Stock Number: 008215 Strain Name: (C57BL/6-Tg(TRAMP)8247Ng/J X FVB/NJ)F1/J

Allele: Tg(TRAMP)-Tg2 Protocol Name: Tg(Tramp2) Method: Standard PCR Version: 1.1 Created: 15-April -2009 Updated: 24-September-2009

Notes: This assay will NOT distinguish hemizygous from homozygous transgenic animals. Expected Results: Transgene = ~650 bp Internal positive control = 324 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 oIMR7084 - GCG CTG CTG ACT TTC TAA ACA TAA G - Transgene oIMR7085 - GAG CTC ACG TTA AGT TTT GAT GTG T - 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 3.87 - 3.87 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 oIMR7084 0.50 0.83 uM 0.50 20 uM oIMR7085 0.50 0.83 uM 0.50 20 uM oIMR7338 0.10 0.17 uM 0.10 20 uM oIMR7339 0.10 0.17 uM 0.10 5 mM DNA Loading Dye 1.66 0.69 mM 1.66 5 U/ul Taq DNA Polymerase 0.15 0.06 U/ul 0.15 DNA 2.00 - 2.00

Cycling  Step # Temp °C Time Note 1 94 3 min - 2 94 30 sec - 3 62 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 003135 C57BL/6-Tg(TRAMP)8247Ng/J 008215 (C57BL/6-Tg(TRAMP)8247Ng/J X FVB/NJ)F1/J 308215 C57BL/6-Tg(TRAMP)8247Ng/J X FVB/NJ)F1 Genotyping resources and troubleshooting This page contains guidelines, frequently asked questions, and helpful external links to genotyping and PCR resources.