Stock Number: 004363 Strain Name: FVB.Cg-Tg(MMTV-vHaras)SH1Led/J

Allele: Tg(MMTV-vHaras)SH1Led Protocol Name: Tg(MMTV-vHaras)SH1Led Method: Standard PCR Version: 1.1 Created: 10-June -2003 Updated: 24-September-2009

Notes: This assay will NOT distinguish hemizygous from homozygous transgenic animals. Expected Results: Transgene = 825 bp Internal positive control = 200 bp

Gel Image

Image Image file View Full Size

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

Protocol Primers

Primer 5' Label Sequence 5' --> 3' 3' Label Primer Type oIMR1779 - CCC AGG GCT TAA GTA AGT TTT TGG - Transgene Forward oIMR1780 - GGG CAT AAG CAC AGA TAA AAC ACT - Transgene Reverse oIMR8744 - CAA ATG TTG CTT GTC TGG TG - Internal Positive Control Forward oIMR8745 - GTC AGT CGA GTG CAC AGT TT - Internal Positive Control Reverse

Number of Reactions

Reaction/Components  Reaction Component Volume (µl) Final Concentration Total Volume (µl) ddH2O 3.16 - 3.16 10 X AB PCR BufferII 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 oIMR1779 0.60 1.00 uM 0.60 20 uM oIMR1780 0.60 1.00 uM 0.60 20 ul oIMR8744 0.40 0.67 ul 0.40 20 uM oIMR8745 0.40 0.67 uM 0.40 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 1 min - 4 72 1 min 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 004363 FVB.Cg-Tg(MMTV-vHaras)SH1Led/J Genotyping resources and troubleshooting This page contains guidelines, frequently asked questions, and helpful external links to genotyping and PCR resources.