful external links to genotyping and PCR resources.

Cycling  Step # Temp °C Time Note 1 94 3 min - 2 94 30 sec - 3 61 1 min - 4 72 1 min repeat steps 2-4 for 35 cycles 5 72 2 min - 6 10 - hold