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This assay will NOT distinguish hemizygous from homozygous transgenic animals.
Transgene = 187 bp
Internal positive control = 415 bp
Primer | 5' Label | Sequence 5' → 3' | 3' Label | Primer Type | Reaction | Note |
---|---|---|---|---|---|---|
21238 | CTG TCC CTG TAT GCC TCT GG | Internal Positive Control Forward | A | |||
21239 | AGA TGG AGA AAG GAC TAG GCT ACA | Internal Positive Control Reverse | A | |||
24712 | CTT GGA ATA AGG CCG GTG T | Transgene Forward | A | |||
24713 | AGC TTC CAG AGG AAC TGC TTC | Transgene Reverse | A |
Component | Final Concentration |
---|---|
ddH2O | |
Kapa 2G HS buffer | 1.30 X |
MgCl2 | 2.60 mM |
dNTP KAPA | 0.26 mM |
21238 | 0.50 uM |
21239 | 0.50 uM |
24712 | 0.50 uM |
24713 | 0.50 uM |
Glycerol | 6.50 % |
Dye | 1.00 X |
Kapa 2G HS taq polym | 0.03 U/ul |
DNA |
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 |
Stock Number | Strain Name |
---|---|
031824 | B6(FVB)-Tlr4tm1.1Gbrt/J |
033394 | B6.129-Ntsr2tm1.1(cre)Giml/J |
031559 | B6.Cg-Crhtm1.1(flpo)Bsab/J |
030211 | B6.Cg-Npytm1.1(flpo)Hze/J |
017580 | C.129S4(B6)-Ifngtm3.1Lky/J |
031289 | STOCK Lmx1btm2(cre/ERT2)Rjo/J |
029803 | STOCK Mki67tm2.1(cre/ERT2)Cle/J |
029429 | STOCK Or8a1tm26Mom Tg(Or8a1,-MAPT/mCherry)5MGaFeins/J |
006677 | STOCK Or8a1tm28(cre)Mom/MomJ |
036906 | STOCK Tmem178btm1c(KOMP)Wtsi Tmem178tm1.1Sud/J |
026703 | STOCK Vsx1tm2Mci/ChowJ |
026707 | STOCK Tg(Grm6-lacZ)1Chow/J |
018281 | STOCK Tg(Wnt7a-EGFP/cre)#Bhr/Mmjax |
13 strains use this protocol |