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Aarons PCR genotyping

NF1flox (480 bp band, flox = 350 bp band)

Yuan10 5' ACC TCT CTA GCC TCA GGA ATG A 3'
Yuan11 5' CTT CAG ACT GAT TGT TGT ACC TGA 3'
Yuan17 5' TGA TTC CCA CTT TGT GGT TCT AAG 3'

NF1KO (wt = 194 band, ko = 340 bp band)

NF31a 5 GTA TTG AAT TGA AGC ACC TTT GTT TGG 3
NEOTKP 5 GCG TGT TCG AAT TCG CCA ATG 3
NF31b 5 CTG CCC AAG GCT CCC CCA G 3

Tie2-Cre* (cre+ = 450 bp band)

Tie2creF 5 CGC ATA ACC AGT GAA ACA GCA TTG C 3
Tie2creR 5 CCC TGT GCT CAG ACA GAA ATG AGA 3

*these primers are specific for tie2-cre and do not work for other cre transgenes.

 

Use above primers in standard PCR mix and run program NewSP
Run PCR sample on 1.5% agarose gel

1. 94°C 2 minutes
2. 94°C 1 minute
3. 60°C 1 minute
4. 72°C 1 minute
5. GOTO step #2 35 times
6. 72°C 5 minutes
7. END

 

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