ThepET-20bvectorcarriesanN-terminalpelBsignalsequenceforpotentialperiplasmiclocalization,plusoptionalC-terminalHis•Tag sequence.Uniquesitesareshownonthecirclemap.NotethatthesequenceisnumberedbythepBR322convention,sotheT7expressionregionisreversedonthecircularmap.Thecloning/expressionregionofthecodingstrandtranscribedbyT7RNApolymeraseisshownbelow.Thef1originisorientedsothatinfectionwithhelperphagewillproducevirionscontainingsingle-strandedDNAthatcorrespondstothecodingstrand.Therefore,single-strandedsequencingshouldbeperformedusingtheT7terminatorprimer. |