Plant Molecular Biology 40: 289–296, 1999.
© 1999 Kluwer Academic Publishers. Printed in the Netherlands.
Identiﬁcation of disease response genes expressed in Gossypium hirsutum
upon infection with the wilt pathogen Verticillium dahliae
Melissa K. Hill, Karin J. Lyon and Bruce R. Lyon
Cooperative Research Centre for Sustainable Cotton Production, School of Biological Sciences A12, The
University of Sydney, Sydney NSW 2006, Australia
Received 7 July 1998; accepted in revised form 27 January 1999
Key words: gene expression, Gossypium hirsutum, plant defence response, Verticillium dahliae
Verticillium wilt is a vascular disease of cotton (Gossypium spp.) caused by the fungal pathogen Verticillium
dahliae. To begin to understand the molecular mechanisms of the disease response in cotton cultivars that display
superior wilt tolerance, such as Gossypium hirsutum cv. Sicala V-1, a cDNA library was constructed with mRNA
isolated from root tissue of Sicala V-1, 24 h after inoculation with V. dahliae. The library was screened by a
differential screening technique which was successful in identifying differences in gene expression between unin-
fected and V. dahliae-infected G. hirsutum root tissue. Among the differentially expressed clones, 51% represented
up-regulated genes which had the potential to be involved in the defence response of G. hirsutum. The temporal
expression patterns of nine suspected defence response genes were examined by northern blot analysis at several
time intervals after inoculation with V. dahliae. The rapid increase in mRNA transcripts corresponding to each
of these clones upon infection suggests a role for these genes in the defence response of G. hirsutum. Genes
not previously associated with the defence response of the cotton plant, such as those for a 14-3-3-like protein and
pathogenesis-related (PR) proteins, have been identiﬁed together with presumably novel genes, for which a deﬁnite
function could not be ascribed.
Strains of the pathogenic fungus Verticillium dahliae
Kleb. are the causal agents of a vascular disease of
cotton (Gossypium spp.) known as Verticillium wilt.
The fungus colonises the root surface, then hyphae
penetrate the root cortex, the endodermis and the
xylem. Systemic invasion of the plant results when
successive generations of conidia are produced and
then transported by the xylem’s transpiration stream
to the aerial parts of the plant (Garas et al., 1986).
The cotton plant deploys a coordinated response that
is speciﬁcally directed towards the rapid containment
of the pathogen to halt the colonisation of the vascu-
lar system. Accordingly, the success of the defence
response appears to hinge on the ability of the cot-
ton plant to rapidly negate the systemic spread of V.
The nucleotide sequence data reported will appear in the NCBI
database dbEST under the accession numbers listed in Table 1.
dahliae with a combination of constitutive and in-
duced defence mechanisms (Bell, 1994). Research
into the defence mechanisms employed by the cot-
ton plant has thus far been concentrated on the role
of antibiotic ﬂavonoids and terpenoids, and structural
defences such as vascular occlusion. Recent advances
in molecular techniques have enabled the investigation
of genes underlying the observed defence response
in Gossypium spp. Characterisation of 3-hydroxy-3-
methylglutaryl coenzyme A reductase (HMGR), the
key regulatoryenzyme of the mevalonate pathway that
synthesises terpenoid phytoalexins, found that tran-
scripts were induced in cultivars of both G. hirsutum
and G. barbadense upon V. dahliae stem inoculation
(Joost et al., 1995). Several enzymes of phenyl-
propanoid metabolism that are thought to be involved
in the synthesis of lignin, for cell wall strengthening
(Smit and Dubery, 1997) and the pathogenesis-related
(PR) proteins chitinase and β-1,3-glucanase (Dubery