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Use of biochemical methods in the study of plant metabolism with emphasis on the defense system
Bělonožníková, Kateřina ; Ryšlavá, Helena (advisor) ; Luhová, Lenka (referee) ; Plchová, Helena (referee)
As sessile organisms, plants cannot avoid being exposed to various biotic and abiotic stress factors. To defend themselves plants have developed complex and sophisticated defense mechanisms. In agriculture, pesticides are used for plant protection, but they can have undesirable side effects, and therefore attention is being paid to biological control agents. One such agent is Pythium oligandrum, a soil oomycete that has a positive effect on plants at several levels. On the one hand, it secretes elicitins, which stimulate defensive responses and prepare the plant for possible infection (i.e., priming effect), and on the other hand, it produces tryptamine, a precursor of auxin, thus supporting the plant's growth. Furthermore, P. oligandrum directly acts as pathogen mycoparasite or evokes antibiosis, or competition for nutrients and space with other microorganisms. The key research topic of this thesis was the analysis of the biochemical properties of eleven Pythium strains and their impact on plant metabolism. Our results showed that even closely related P. oligandrum strains significantly differ in the content of compounds secreted into the medium, including proteins, amino acids, tryptamine, and hydrolytic enzymes capable of degrading cell walls (endo-β-1,3-glucanase, chitinase, and cellulase),...
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Viral vectors based on tobacco mosaic virus for the expression of recombinant proteins in plants.
Kroupa, Daniel ; Moravec, Tomáš (advisor) ; Plchová, Helena (referee)
(eng): Over the past few decades, plant viruses and vectors derived from them have become not only an integral part of the work in laboratories, but also began to be used commercially for the production of vaccines, drugs, enzymes and other proteins / peptides. One of the most commonly used viruses for the expression of heterologous proteins is tobacco mosaic virus. In this work, I would like to focus on plant viruses and vectors derived therefrom. Compare it with other expression systems used in plants and describe how their benefits could be further used and combined with modern methods of cloning. Key words: viral vector, expression, Tobacco mosaic virus, Cowpea mosaic virus, Golden gate cloning
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Acetyl - CoA carboxylase - evolution and inhibition
Chalupská, Dominika ; Čeřovská, Noemi (advisor) ; Plchová, Helena (referee) ; Vlček, Čestmír (referee)
Abstract
Acetyl-CoA
carboxylase
(ACC)
is
a
key
enzyme
of
fatty
acid
metabolism
with
multiple
isozymes
often
expressed
in
different
eukaryotic
cellular
compartments.
In
agriculture,
inhibitors
of
plastid
ACC
are
used
as
efficient
herbicides
against
grass
weed.
However,
grass
weed
populations
resistant
to
aryloxyphenoxypropionate
(APP)
and
cyclohexanedione
(CHD)
herbicides
represent
a
major
problem
for
sustainable
agriculture.
Using
PCR
and
sequencing
it
was
found
out
that
five
amino
acid
substitutions
in
plastid
ACC
were
correlated
with
herbicide
resistance
of
Avena
sterilis
ssp.
ludoviciana
Durieu
populations
from
the
northern
grain-growing
region
of
Australia:
Trp-1,999-Cys,
Trp-2,027- Cys,
Ile-2,041-Asn,
Asp-2,078-Gly
and
Gly-2,096-Ala.
We
showed,
using
a
yeast
gene- replacement
system,
that
these
single-site
mutations
also
confer
herbicide
resistance
to
wheat
plastid
ACCase:
Asp-2,078-Gly
confers
resistance
to
APPs
and
CHDs,
Trp-2,027-Cys
and
Ile-2,041-Asn
confer
resistance
to
APPs,
and
Trp-1,999-Cys
confers
resistance
only
to
fenoxaprop.
These
mutations
are
very
likely
to
confer
resistance
to
any
grass
weed
species
under
selection
imposed
by
the
extensive
agricultural
use
of
the
herbicides.
...
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Produkce polyklonálních protilátek proti rekombinantnímu proteinu X viru bramboru
Čeřovská, Noemi ; Plchová, Helena ; Moravec, Tomáš ; Filigarová, Marie ; Dědič, P.
The gene encoding the coat protein (CP) of Potato virus X was cloned into expression vector pMPM-4A Omega. PVX CP was expressed in Escherichia coli and was used for raising rabbit polyclonal antibodies. For immunization we purified CP fractions from bacterial lysate either by simple fractionation or by excision from SDS gel. The obtained sera and antibodies were tested for the detection of PVX in laboratory hosts Nicotiana tabacum, cv. Petit Havana SR 1 and N.benthamiana and in various cultivars of natural host Solanum tuberosum by ELISA as well as by Western blots. The antibodies were specific for PVX and are usable for the detection of the broad isolates spectrum of the Potato virus X by ELISA and Western blot.
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