Plant Biotechnology and Genetics: Principles, Techniques and Applications

explants, there would be little chance of recovering transgenic cells without selectable
markers. Selectable marker genes fall into two separate families: those that provide positive
and negative selection (Table 9.1).Positiveselectable marker genes confer a selective
growth advantage on plant cells so that transformed cells can outgrow the nontransformed
cells. The mode of action may also be more severe through the use of toxic chemicals
placed in the growth media. The chemicals are designed to selectively kill nontransformed
cells, whereas transformed cells are allowed to live through the action of detoxification or
resistance mechanisms encoded by the selectable marker genes.Negativeselectable marker
genes encode systems that are toxic to transgenic plant cells and selectively kill them. It is
the positive selection systems that are used in plant biotechnology for the recovery of trans-
genic plants as the transformation process is seldom very efficient.
Conditional positive selectable marker genes are the most important family used in the
recovery of transgenic plants and are key components of most transformation systems.
Indeed, it is difficult to imagine how plant biotechnology could have progressed without
these marker genes. They are usually incorporated into the basic design of transformation
vectors used to insert genes into plant genomes and thus accompany other genes of interest
or genetic elements fused to reporter genes in the transgenic plants (Fig. 9.1). They orig-
inate from a variety of sources, including plant and nonplant species but most are bacterial
in origin and introduce a novel resistance trait (typically antibiotic resistance) into the plant
(Table 9.1). To be effective, they should not interact with specific targets within the plant or
alter signal transduction pathways in a way that changes the plant. If they create such
changes it would be difficult to identify the phenotypes associated with the gene of interest
or the factors affecting their expression. They also act as dominant genetic markers in the
homozygous and hemizygous states.
For expression in plant cells selectable marker genes from bacteria must be extensively
modified because the signals on the bacterial gene will not be correctly recognized by the

TABLE 9.1. Categories of Marker Genes Used in Plants with Selected Examples

Classes of Marker Genes Examples of Genes Source of Genes Selective Agent

Selectable marker genes
Positive
Conditional nptII, neo, aphII Escherichia coliTn 5
(bacteria)

Kanamycin

hpt, hph, aphIV E. coli(bacteria) Hygromycin

bar Streptomyces hygroscopicus

(bacteria)

Phosphinothricin

manA E. coli(bacteria) Mannose

Non-conditional ipt Agrobacterium tumefaciens

(bacteria)

N/Aa

Negative

Conditional codA E. coli(bacteria) 5-Fluorocytosine

Nonconditional barnase Bacillus amyloliquefaciens

(bacteria)

N/A

Non selectable (reporter) genes
Conditional uidA, gusA E. coli(bacteria) MUG, X-gluc
Nonconditional gfp Aequorea victoria(jellyfish) N/A
aNonapplicable.

9.2. DEFINITION OF MARKER GENES 219