Stock Number: 008277 Strain Name: B6.Cg-Tg(tetO-Clockm1Jt)CL57Jt/J

Allele: Tg(tetO-Clockm1Jt)CL57Jt Protocol Name: Tg(tetO-Clock) Method: Standard PCR Version: 1.0 Created: 10-September-2009 Updated: 30-October -2009

Expected Results: TG = 285 bp internal control = 200 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 oIMR8228 - ATA TGC AAG GCC AGG TTG TC - Transgene Forward oIMR8229 - TCT GTG GCA TAC TGG ATG GA - Wild type 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 3.36 - 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 oIMR8228 0.60 1.00 uM 0.60 20 uM oIMR8229 0.60 1.00 uM 0.60 20 oIMR8744 0.30 0.50 0.30 20 uM oIMR8745 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 56 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 008277 B6.Cg-Tg(tetO-Clockm1Jt)CL57Jt/J Genotyping resources and troubleshooting This page contains guidelines, frequently asked questions, and helpful external links to genotyping and PCR resources.