Stock Number: 008596 Strain Name: B6.Cg-Tg(Prnp-Abca1)EHol/J

Allele: Tg(Prnp-Abca1)EHol Protocol Name: Tg(Prnp-Abca1) Method: Standard PCR Version: 1.1 Created: 24-February -2009 Updated: 24-September-2009

Notes: This assay will NOT distinguish hemizygous from homozygous transgenic animals. Expected Results: Transgene = 765 bp Internal positive 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 oIMR8744 - CAA ATG TTG CTT GTC TGG TG - Internal Positive Control Forward oIMR8745 - GTC AGT CGA GTG CAC AGT TT - Internal Positive Control Reverse oIMR9291 - CCT GCA GCA CAA TTT GTC CCT TCC AAG ATC - Forward oIMR9292 - AGG TCA TGG AAC ACA GCC AAC TCC TGA AAG - Reverse

Number of Reactions

Reaction/Components  Reaction Component Volume (µl) Final Concentration Total Volume (µl) ddH2O 3.78 - 3.78 10 X High Fidelity Buffer 1.20 1.00 X 1.20 50 mM MgCl2 0.96 4.00 mM 0.96 2.5 mM dNTP 0.96 0.20 mM 0.96 20 uM oIMR9291 0.60 1.00 uM 0.60 20 uM oIMR9292 0.60 1.00 uM 0.60 20 oIMR8744 0.08 0.13 0.08 20 uM oIMR8745 0.08 0.13 uM 0.08 5 mM DNA Loading Dye 1.66 0.69 mM 1.66 5 U/ul High Fidelity Platinum Taq 0.08 0.03 U/ul 0.08 DNA 2.00 - 2.00

Cycling  Step # Temp °C Time Note 1 94 3 min - 2 94 30 sec - 3 60 45 sec - 4 72 45 sec 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 008596 B6.Cg-Tg(Prnp-Abca1)EHol/J Genotyping resources and troubleshooting This page contains guidelines, frequently asked questions, and helpful external links to genotyping and PCR resources.