Stock Number:005130
Strain Name:STOCK Gt(ROSA)26Sortm1(Smo/EYFP)Amc/J
Gt(ROSA)26Sortm1(Smo/EYFP)Amc
Gt(ROSA)26Sortm1(Smo/EYFP)Amc
Separated PCR
3.0
09-September-2011 09-September-2011
Expected Results:

Mutant = ~490 bp
Heterozygote = ~490 bp and ~410 bp
Wild type = ~410 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
oIMR0872 - AAG TTC ATC TGC ACC ACC G - Mutant
oIMR3621 - CGT GAT CTG CAA CTC CAG TC - Wild type Forward
oIMR3999 - TGC TCA GGT AGT GGT TGT CG - Mutant
oIMR8546 - GGA GCG GGA GAA ATG GAT ATG - Wild type Reverse
Reaction/Components 
Reaction Component Volume (µl) Final
Concentration		 Total Volume (µl)
5 U/ul KAPATaq DNA Polymerase 0.06 0.03 U/ul 0.06
ddH2O 2.76 - 2.76
2.5 mM dNTP 0.96 0.20 mM 0.96
20 uM oIMR0872 0.60 1.00 uM 0.60
5 mM DNA Loading Dye 1.66 0.69 mM 1.66
DNA 2.00 - 2.00
20 uM oIMR3999 0.60 1.00 uM 0.60
5 X KAPA LR HotStart Buffer 2.40 1.00 X 2.40
25 mM MgCl2 0.96 2.00 mM 0.96
Cycling 
Step # Temp °C Time Note
1 94 3 min -
2 94 30 sec -
3 65 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 2.70 - 2.70
5 X KAPA LR HotStart Buffer 2.40 1.00 X 2.40
25 mM MgCl2 0.96 2.00 mM 0.96
2.5 mM dNTP 0.96 0.20 mM 0.96
20 uM oIMR3621 0.60 1.00 uM 0.60
20 uM oIMR8546 0.60 1.00 uM 0.60
5 mM DNA Loading Dye 1.66 0.69 mM 1.66
2.5 U/ul KAPA LR HotStart Taq 0.12 0.03 U/ul 0.12
DNA 2.00 - 2.00
This genotyping protocol is used for the following strains:
 
Stock Number Strain Name
005130 STOCK Gt(ROSA)26Sortm1(Smo/EYFP)Amc/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 3 min -
2 94 30 sec -
3 67 30 sec -
4 72 1 min repeat steps 2-4 for 35 cycles
5 72 2 min -
6 10 - hold