Root colonization of oil palm (Elaeis guineensis Jacq.) Using GFP-Expressing Ganoderma boninense and effects of lignin on disease progression
Oil palm is the world’s most efficient oil-bearing tree. Major diseases impeding the oil palm productivity have been caused by fungi, particularly Ganoderma boninense, the causal agent of basal stem rot (BSR). Visible symptoms can only be observed nearing the plant death stage while early penetra...
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| Format: | Thesis |
| Language: | English |
| Published: |
2016
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| Subjects: | |
| Online Access: | http://psasir.upm.edu.my/id/eprint/69900/1/ITA%202016%204%20IR.pdf |
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| Summary: | Oil palm is the world’s most efficient oil-bearing tree. Major diseases impeding the oil
palm productivity have been caused by fungi, particularly Ganoderma boninense, the
causal agent of basal stem rot (BSR). Visible symptoms can only be observed nearing
the plant death stage while early penetration and infection strategy remain cryptic due
to the fungal hyaline nature. The underlying principles on how Ganoderma penetrates
and infects oil palm roots are unknown. Therefore, a tagged G. boninense harbouring
GUS-GFP fusion gene would ideally serve as a tool to unravel early pathogenesis of G.
boninense. Lignin, a heterogeneous complex polymer is poorly understood during BSR
development and thus, assessments of lignin content and composition in different
planting materials and during the disease development were performed. In addition,
enzyme activity and gene expression of key components in the phenylpropanoid
pathway were investigated. Lignin content and composition were screened in oil palm
lines with differential tolerance to BSR. Both parameters were associated to growth
factors (height, weight and girth) and micronutrients depositions were measured using
X-Ray Fluorescence (XRF). Efficient Agrobacterium-mediated transformation protocol
was established via optimization of several parameters; Agrobacterium strain
(LBA4404, GV3101, EHA101 and EHA105), explants (mycelia, spore and protoplast),
vir gene induction period and modification of binary vector. The transformant was
utilized to discern early stage colonization of BSR using the confocal microscopy.
Glass-house trial on BSR development was performed to evaluate enzyme activities
and gene expression of the following defence genes; phenylalanine ammonia lyase
(PAL), cinnamyl alcohol dehydrogenase (CAD) and peroxidase (POD) using enzyme
assay and quantitative real-time PCR respectively. Lignin content and composition
were significantly different among the oil palm seedlings with different tolerance to
BSR. The susceptible and intermediate lines showed significantly higher lignin content
in comparison to the tolerant line, while lignin composition denoted as S/G ratio was
higher in tolerant line in comparison to both susceptible and intermediate lines.
Apparent lignin accumulation was supported by micronutrients deposition which
comprised copper, silicon, titanium and sulphur. A successful transformation system
was developed for G. boninense using protoplast and Agrobacterium strain LBA 4404.
The binary vector pCAMBIA 1304, modified to harbour GPD fungal promoter from plasmid p416 improved the expression of GUS-GFP fusion protein. Colonization
pattern was initiated with active differentiation of the tagged G. boninense into microhyphae.
The needle-like structure was able to penetrate the epidermis layer randomly
and progressed longitudinally into exodermis and cortex region. Induced lignification
during defense showed great participation from both PAL and CAD genes and
enzymes. The S/G ratio increased significantly in the induced lignin as compared to
constitutive lignin indicated alterations employed by host as part of their defense
strategy during Ganoderma infection. Low lignin content supported growth without
compromising oil palm biomass while creating an avenue for greater proportion of
induced lignin which consists of S monomer during G. boninense infection. The
findings can be adopted in oil palm breeding strategies aimed to produce resistant
planting materials. |
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