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. 2009 Jul 9;11(1):33–41. doi: 10.1111/j.1364-3703.2009.00566.x

Figure 1.

Figure 1

(A) Outline of the Citrus tristeza virus (CTV) genome organization (Karasev et al., 1995), with the numbers at the top indicating open reading frames (ORFs) and the boxes the corresponding proteins or catalytic domains within the polyprotein encoded by ORF 1a (see text for further details). (B) Diagram of the T‐DNA from the binary vector pBin19‐sgfp and constructs expressing a 3′‐moiety of p23 and the 3′UTR (3′p23+3′UTR) in sense, antisense or intron‐hairpin format. The 3′p23+3′UTR transgenes are controlled by the double‐enhanced 35S promoter of Cauliflower mosaic virus (CaMV) and the nopaline synthase terminator (nos‐ter), and flanked by the neomycin phosphotransferase II gene (nptII) between the nos promoter (nos‐pro) and nos‐ter, and by the green fluorescent protein gene (sgfp) between the 35S promoter and nos‐ter. A, E, H and N denote ApaI, EcoRI, HindIII and NheI restriction sites, respectively. (C) Southern blot hybridization of nucleic acid preparations from lime plants transformed with the 3′p23+3′UTR antisense construct (lines 24, 23, 16, 10, 17, 5 and 1), and with the empty vector pBin19‐sgfp (EV). (D) Southern blot hybridization of nucleic acid preparations from lime plants transformed with the 3′p23+3′UTR sense construct (lines 14, 15A, 13, 38, 39, 12 and 6), and with the empty vector pBin19‐sgfp (EV). In (C) and (D), DNA aliquots (20 µg) were digested with EcoRI, which cuts once in the T‐DNA near the left border (LB), or with HindIII, which excises the expression cassette. (E) Southern blot hybridization of nucleic acid preparations from lime plants transformed with the 3′p23+3′UTR intron‐hairpin construct (lines 32, 6, 38, 25, 46, 28, 51 and 20), and with the empty vector pBin19‐sgfp (EV). DNA was digested with NheI, which cuts once at the right of the intron‐hairpin cassette, or with HindIII, which excises the expression cassette. Digoxigenin‐labelled DNA probes were derived from the 3′p23+3′UTR fragment for the sense and antisense constructs, and from the Fad2 intron for the intron‐hairpin construct. M, DNA molecular weight marker II (Roche) (C, D and E).