Entwicklung eines "plant-based" Impfstoffs gegen den Lungenwurm des Rindes

Masterarbeit aus dem Jahr 2013 im Fachbereich Biologie - Genetik / Gentechnologie, Note: 1,0, Gottfried Wilhelm Leibniz Universität Hannover (Pflanzen Genetik), Sprache: Deutsch, Abstract: Genetically modified plants, which are used in the production of industrial materials or

pharmaceuticals instead of feed and food production, are nowadays referred to as

pharmaceutical plants. Due to genetic modification, these pharmaceutical plants

produce important proteins (vaccines or antibodies) for pharmaceutical use as

enzymes for different applications. In order to produce these proteins/enzymes the

coding DNA are incorporated into the genome of plant where the plants will express

the protein to high level producing low cost molecules compared with other

expression systems like cell culture, hypridoma, fermentas. As well as, when the

target proteins has a lethal effect on E.coli hindering the overexpression. In temperate

climates such as Northern Germany the lungworm (Dictyocaulus viviparous) is one of

the economically most important parasites of cattle. This parasite causes parasitic

bronchitis disease, which occurs in varying intensity and¿in some cases¿can lead to

death of the infected animals.

It was possible to control a lung worm infection with anthelmintics and even in some

countries with a live vaccine, which consists of X-ray attenuated larvae and pasturage

technical measures. This vaccine can only induce a robust immunity in cattle for a

few months. In addition, the production of these vaccines is very costly and suffers

from limited durability. Therefore, an attempt was made in this work to produce

recombinant proteins in high purity and in large amounts by using tobacco plants and

to use these proteins instead of the previously available immunization as a vaccine

with high efficiency.

First, the vectors were constructed harbouring SOD (superoxide dismutase

pGIIMH35S-SOD vector size approximately 6138 bp). and NMT (N-methyl)

(pGIIMH35S NMT-HIS vector size approximately 6322 bp) since expression of SOD

and NMT showed a toxic effect in E. coli, alternative system is needed like plantAbstract

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based expression. Therefore, they were genetically transfered into tobacco plants

using Agrobacterium tumefaciens mediated transformation. Different molecular

biology methods were used to confirm the integration of the genes into gDNA of

tobacco like PCR and RT-PCR using different sets of primers at DNA and RNA level

and functional assays using Basta herbicide and enzymatic assys at protein level.

The desired proteins were detected by western blot and their enzyme activity was

measured. [...]