Stock Number: 002878 Strain Name: B6;129-Apobtm1Sgy Apoetm1Unc/J

Allele: Apobtm1Sgy Protocol Name: Apobtm1Sgy Method: Restriction Enzyme Digest Version: 2.1 Created: 30-January -2001 Updated: 24-September-2009

Expected Results: Mutant = 181 bp and 100 bp Heterozygote = 100 bp, 181 bp and 281 bp Wild type = 281 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 oIMR0671 - CTG AAT GCA TCT GAC TGG GAG AGA - Common oIMR0672 - CGG ATA TGA TAC TGT TCA TCA AGA A - Common

Number of Reactions

Reaction/Components  Reaction Component Volume (µl) Final Concentration Total Volume (µl) ddH2O 4.23 - 4.23 10 X AB PCR BufferII 1.20 1.00 X 1.20 25 mM MgCl2 0.72 1.50 mM 0.72 2.5 mM dNTP 0.96 0.20 mM 0.96 20 uM oIMR0671 0.60 1.00 uM 0.60 20 uM oIMR0672 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 0.03 0.01 U/ul 0.03 DNA 2.00 - 2.00

Cycling  Step # Temp °C Time Note 1 94 3 min - 2 94 20 sec - 3 64 30 sec* *-0.5 degrees per cycle 4 72 35 sec repeat steps 2-4 for 12 cycles 5 94 20 sec - 6 58 30 sec - 7 72 35 sec repeat steps 5-7 for 25 cycles 8 72 2 min - 9 10 - hold**

Reaction/Components  Reaction Component Volume (µl) Final Concentration Total Volume (µl) 10 X AB PCR BufferII 1.20 - 1.20 ddH2O 5.68 - 5.68 10 mg/ml BSA 0.12 - 0.12 Spe I 1.00 - 1.00 This genotyping protocol is used for the following strains:   Stock Number Strain Name 002876 B6;129S-Apobtm1Sgy/J 002878 B6;129-Apobtm1Sgy Apoetm1Unc/J Genotyping resources and troubleshooting This page contains guidelines, frequently asked questions, and helpful external links to genotyping and PCR resources.