Figure 6.

Molecular analysis of partially rescued rsy3 plants. A, Diagram of the construct used in the partial complementation. The EcoRI fragment (see Fig. 2A) was blunt-end ligated into the NotI site of the T-DNA vector pGHS166N (see “Materials and Methods”). P2 is the mannopine synthase promoter contained within pGSH166N vector. B, Genomic DNA restriction analysis of some F₂ segregants generated from the complementation cross between rsy3/RSY3 and rsy3/RSY3;tE989 lines. Two separate sets of genomic DNAs were digested with EcoRI or NotI and were size fractionated by electrophoresis in a 1% (w/v) agarose gel; the resulting blots were hybridized with the 2.5-kb EcoRI fragment of the RSY3 genomic clone (see Fig. 2A). C, RNA analysis of RSY3 transcripts in different tissues of wild-type (left panel) and in seedlings generated from partially complemented rsy3 mutant (rsy3/rsy3;tE989) plants (right panel). The length of exposure for the left panel was 3 d, and for the right panel was 1 d. D, Portion of the cDNA sequence generated from the reverse transcriptase-PCR analysis using mRNA samples from partially rescued rsy3 mutant (rsy3/rsy3;tE989) plants (see “Materials and Methods”). The series of dots (…) represents internal sequences identical to the sequence shown in Figure 3A (see also accession no. AY077630).