A total of 17 different constructs were used for biolistic
transformation. Two plasmids, pMON25496 and pMON25497,
were used to generate transgenic events that were characterized by
Southern analysis (Fig. 2). Both plasmids contain two gene
cassettes that are tandemly joined in a head-to-tail orientation
within a pUC119-derived (Vieira and Messing 1987) high-copy
plasmid backbone. The first transgene cassette of plasmid
pMON25496 contains the rice actin 1 promoter and intron
(McElroy et al. 1990) followed by the Arabidopsis thaliana EPSPS
transit peptide (Shah et al. 1986) linked to the aroA:CP4 gene and
the 30 non-translated region of the nopaline synthase from
Agrobacterium T-DNA (Fraley et al. 1983). The second cassette
is the same as the first except that the rice actin 1 promoter and
intron are replaced by the cauliflower mosaic virus-enhanced 35S
promoter (Kay et al. 1987) and maize heat-shock protein 70 intron
(Brown and Santino 1994). Plasmid pMON25497 is identical to
pMON25496 except that the maize heat-shock protein 70 intron is
replaced by the rice actin 1 intron. These plasmids were digested
with MluI to liberate the fragment containing both aroA:CP4
cassettes, and the vector backbone was purified away by preparative gel electrophoresi