Berglund:classic method

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Classic Transcription Method:

60µl 25mM rNTP (mixed, each rNTP is 25mM)
60µl 5x transcription buffer
30µl yeast pyrophosphatase
60µl T7 polymerase
2µl gamma-CTP
30µl 100mM DTT
20µl PCR or cut plasmid product (~300ng)
37.5µl water

fv= 300µl

2-3 hours at 37°C

Phenol/chloroform/ethanol precipitate

Run product on a 6%-15& denaturing gel (based on size of product)
Cut band out, elute from gel piece.
Ethanol precipitate.

Dry pellet briefly (2 minutes)
Add store as a pellet or add low TE; either way store in the -20 in plexiglass until you need it.

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