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Fig. 1 | Virology Journal

Fig. 1

From: A reverse genetics system for avian coronavirus infectious bronchitis virus based on targeted RNA recombination

Fig. 1

Coronavirus genome organization and schematic overview of targeted RNA recombination. a Schematic genome representations of IBV (blue) and MHV (red). The first two-third of the genome is truncated, the structural and accessory genes are drawn to scale. The lengths of the first two-thirds and last one-third of the IBV genome are indicated at the top. The different domains of the spike gene are indicated: ss = signal sequence; ec = ectodomain; tm = transmembrane domain; en = endodomain. PCR amplicons are depicted as black bars drawn to scale above the genomes, with encircled letters referring to the primer sets in Table 3 and Fig. 3. b Stage 1 in targeted RNA recombination: an interspecies chimeric murinized IBV with a MHV spike ectodomain (mIBV) is generated by a single recombination event of IBV genomic RNA with synthetic RNA transcribed from donor plasmid p-mIBV in the 3′-end region of the 1b gene (indicated by a black curved line). Murinized IBV is selected on murine LR7 cells. Plasmid inserts are indicated above p-mIBV, with numbers in black circles referring to the plasmid junctions. c Stage 2 in targeted RNA recombination: a recombinant IBV with the IBV spike gene (rIBV) is recreated by a single recombination event of mIBV with synthetic RNA transcribed from donor plasmid p-IBV. Recombinant IBV is selected on embryonated chicken eggs. d Nucleotide sequences of the plasmid junctions, marked with corresponding numbers in the schematic donor plasmid drawings. Nucleotide sequences are indicated for wild-type IBV and donor plasmids p-IBV and p-mIBV, with restriction enzyme sites in italics, MHV spike gene sequences in lower case, and spike domains (i.e. ss, ec, tm and en) separated by vertical dashes. Stop codons are highlighted in red. Open reading frames (ORFs) are underlined, overlapping ORFs are double-underlined, and ORF translations are indicated as amino acids below the nucleotide sequences if applicable

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