Stock Number:017928
Strain Name:FVB/N-Tg(Mpz-cre)26Mes/J
Tg(Mpz-cre)1Mfel
Tg(Mpz-cre)1Mfel

Separated MCA

1.0
28-March -2012 28-March -2012
Notes:

This assay will NOT distinguish hemizygous from homozygous transgenic animals.
Expected Results:

Transgene = 500 bp

Internal positive control = 324 bp

 

Transgene     Tm = 88  Area =  63  plus WT  peak  Tm = 81  Area = 66

Internal positive control  Tm= 82  Area = 61
Gel Image 
Image Image file
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Protocol Primers 
Primer 5' Label Sequence 5' --> 3' 3' Label Primer Type
14203 - CCA CCA CCT CTC CAT TGC AC - Transgene Forward
oIMR7338 - CTA GGC CAC AGA ATT GAA AGA TCT - Internal Positive Control Forward
oIMR7339 - GTA GGT GGA AAT TCT AGC ATC ATC C - Internal Positive Control Reverse
oIMR9377 - ATG TTT AGC TGG CCC AAA TG - Mutant Reverse
Reaction/Components 
Reaction Component Volume (µl) Final
Concentration		 Total Volume (µl)
ddH2O 3.95 - 3.95
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 dNTP KAPA 0.24 0.20 mM 0.24
20 uM 14203 0.30 0.25 uM 0.30
20 uM oIMR9377 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
20 X EvaGreen 0.60 1.00 X 0.60
DNA 2.00 0.00 2.00
Cycling 
Step # Temp °C Time Note
1 94 2 min -
2 94 20sec -
3 65 15sec -1.5 C per cycle
4 68 10sec -
5 - - Go to step 2-4, 10 times
6 94 15sec -
7 50 15sec -
8 72 10sec -
9 - - Go to step 6-8, 28 times
10 72 2 min -
11 10 - hold
Reaction/Components 
Reaction Component Volume (µl) Final
Concentration		 Total Volume (µl)
ddH2O 3.95 - 3.95
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 dNTP KAPA 0.24 0.20 mM 0.24
20 uM oIMR7338 0.30 0.25 uM 0.30
20 uM oIMR7339 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
20 X EvaGreen 0.60 1.00 X 0.60
DNA 2.00 0.00 2.00
This genotyping protocol is used for the following strains:
 
Stock Number Strain Name
017928 FVB/N-Tg(Mpz-cre)26Mes/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 -1.5 C per cycle
4 68 10sec -
5 - - Go to Step 2-4, 10 times
6 94 15sec -
7 50 15sec -
8 72 10sec -
9 - - Go to Step 6-8, 28 times
10 72 2 min -
11 10 - hold