Stock Number: 002287 Strain Name: B6.129S7-Ifngtm1Ts/J

Allele: Ifngtm1Ts Protocol Name: Ifngtm1Ts Method: Standard PCR Version: 2.0 Created: 01-October -2009 Updated: 10-November -2009

Expected Results: Mutant = ~500 bp Heterozygote = ~210 bp and ~500 bp Wild type = ~210 bp

Gel Information: Separated by gel electrophoresis on a 1.5% agarose gel.

Protocol Primers

Primer 5' Label Sequence 5' --> 3' 3' Label Primer Type oIMR6218 - CCT TCT ATC GCC TTC TTG ACG - - oIMR8284 - AGA AGT AAG TGG AAG GGC CCA GAA G - - oIMR8285 - AGG GAA ACT GGG AGA GGA GAA ATA T - -

Number of Reactions

Reaction/Components  Reaction Component Volume (µl) Final Concentration Total Volume (µl) ddH2O 3.36 - 3.36 25 mM MgCl2 0.96 2.00 mM 0.96 2.5 mM dNTP 0.96 0.20 mM 0.96 20 uM oIMR8284 0.20 0.33 uM 0.20 20 uM oIMR8285 0.20 0.33 uM 0.20 5 mM DNA Loading Dye 1.66 0.69 mM 1.66 DNA 2.00 - 2.00 20 uM oIMR6218 1.40 2.33 uM 1.40 10 X High Fidelity Buffer 1.20 1.00 X 1.20 5 U/ul High Fidelity Platinum Taq 0.06 0.03 U/ul 0.06

Cycling  Step # Temp °C Time Note 1 94 3 min - 2 94 30 sec - 3 61 1 min *35 cycles 4 72 1 min - 5 72 2 min - 6 10 - hold

This genotyping protocol is used for the following strains:   Stock Number Strain Name 002286 C.129S7(B6)-Ifngtm1Ts/J 002287 B6.129S7-Ifngtm1Ts/J 002575 NOD.129S7(B6)-Ifngtm1Ts/DvsJ 008228 C3.129S7(B6)-Ifngtm1Ts/J Genotyping resources and troubleshooting This page contains guidelines, frequently asked questions, and helpful external links to genotyping and PCR resources.