Master Protocol Details



Notes:	Validation/LC conversion data attached to rec 134991
Expected Results:	Transgene Tm = 87.75°C +/- 0.75°C Internal positive control Tm = 81.5°C+/- 0.5°C

Gel Image

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Gel Information:	Separated by gel electrophoresis on a 3.0% agarose gel.

Protocol Primers

Primer	5' Label	Sequence 5' --> 3'	3' Label	Primer Type
oIMR4110	-	CCC CGT AAT GCA GAA GAA GA	-	Transgene
oIMR4111	-	CTT GGC CAT GTA GGT GGT CT	-	Transgene
oIMR8744	-	CAA ATG TTG CTT GTC TGG TG	-	Internal Positive Control Forward
oIMR8745	-	GTC AGT CGA GTG CAC AGT TT	-	Internal Positive Control Reverse

Reaction/Components

Reaction Component	Volume (µl)	Final Concentration	Total Volume (µl)
ddH2O	2.20	-	2.20
2 X SYBR Green Master Mix (Roche)	6.00	1.00 X	6.00
20 uM oIMR4110	0.60	1.00 uM	0.60
20 uM oIMR4111	0.60	1.00 uM	0.60
20 ul oIMR8744	0.30	0.50 ul	0.30
20 uM oIMR8745	0.30	0.50 uM	0.30
DNA	2.00	-	2.00

Cycling

Step #	Temp °C	Time	Note
1	94	5 min	-
2	94	30 sec	-
3	63	1 min	-
4	72	30 sec	repeat steps 2-4 for 35 cycles
5	72	2 min	-
6	10	-	hold

This genotyping protocol is used for the following strains:

Stock Number	Strain Name
005645	STOCK Tg(CAG-mRFP1)1F1Hadj/J
005884	B6.Cg-Tg(CAG-mRFP1)1F1Hadj/J

Genotyping resources and troubleshooting
This page contains guidelines, frequently asked questions, and helpful external links to genotyping and PCR resources.