To efficiently fight against virus causing diseases vaccines against them are needed, which cause immunoreactions in organism. One of modern vaccine types is chimeric vaccine – infection non-causing viruses or nucleic acid non-containing virus-like particles (VLP) carrying on their capsids foreign virus antigens.

The aim of my course work is to acquire gene-engineering methods and carry out the experiments to create a cDNA vector based on monocot plant virus suitable for the future preparation of new chimeric plant viruses – vaccine candidates.

My course work has been worked out from February 2002 till September 2002 in Latvian University Biomedicine study and research centre in the frame of project “Plant sobmeoviruses vectors for the construction of chimeric vaccines” (Nr. 01.0223) under supervision of Andris Zeltiņš. The main topic of this group are studies on chimeric vaccines from plants and virus like particles from virus capsids in microorganisms, using as a model Cocksfoot mottle virus (CfMV) and Rice yellow mottle virus (RYMV).

In this work and in my bachelor work in future several cDNA vector constructions will be prepared, which will be tested on their ability to infect oat plants. In case of success the best cDNA vector will be used for creating of plant virus based vaccine candidates.

In the frame of this work plant virus vector candidate containing two Cauliflower mosaic virus (CaMV) 35S promoters and CfMV cDNA has been constructed.

For cloning 35S sequences before CfMV cDNA a suitable restriction site directly before CfMV cDNA was inserted using PCR mutagenesis. The cDNA identity of the fragment amplified by PCR was verified by sequencing.

For the constructing of cDNA vector two strategies has been chosen. The first construction was prepared by subcloning of CfMV cDNA in a vector containing two 35S promoters. Unfortunately this construction was structurally unstable. For this reason similar construction was prepared in a different way: in the plasmid bearing the CfMV cDNA two 35S promoter sequences were subcloned. For this construction no structural unstability has been observed as seen from restriction analysis experiments. This CfMV cDNA vector will be used in the plant experiments in my future work.