Stock Number: 008075 Strain Name: B6CBA(FVB)-Tg(Prnp-TBP*)105Xjl/J

Allele: (Prnp-TBP*)105Xjl Protocol Name: (Prnp-TBP*)105Xjl Method: Standard PCR Version: 1.0 Created: 22-June -2009 Updated: 24-September-2009

Notes: AM Bufer:  670 mM TrisHCl pH 8.8, 166 mM (NH4)2SO4, 20 mM MgCl2, 1.7 mg/ml BSA, 10 mM 2-mercaptoethanol (Add 2-mercaptoethanol to 10 mM just prior to use. Do not store with 2-mercaptoethanol in buffer.) Size on ABI3730. Expected Results: Transgene = 545 bp C57BL/6J = 276 bp Human = 341 bp

Protocol Primers

Primer 5' Label Sequence 5' --> 3' 3' Label Primer Type TBP-A629 - AGA AGC TGG TGT GGC AGG AGT GAT - - TBP-S365 6-FAM CCA CAG CCT ATT CAG AAC ACC - -

Number of Reactions

Reaction/Components  Reaction Component Volume (µl) Final Concentration Total Volume (µl) ddH2O 6.91 - 6.91 10 X AM Buffer 1.31 1.09 X 1.31 2.5 mM dNTP 1.04 0.22 mM 1.04 6.6 uM TBP-S365 with 5 0.24 0.13 uM 0.24 6.6 uM TBP-A629 0.24 0.13 uM 0.24 DMSO 0.60 - 0.60 5 M Betaine 0.60 0.25 M 0.60 5 U/ul 5prime HotMaster Taq 0.06 0.03 U/ul 0.06 DNA (crude prep diluted 1:1 in water) 1.00 - 1.00

Cycling  Step # Temp °C Time Note 0 0 - Put samples into preheated block. 1 96 3 min - 2 96 45 sec - 3 59.9 45 sec - 4 72 1 min repeat steps 2-4 for 38 cycles 5 72 5 min - 6 10 - hold

This genotyping protocol is used for the following strains:   Stock Number Strain Name 008075 B6CBA(FVB)-Tg(Prnp-TBP*)105Xjl/J Genotyping resources and troubleshooting This page contains guidelines, frequently asked questions, and helpful external links to genotyping and PCR resources.