Department of Molecular Biology and Biotechnology

Head of the Department: Ing. Jana Libantová, CSc.

Other members

Ing. Eva Boszorádová, PhD.
Ing. Martin Jopčík, PhD.
RNDr. Radoslava Matúšová, PhD.

PhD students
Ing. Monika Frátriková
Ing. Miroslav Rajninec

Technical assistance
Anna Fábelová

The research activity of the Department of Molecular Biology and Biotechnology is focused on the study of genes and their regulatory sequences with the aim to increase the resistance of agronomically important crops to stress factors. It covers two basic areas:
(1) identification and isolation of genes involved in the biotic and abiotic stress; and the impact analyses when they are overexpressed in the transgenic organism
(2) modulation of the transgene expression in plants with the goal to make them safer for the environment and more acceptable to the consumers

The first part covers the issue of abiotic stress factor influence (heavy metals, drought, excess of nitrogen in the soil)  on defense mechanisms of selected varieties of soybean (Glycine max L.), and wheat (Triticum aestivum L.), respectively. The obtained results have shown that the varieties with the increased potential to accumulate heavy metals can be cheaply and rapidly screened by the method involving SSR markers. Next, the differences in the quantitative and qualitative profile of specific chitinases - defense stress proteins - depend on the type of metal and variety, respectively. Chitinase isoforms identified as responsive to heavy metal stress factors have a potential application in biotechnology phytoremediation programs.

(A) Illustration of protein spectra in plants and spectra of proteins with the chitinolytic activity
(B) Graph showing quantitative and qualitative differences of specific chitinases as a result of heavy metal exposure effect

Within the biotic stress, we focus on the isolation and characterization of genes encoding hydrolytic enzymes, that are expected to increase the defense machinery against pathogens following their introduction into the target organism. For this purpose, the carnivorous plant, Drosera rotundifolia L., was chosen as a genetic source, while the genes involved in the digestive processes represent the subject of our interest. Following their transfer into the plant model organism - tobacco - using the genetic engineering methods  we test whether candidate  genes are able to support recipient plant defense capability.

Isolation gene for hydrolytic enzyme from D. rotundifolia and the construct preparation for expression in plants
(B) Transformation of tobacco leaf explants using agrobacteria
(C) Illustration of inhibition of phytopathogenic fungi growth by protein expracts isolated from transgenic plants

Parasitic plants represent specific type of biotic stress for plants. Several Striga (witchweed), Phelipanche and Orobanche spp. (broomrapes) belong to the group of economically important parasitic weeds of the Orobanchaceae family with negative impact on crop production. Infected crops can be heavily damaged before the parasites emerge above the soil. They attach to the host root and acquire all nutrients and water from the roots of the host. The effective method of their control does not exist due to their specific nature. Therefore the research is focusing on the molecular basis of the interaction between the host plant and the parasite and on key signalling molecules involved in their interactions. The seeds of these parasitic plants will only germinate after induction by a chemical signal exuded from the roots of the host plant. Strigolactones, a new class of carotenoid-derived phytohormones, play a key function in this interaction, although other compounds exuded from the roots of the host may be involved in the interaction. We are interested in the biosynthesis of strigolactones, their detection and functions in the plants and their role in communication with the parasitic plants and other microorganisms.

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Phelipanche ramosa acquires nutrients and water from the roots of tomato (Lycopersicon esculentum)

Within the second area comprising the optimalisation of transgene expression in plants, we focus on the isolation of tissue-specific promoters from the plant source with  the sequenced genome (Arabidopsis thaliana L.). Next we test their ability to maintain a specific expression profile in the transgenic organism. It appears that the CaMV35S promoter, which is in most cases used to drive expression of the selectable marker gene in transgenic plants, is able to change the tissue-specific profile of many promoters to constitutive when they are closely located. Promoters that are resistant to such influence, have a preferential application in plant biotechnology as they are able to maintain the tissue/developmental specific character of the transgene expression.

Figure shows maintenance of pollen and embryo specific expression profile of transgene directed by one of tested promoters in transgenic tobacco plants

Currently, the transgenic plants have a lot of supporters as well as opponents. One of the arguments of the opponents is the question of their risk assessment for humans and the environment. The aim of our research is to test the strategies leading to the increased bio-safety of transgenic plants. One of the promoters, that had been previously isolated, was used for control of the specific splicing of the selectable marker (SM) gene encoding resistance to the antibiotics in the pollen and the embryo, when the transgenic plants from the non-transgenic had been successfully selected. Progeny of such plants contain only the target gene. A key element in terms of efficiency excision event  appears to be the  promoter  that triggers  cre recombinase expression that represents the molecular scissors for splicing of the loxP cassette.

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(A) Scheme of sellection marker gene excision in transgenic plants transformed by vector construct containing self-excision cassette
(B) Testing of functionality SM gene excision was carried out on model plant species for transformation - tobacco