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L4440 double-T7 vector inside HT115 RNase-deficient E. coli. A fragment from the gene of interest is amplified by PCR and cloned into the L4440 double-T7 vector, which has two T7 promoters in inverted orientation flanking the multiple cloning site [4]. Cloned plasmids are transformed into HT115(DE3), an RNase III-deficient E. coli strain with IPTG-inducible expression of T7 polymerase (L. Timmons and A. Fire, personal communication).
