Stock Number: 006579 Strain Name: C57BL/6-Tg(Camk2a-Bdnf)A9Stl/J

Allele: Generic SV40 Protocol Name: Generic SV40 Method: Fast MCA Version: 2.0 Created: 31-August -2010 Updated: 31-August -2010

Notes: Melting curve analysis is done using a Roche Light Cycler 480. This assay will NOT distinguish hemizygous from homozygous transgenic animals. Expected Results: Transgene Tm = 80 Internal positive control Tm = 85.5

Protocol Primers

Primer 5' Label Sequence 5' --> 3' 3' Label Primer Type oIMR1201 - GTG AAG GAA CCT TAC TTC TGT GGT G - - oIMR1202 - GTC CTT GGG GTC TTC TAC CTT TCT C - - oIMR9020 - AAG GGA GCT GCA GTG GAG TA - - oIMR9021 - CCG AAA ATC TGT GGG AAG TC - -

Number of Reactions

Reaction/Components  Reaction Component Volume (µl) Final Concentration Total Volume (µl) ddH2O 1.60 - 1.60 2 X KAPA SYBR Fast 6.00 1.00 X 6.00 20 uM oIMR1201 0.60 1.00 uM 0.60 20 uM oIMR1202 0.60 1.00 uM 0.60 DNA 2.00 - 2.00 20 uM oIMR9020 0.60 1.00 uM 0.60 20 uM oIMR9021 0.60 1.00 uM 0.60

Cycling  Step # Temp °C Time Note 1 94 5 min KF 61-45 2 94 15 sec - 3 61 45 S - 5 72 2 min - 6 10 - hold

This genotyping protocol is used for the following strains:   Stock Number Strain Name 003135 C57BL/6-Tg(TRAMP)8247Ng/J 006579 C57BL/6-Tg(Camk2a-Bdnf)A9Stl/J 006662 C57BL/6-Tg(ACTB-MAP2K1*K97M)1Stl/J 008215 (C57BL/6-Tg(TRAMP)8247Ng/J x FVB/NJ)F1/J 008247 B6.Cg-Tg(Ela1-TAg*)289Mjt/J Genotyping resources and troubleshooting This page contains guidelines, frequently asked questions, and helpful external links to genotyping and PCR resources.