Stock Number: 003295 Strain Name: B6;129-Ednrbtm1Ywa/J

Allele: Ednrbtm1Ywa Protocol Name: Ednrbtm1Ywa Method: Standard PCR Version: 1.1 Created: 28-July -2008 Updated: 24-September-2009

Expected Results: Mutant = 244 bp Heterozygote = 244 bp and 444 bp Wild type = 444 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 oIMR6448 - CAT GGT CTT GTT CCT GAT GC - Common oIMR6449 - GAT GAA CCT GCT CAG TGC AA - - oIMR7770 - ATA GAT TCG CCC TTG TGT CC - 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 oIMR6448 0.60 1.00 uM 0.60 20 uM oIMR6449 0.60 1.00 uM 0.60 20 uM oIMR7770 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 003295 B6;129-Ednrbtm1Ywa/J Genotyping resources and troubleshooting This page contains guidelines, frequently asked questions, and helpful external links to genotyping and PCR resources.