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Taqman qPCR protocols are run on a real time PCR instrument. Use an appropriate instrument specific Fluorophore/Quencher combination.
Mutant = 116 bp
Heterozygote = 116 bp and 148 bp
Wild type = 148 bp
This assay is capable of distinguishing hemi from hom. Integration site is known to be on mChr2.
>chr2:100921800-100921947 148bp AGCACTAGACTTCTTGTCAGAAACT TTTCTGTCAGTTAGCTTTATGCTTT
Primer | 5' Label | Sequence 5' → 3' | 3' Label | Primer Type | Reaction | Note |
---|---|---|---|---|---|---|
37999 | CTC CTG ACC TCA TGA TCC AC | Transgene Forward | A | FCGRT | ||
38003 | Fluorophore-1 | TCT GCC TCC CAA AGG GGT | Quencher-1 | Tg Probe | FCGRT | |
43800 | AGC ACT AGA CTT CTT GTC AGA AAC T | Wild type Forward | A | mChr2 | ||
43801 | TTT CTG TCA GTT AGC TTT ATG CTT T | Common | A | mChr2 | ||
43802 | Fluorophore-2 | TGG AAG CAA GAG GAG GGT AGA | Quencher-2 | WT Probe | mChr2 |
Component | Final Concentration |
---|---|
Kapa Probe Fast QPCR | 1.00 X |
ddH2O | |
37999 | 0.40 uM |
43800 | 0.40 uM |
43801 | 0.40 uM |
Wt Probe | 0.15 uM |
Mutant Probe | 0.15 uM |
DNA |
Step | Temp °C | Time | Note |
---|---|---|---|
1 | 95.0 | -- | |
2 | 95.0 | -- | |
3 | 60.0 | -- | |
4 | -- | repeat steps 2-3 for 40 cycles | |
5 | 40.0 | -- | Forever |
Stock Number | Strain Name |
---|---|
018441 | B6.Cg-Fcgrttm1Dcr Prkdcscid Tg(FCGRT)32Dcr/DcrJ |
031644 | B6.Cg-Tg(FCGRT)32Dcr Albem12Mvw Fcgrttm1Dcr Prkdcscid /J |
025201 | B6.Cg-Tg(FCGRT)32Dcr Albem12Mvw Fcgrttm1Dcr/MvwJ |
035448 | B6.Cg-Tg(FCGRT)32Dcr Fcgrttm1Dcr Ace2em4Lutzy/J |
029686 | B6.Cg-Tg(FCGRT)32Dcr Fcgrttm1Dcr Ighg1em2(IGHG1)Mvw/MvwJ |
037043 | B6.Cg-Tg(K18-ACE2)2Prlmn Tg(FCGRT)32Dcr Fcgrttm1Dcr/2J |
034902 | B6.Cg-Tg(K18-ACE2)2Prlmn Tg(FCGRT)32Dcr Fcgrttm1Dcr/J |
028615 | NOD.Cg-Fcgrttm1Dcr Prkdcscid Il2rgtm1Wjl Tg(FCGRT)32Dcr/J |
8 strains use this protocol |