Stock Number: 006999 Strain Name: STOCK Dbttm1Geh Tg(tTALap)5Bjd Tg(tetO-DBT)A1Geh/J

Allele: Tg(tetO-DBT)A1Geh Protocol Name: Tg(tetO-DBT)A1Geh Method: Standard PCR Version: 1.1 Created: 03-July -2008 Updated: 30-October -2009

Notes: This assay will NOT distinguish hemizygous from homozygous transgenic animals. Expected Results: Transgene=239 bp

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

Protocol Primers

Primer 5' Label Sequence 5' --> 3' 3' Label Primer Type oIMR6991 - GCA GGG TTT GAT TGT CCC TA - Transgene oIMR6992 - GGG GAA TGG CCT TAA TTG AT - Transgene

Number of Reactions

Reaction/Components  Reaction Component Volume (µl) Final Concentration Total Volume (µl) ddH2O 4.56 - 4.56 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 oIMR6991 0.30 0.50 uM 0.30 20 uM oIMR6992 0.30 0.50 uM 0.30 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 1.5 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 006999 STOCK Dbttm1Geh Tg(tTALap)5Bjd Tg(tetO-DBT)A1Geh/J Genotyping resources and troubleshooting This page contains guidelines, frequently asked questions, and helpful external links to genotyping and PCR resources.