Stock Number: 002459 Strain Name: B6D2-Tg(MMTVTGFA)254Rjc/J

Allele: Tg(MMTVTGFA)254Rjc Protocol Name: Tg(MMTVTGFA) Method: Standard PCR Version: 2.1 Created: 17-April -2009 Updated: 24-September-2009

Expected Results: TG = 596 wt= 324

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

Protocol Primers

Primer 5' Label Sequence 5' --> 3' 3' Label Primer Type oIMR0199 - GAT CTT TTC TAT GGA ATA AGG AAT GGA - Transgene oIMR0200 - GAT CCA GTG TGA CCT AGA GAA GAA AT - Transgene oIMR7338 - CTA GGC CAC AGA ATT GAA AGA TCT - Internal Positive Control Forward oIMR7339 - GTA GGT GGA AAT TCT AGC ATC ATC C - Internal Positive Control Reverse

Number of Reactions

Reaction/Components  Reaction Component Volume (µl) Final Concentration Total Volume (µl) ddH2O 4.80 - 4.80 10 X PCR Buffer II (ABI) 1.20 1.00 X 1.20 25 mM MgCl2 0.72 1.50 mM 0.72 2.5 mM dNTP 0.96 0.20 mM 0.96 20 uM oIMR0199 0.60 1.00 uM 0.60 20 uM oIMR0200 0.60 1.00 uM 0.60 5 mM DNA Loading Dye 1.66 0.69 mM 1.66

Cycling  Step # Temp °C Time Note 1 94 3 min - 2 94 30 sec - 3 64 30 sec - 4 72 45 sec 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 4.80 - 4.80 10 X PCR Buffer II (ABI) 1.20 1.00 X 1.20 25 mM MgCl2 0.72 1.50 mM 0.72 2.5 mM dNTP 0.96 0.20 mM 0.96 20 uM oIMR7338 0.60 1.00 uM 0.60 20 uM oIMR7339 0.60 1.00 uM 0.60 5 mM DNA Loading Dye 1.66 0.69 mM 1.66 This genotyping protocol is used for the following strains:   Stock Number Strain Name 002373 B6D2-Tg(MMTVTGFA)29Rjc/J 002459 B6D2-Tg(MMTVTGFA)254Rjc/J 002953 FVB.Cg-Tg(MMTVTGFA)254Rjc/J Genotyping resources and troubleshooting This page contains guidelines, frequently asked questions, and helpful external links to genotyping and PCR resources.

Cycling  None found.