plant gene expression machinery. The modifications could include changes to the codons
favored by plants and elimination of cryptic sites that could result in aberrant processing
of transcripts. They almost always include swapping of the upstream and downstream regu-
latory elements with plant sequences to create “chimeric genes” that will be recognized by
the plant transcriptional and translational systems. Such chimeric genes are designed to
express efficiently in plant cells (Fig. 9.1). The first chimeric genes created for plants
were selectable marker genes, and these were directly responsible for the development of
transformation technologies for plants (Fraley et al. 1983; Bevan et al. 1983; Herrera-
Estrella et al. 1983). These include antibiotic- or herbicide-resistant markers.
Figure 9.2 compares the use of a typical conditional positive selectable marker gene
system with newer nonconditional positive selectable marker gene systems being developed
for the selection of transgenic plants. Cells that express the conditional positive selectable
marker gene are supplied with a novel resistance trait, which is the ability to detoxify a toxic
substrate in the tissue culture media used to culture plant cells and regenerate plants. Only
transformed tissue can grow normally and regenerate into plants because the untransformed
cells are prevented from growing or are killed by the substrate. The nonconditional systems
allow transformed cells to be distinguished by alterations in growth and development
Figure 9.1.Functional organization of selectable marker genes and reporter genes on transformation
vectors used to transfer DNA to plant cells. The selectable marker genes are a fundamental component
of the transformation vectors as they are needed for the recovery of transgenic material. The vectors
are used for many purposes, including the study of plant genes and their regulatory elements. Often
the function of genes emerging from genomics studies are unknown and the transgenic plant provides
an experimental model for gaining functional understanding. (a) The gene of interest can be examined
in many ways. The regulatory elements are often found in the noncoding regions of the gene. For
example, the promoter (Px) is found in the 5^0 upstream region and includes a number elements
needed for transcription, including the core promoter and often enhancer or repressor elements.
Some of these elements may also exist in the 3^0 end region. (b) By fusing the 5^0 and 3^0 noncoding
regions to a reporter gene and inclusion of the chimeric gene in the transformation vector, the patterns
of gene regulation can be assessed in transgenic plants. (c) Gain-of-function experiments can be per-
formed by the overexpression of the coding region using a strong constitutive promoters (Pc), such as
the 35Spromoter, and 3^0 ends needed for termination and polyadenylation (3^0 ), such as those from the
nosgene or 35Stranscript. A phenotype in the transgenic plant may reveal function. (d) A mutant
phenotype may also be mimicked by eliminating or reducing the expression of the gene of interest
by creating an antisense transcript in the transgenic plant. (e) This may also be achieved by creating
a vector with inverted repeats of the gene of interest, which may induce gene silencing. In each case,
the selectable marker and the reporter genes serve different purposes.
220 MARKER GENES AND PROMOTERS