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

Allele: Dbttm1Geh Protocol Name: Dbttm1Geh Method: Standard PCR Version: 2.1 Created: 05-May -2009 Updated: 24-September-2009

Expected Results: Mutant = ~ 500 bp Heterozygote = 210 bp and ~500 bp Wild type = 210 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 oIMR2060 - CAC GAG ACT AGT GAG ACG TG - Mutant oIMR6993 - GGC TGC ACT GTT ACA AGC AT - Wild type Forward oIMR6994 - GTG CTG CAT TCA CAA TGA CC - Common

Number of Reactions

Reaction/Components  Reaction Component Volume (µl) Final Concentration Total Volume (µl) ddH2O 3.36 - 3.36 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 oIMR6993 0.60 1.00 uM 0.60 20 uM oIMR6994 0.60 1.00 uM 0.60 20 uM oIMR2060 0.60 1.00 uM 0.60 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(Cebpb-tTA)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.