Protocol 30027: QPCR Assay - Tg(SMN2*)

FVB.Cg-Grm7^{Tg(SMN2)89Ahmb} Smn1^tm1Msd Tg(SMN2*delta7)4299Ahmb/J

Stock No: 005025

Version 10.0

Notes

Taqman qPCR protocols are run on an ABI 7500, 7700, 7900 or the Roche LightCycler 480. Use an appropriate instrument-specific Fuorophore/Quencher combination.  The transgene genotype is determined by comparing ΔCt values of each unknown sample against known homozygous and hemizygous controls, using appropriate endogenous references.

The genotyping protocol(s) presented here have been optimized for reagents and conditions used by The Jackson Laboratory (JAX). To genotype animals, JAX recommends researchers validate the assay independently upon receipt of animals into their facility. Reaction cycling temperature and times may require additional optimization based on the specific genotyping reagents used.

Expected Results

JAX Protocol

Protocol Primers

Primer	5' Label	Sequence 5' → 3'	3' Label	Primer Type	Reaction	Note
13739	Fluorophore-1	CTT CTG GAC CAC CAA TAA TTC CCC CAC C	Quencher-1	Tg Probe
oIMR1544		CAC GTG GGC TCC AGC ATT		Internal Positive Control Forward	A
oIMR3580		TCA CCA GTC ATT TCT GCC TTT G		Internal Positive Control Reverse	A
oIMR3675		TGC TGG CTG CCT CCA TTT		Transgene	A
oIMR3676		GCA TCA TCA AGA GAA TCT GGA CAT		Transgene	A
TmoIMR0105	Fluorophore-2	CCA ATG GTC GGG CAC TGC TCA A	Quencher-2	IC Probe

Reaction A

Component	Final Concentration
ddH2O
Kapa Probe Fast QPCR	1.00 X
oIMR1544	0.40 uM
oIMR3580	0.40 uM
oIMR3675	0.40 uM
oIMR3676	0.40 uM
Tg Probe	0.15 uM
IC Probe	0.15 uM
DNA

Cycling

Step	Temp °C	Time	Note
1	95.0	--
2	95.0	--
3	60.0	--	repeat steps 2-3 for 40 cycles

JAX uses a very high speed Taq (~1000 bp/sec), use cycling times recommended for your reagents.

Strains Using This Protocol

Stock Number	Strain Name
006385	B6;129P2-Syt1^tm3Sud/J
006386	B6;129P2-Syt1^tm5Sud/J
008209	FVB.Cg-Grm7^Tg(SMN2)89Ahmb Smn1^tm1Msd Tg(ACTA1-SMN)69Ahmb/J
005025	FVB.Cg-Grm7^Tg(SMN2)89Ahmb Smn1^tm1Msd Tg(SMN2*delta7)4299Ahmb/J
030214	FVB.Cg-Grm7^Tg(SMN2)89Ahmb Smn1^tm1Msd Tg(Prnp-PLS3)14Ahmb Tg(SMN2*delta7)4299Ahmb/J
030970	FVB.Cg-Mapk10^tm1Flv Grm7^Tg(SMN2)89Ahmb Smn1^tm1Msd Tg(SMN2*delta7)4299Ahmb/LganJ
031909	FVB.Cg-Tg(SMN2*A111G)588Ahmb Tg(SMN2-SMN1*Q282A)2Ahmb Smn1^tm1Msd/J
008203	STOCK Grm7^Tg(SMN2)89Ahmb Smn1^tm1Msd Tg(ACTA1-SMN)63Ahmb/J
032003	STOCK Grm7^Tg(SMN2)89Ahmb Smn1^tm1Msd Tg(Hlxb9-GFP)1Tmj Tg(SMN2*delta7)4299Ahmb/J
018916	STOCK Grm7^Tg(SMN2)89Ahmb Smn1^tm1Msd Tg(SMN1-SMN2*)16Cll/CllJ
008783	STOCK Grm7^Tg(SMN2)89Ahmb Smn1^tm3(SMN2/Smn1)Mrph Tg(CAG-cre/Esr1*)5Amc Tg(SMN2*delta7)4299Ahmb/J
007951	STOCK Grm7^Tg(SMN2)89Ahmb Smn1^tm3(SMN2/Smn1)Mrph Tg(SMN2*delta7)4299Ahmb/J
017596	STOCK Gt(ROSA)26Sor^tm1.1(rtTA,EGFP)Nagy Grm7^Tg(SMN2)89Ahmb Smn1^tm1Msd Tg(SMN2*delta7)4299Ahmb Tg(tetO-SMN2,-luc)#aAhmb/J
13 strains use this protocol