Protocol 27394: Standard PCR Assay - Grm7<Tg(SMN2)89Ahmb> Alternate1
Version 1.2

Notes

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

Mutant = 107 bp
Heterozygote = 107 bp and 292 bp
Wild type = 292 bp

The integration site of this Tg is known to be on Mouse Chr 6 so the standard PCR assay can distinguish hemi from hom.

JAX Protocol

Protocol Primers

Primer 5' Label Sequence 5' → 3' 3' Label Primer Type Reaction Note
23139 GAA ATA TGT AGC CTC CCA AAT TGT Common A Grm7
23140 TGC AAA TGT AGC CTA AAG CTG GT Wild type Reverse A Grm7
23141 TGT ACC TAC GTT CAG AAG ACA GAC A Transgene Reverse A SMN2 upstream

Reaction A

Component Final Concentration
ddH2O
Kapa 2G HS buffer 1.30 X
MgCl2 2.60 mM
dNTP KAPA 0.26 mM
23139 0.50 uM
23140 0.50 uM
23141 0.50 uM
Glycerol 6.50 %
Dye 1.00 X
Kapa 2G HS taq polym 0.03 U/ul
DNA

Cycling

Step Temp °C Time Note
1 94.0 --
2 94.0 --
3 65.0 -- -0.5 C per cycle decrease
4 68.0 --
5 -- repeat steps 2-4 for 10 cycles (Touchdown)
6 94.0 --
7 60.0 --
8 72.0 --
9 -- repeat steps 6-8 for 28 cycles
10 72.0 --
11 10.0 -- hold
JAX uses a very high speed Taq (~1000 bp/sec), use cycling times recommended for your reagents.
JAX uses a 'touchdown' cycling protocol and therefore has not calculated the optimal annealing temperature for each set of primers.

Strains Using This Protocol

Stock Number Strain Name
007222 B6.Cg-Grm7Tg(SMN2)89Ahmb Smn1tm1Msd Tg(SMN1*A2G)2023Ahmb/J
006964 B6.Cg-Grm7Tg(SMN2)89Ahmb Smn1tm1Msd Tg(SMN2*delta7)4299Ahmb/J
006773 B6.Cg-Grm7Tg(SMN2)89Ahmb Smn1tm1Msd/J
006385 B6;129P2-Syt1tm3Sud/J
007968 FVB.Cg-Grm7Tg(SMN2)89Ahmb Smn1tm1Msd Tg(SMN1*A2G)2023Ahmb/2J
005026 FVB.Cg-Grm7Tg(SMN2)89Ahmb Smn1tm1Msd Tg(SMN1*A2G)2023Ahmb/J
007952 FVB.Cg-Grm7Tg(SMN2)89Ahmb Smn1tm1Msd Tg(SMN2*delta7)4299Ahmb/2J
031906 FVB.Cg-Grm7Tg(SMN2)89Ahmb Smn1tm1Msd Tg(SMN2-SMN1*T274I)5Ahmb/J
007949 FVB.Cg-Grm7Tg(SMN2)89Ahmb Smn1tm1Msd/2J
005024 FVB.Cg-Grm7Tg(SMN2)89Ahmb Smn1tm1Msd/J
030214 FVB.Cg-Grm7Tg(SMN2)89Ahmb Smn1tm1Msd Tg(Prnp-PLS3)14Ahmb Tg(SMN2*delta7)4299Ahmb/J
030970 FVB.Cg-Mapk10tm1Flv Grm7Tg(SMN2)89Ahmb Smn1tm1Msd Tg(SMN2*delta7)4299Ahmb/LganJ
031907 STOCK Grm7Tg(SMN2)89Ahmb Tg(SMN2-SMN1*Q282A)2Ahmb Smn1tm1Msd/J
032003 STOCK Grm7Tg(SMN2)89Ahmb Smn1tm1Msd Tg(Hlxb9-GFP)1Tmj Tg(SMN2*delta7)4299Ahmb/J
006570 STOCK Grm7Tg(SMN2)89Ahmb Smn1tm1Msd Tg(Hlxb9-GFP)1Tmj/J
008212 STOCK Grm7Tg(SMN2)89Ahmb Smn1tm1Msd Tg(Prnp-SMN)92Ahmb/J
018916 STOCK Grm7Tg(SMN2)89Ahmb Smn1tm1Msd Tg(SMN1-SMN2*)16Cll/CllJ
031908 STOCK Grm7Tg(SMN2)89Ahmb Smn1tm1Msd Tg(SMN2-SMN1*D44V)10Ahmb/J
008783 STOCK Grm7Tg(SMN2)89Ahmb Smn1tm3(SMN2/Smn1)Mrph Tg(CAG-cre/Esr1*)5Amc Tg(SMN2*delta7)4299Ahmb/J
007951 STOCK Grm7Tg(SMN2)89Ahmb Smn1tm3(SMN2/Smn1)Mrph Tg(SMN2*delta7)4299Ahmb/J
007022 STOCK Mnx1tm4(cre)Tmj Grm7Tg(SMN2)89Ahmb Smn1tm1Msd Tg(SMN2*delta7)4299Ahmb/J
017600 STOCK Tg(tetO-SMN2,-luc)#bAhmb/J
22 strains use this protocol