In order to develop a stable strain of Bacillus megaterium for Keratinase production, the Keratinase gene (KerA) of Bacillus lichiniformis ATCC 53757 and SPlipA gene from plasmid pHIS1525.SPlipA (Bacillus megaterium origin) were PCR amplified and constructed to give a gene cassette called SPK . Then the gene cassette SPK was cloned into the Integration vector, pMUTIN-GFP+ 6192bps and transformed in Bacillus megaterium ATCC 14945. The chromosomal integration was created using homologous single crossing over mechanism. The strong natural promoter from the chromosomal locus of the SPK not only produced the increased extracellular enzyme, but also functions as a non inducible promoter which does not require any inducer for the production of the enzyme in the new integrant strain. The integrant strain was subjected to feather degradation test and found that it could totally digest the feather meal in complete seven days, resulting in a rich fermentation broth.

8 Abstract Human Herpes Virus 6 (HHV-6) consists of two closely related DNA viruses: HHV-6A and HHV-6B. The primoinfection proceeds at an early childhood usually as sixth exanthem disease or without any clinical symptoms. Both HHV-6 viruses are able to integrate to human genome using recombinant mechanisms which is unique compared to other human herpesviruses. The aim of the thesis is to study Ci-HHV-6 in a group of patients with malign disease and in the healthy population. We analysed 812 patients with malign disease and 420 healthy subjects from general population. The Ci-HHV-6 was assessed by real-time PCR, the specific localization of Ci-HHV-6 was determined using fluorescent in situ hybridization (FISH). Using comparative study, we did not identify significant difference between frequency of Ci-HHV-6 in patients with malign disease (1.11%) and healthy subjects (0.95%) (P-value 0.8). Consequently, we proved the heritability of Ci-HHV-6 in affected families. We determined the localization of Ci-HHV-6 to telomeric regions of chromosomes 2 and 18. We studied the production of viral proteins in the subjects with Ci-HHV-6. In this work, we conducted the first epidemiological study of Ci-HHV-6 in the Czech Republic. We also introduced novel methods which contribute to better characterization of this phenomenom.

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