FIG. 1.
Construction of minigenomes. The minigenomes were inserted in the transcription vector 2,0 (gray segments) between the T7 RNA polymerase promoter (T7) and the hepatitis delta virus ribozyme (striped segments). (A) Diagram of MBGV-specific minigenome 3M-5M (previously designated 215), consisting of 106 nucleotides of the MBGV leader (white; lMBGV), 668 nucleotides of the CAT gene (black), and 439 nucleotides of the MBGV trailer (white). (B) Diagram of the chimeric minigenome 3M-5E consisting of 106 nucleotides of the MBGV leader (white; lMBGV), the CAT gene (black), and 731 nucleotides of the EBOV trailer (white). (C) Diagram of the chimeric minigenome 3E-5M consisting of 472 nucleotides of the EBOV leader (white), the CAT gene (black), and 439 nucleotides of the MBGV trailer (white). (D) Diagram of the EBOV-specific minigenome 3E-5E, consisting of 472 nucleotides of the EBOV leader (white), the CAT gene (black), and 731 nucleotides of the EBOV trailer (white). Above each scheme are indicated the boundary between the T7 RNA polymerase promoter sequence (underlined) and the 5′ ends of the minigenome (negative-sense orientation) (left side) and the boundary between the hepatitis delta virus ribozyme sequence (underlined) and the 3′ end of the minireplicon (negative-sense orientation) (right side). ClaI, NotI, NdeI, and RsrII, restriction enzymes used for cloning; SalI, restriction site used for linearization of the plasmids for in vitro transcription; TL start, translation start codon of the NP gene. Below, in smaller fonts, it is indicated whether the start site originates from NP of MBGV or EBOV. TC start, transcription start site of the NP gene of MBGV or EBOV; TC stop, transcription stop site of the L gene of MBGV or EBOV. The cleavage site of the ribozyme is symbolized by a pair of scissors.