Stock Number:008517
Strain Name:C57BL/6-Gt(ROSA)26Sortm3(CAG-MIR17-92,-EGFP)Rsky/J
Gt(ROSA)26Sortm3(CAG-MIR17-92,-EGFP)Rsky
Gt(ROSA)26Sortm3(CAG-MIR17-92,-EGFP)Rsky Alternate1
Separated PCR
1.0
19-March -2012 29-June -2012
Expected Results:

Mutant = 267 bp

Heterozygote = 267 bp and 604 bp

Wild type = 604 bp
Gel Image 
Image Image file
View Full Size
View Full Size
Protocol Primers 
Primer 5' Label Sequence 5' --> 3' 3' Label Primer Type
13840 - CCA AAG TCG CTC TGA GTT GTT ATC - Wild type Forward
13841 - GAG CGG GAG AAA TGG ATA TG - Wild type Reverse
oIMR8918 - ACC TCC CCC TGA ACC TGA AAC A - Mutant Forward
oIMR8919 - CAG TTT TAC AAG GTG ATG TTC TCT G - Mutant Reverse
Reaction/Components 
Reaction Component Volume (µl) Final
Concentration		 Total Volume (µl)
ddH2O 4.55 - 4.55
5 X Kapa 2G HS buffer 2.40 1.00 X 2.40
25 mM MgCl2 0.96 2.00 mM 0.96
10 mM dNTPS-kapa 0.24 0.20 mM 0.24
20 uM 13840 0.30 0.25 uM 0.30
20 uM 13841 0.30 0.25 uM 0.30
5 mM 10x Loading Dye 1.20 0.50 mM 1.20
2.5 ul Kapa 2G HS taq polymerase 0.05 0.01 ul 0.05
DNA 2.00 - 2.00
Cycling 
Step # Temp °C Time Note
1 94 2 min -
2 94 20sec -
3 65 15sec -0.5 C per cycle
4 68 10sec -
5 - - repeat steps 2-4 for 10 cycles
6 94 15sec -
7 60 15sec -
8 72 10sec -
9 - - repeat steps 6-8 for 28 cycles
10 72 1 min -
11 4 - hold
Reaction/Components 
Reaction Component Volume (µl) Final
Concentration		 Total Volume (µl)
ddH2O 4.55 - 4.55
5 X Kapa 2G HS buffer 2.40 1.00 X 2.40
25 mM MgCl2 0.96 2.00 mM 0.96
10 mM dNTPS-kapa 0.24 0.20 mM 0.24
20 uM oIMR8918 0.30 0.25 uM 0.30
20 uM oIMR8919 0.30 0.25 uM 0.30
5 mM 10x Loading Dye 1.20 0.50 mM 1.20
2.5 ul Kapa 2G HS taq polymerase 0.05 0.01 ul 0.05
DNA 2.00 - 2.00
This genotyping protocol is used for the following strains:
 
Stock Number Strain Name
008517 C57BL/6-Gt(ROSA)26Sortm3(CAG-MIR17-92,-EGFP)Rsky/J
370813 C57BL/6 ?Gt(ROSA)26Sortm3(CAG-MIR17-92,-EGFP)Rsky/J x B6.cg-Tg(cdh5-cre)7Mlia/J
370814 (B6.Cg-Gt(ROSA)26Sortm9(CAG-tdTomato)Hze/J x C57BL/6 ?Gt(ROSA)26Sortm3(CAG-MIR17-92,-EGFP)Rsky/J) x (B6.Cg-Tg(Tek-cre)1Ywa/J x C57BL/6-Gt(ROSA)26Sortm3(CAG-MIR17-92,-EGFP)Rsky/J)
912262 B6.Cg-Gt(ROSA)26Sortm9(CAG-tdTomato)Hze/J x C57BL/6 ?Gt(ROSA)26Sortm3(CAG-MIR17-92,-EGFP)Rsky/J
912263 C57BL/6 ?Gt(ROSA)26Sortm3(CAG-MIR17-92,-EGFP)Rsky/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 20sec -
3 65 15sec -0.5 C per cycle
4 68 10sec -
5 - - repeat steps 2-4 for 10 cycles
6 94 15sec -
7 60 15sec -
8 72 10sec -
9 - - repeat steps 6-8 for 28 cycles
10 72 1min -
11 4 - hold