Stock Number: 002496 Strain Name: BALB/c-Il4tm2Nnt/J

Allele: Il4tm2Nnt Protocol Name: Il4tm2Nnt Method: Separated PCR Version: 1.1 Created: 27-November -2000 Updated: 08-March -2012

Expected Results: Mutant = 464 bp Heterozygote = 175 bp and 464 bp Wild type = 175 bp

Gel Image

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Gel Information: Separated by gel electrophoresis on a 1.5% agarose gel. Do not pool pcr products.

Protocol Primers

Primer 5' Label Sequence 5' --> 3' 3' Label Primer Type oIMR0280 - GTG AGC AGA TGA CAT GGG GC - Common oIMR0281 - CTT CAA GCA TGG AGT TTT CCC - Wild type oIMR0282 - GCC GAT TGT CTG TTG TGC CCA G - Mutant

Number of Reactions

Reaction/Components  Reaction Component Volume (µl) Final Concentration Total Volume (µl) ddH2O 4.56 - 4.56 10 X AB PCR BufferII 1.20 1.00 X 1.20 25 mM MgCl2 0.96 2.00 mM 0.96 2.5 mM dNTP 0.96 0.20 mM 0.96 20 uM oIMR0280 0.30 0.50 uM 0.30 20 uM oIMR0282 0.30 0.50 uM 0.30 5 mM DNA Loading Dye 1.66 0.69 mM 1.66 5 U/ul Taq DNA Polymerase 0.06 0.03 U/ul 0.06 DNA 2.00 - 2.00

Cycling  Step # Temp °C Time Note 1 94 2 min - 2 94 30 sec - 3 64 1 min - 4 72 1 min repeat steps 2-4 for 35 cycles 5 72 2 min - 6 10 - hold

Reaction/Components  Reaction Component Volume (µl) Final Concentration Total Volume (µl) ddH2O 4.56 - 4.56 10 X AB PCR BufferII 1.20 1.00 X 1.20 25 mM MgCl2 0.96 2.00 mM 0.96 2.5 mM dNTP 0.96 0.20 mM 0.96 20 uM oIMR0280 0.30 0.50 uM 0.30 20 uM oIMR0281 0.30 0.50 uM 0.30 5 mM DNA Loading Dye 1.66 0.69 mM 1.66 5 U/ul Taq DNA Polymerase 0.06 0.03 U/ul 0.06 DNA 2.00 - 2.00 This genotyping protocol is used for the following strains:   Stock Number Strain Name 002496 BALB/c-Il4tm2Nnt/J 002518 C57BL/6-Il4tm1Nnt/J Genotyping resources and troubleshooting This page contains guidelines, frequently asked questions, and helpful external links to genotyping and PCR resources.

Cycling  Step # Temp °C Time Note 1 94 2 min - 2 94 30 sec - 3 64 1 min - 4 72 1 min repeat steps 2-4 for 35 cycles 5 72 2 min - 6 10 - hold