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The directed search of genes for secondary metabolites in streptomycetes.
Bakal, Tomáš ; Najmanová, Lucie (advisor) ; Janderová, Blanka (referee)
Discoveries of new natural antibiotics are now relatively rare, therefore the construction of strains producing hybrid substances seems to be a very promising opportunity to gain new interesting biologically active compounds. This work is part of a larger project focused on the preparation of new biologically active substances derived from the antibiotic lincomycin. Lincomycin is composed of saccharide (MTL) and amino acid (propylhygric acid) moieties condensed by amide bond. Various modifications of amino acid moiety, especially of the side alkyl chain, are known to improve the antibiotic properties of final molecule. The bottleneck of biosynthesis of such modified compounds is the condensing enzyme NDL-synthetase, and especially its A-domain, which, similarly to nonribosomal peptide synthetases (NRPS), specifically recognizes and activates the amino acid precursor. In this work a set of degenerate primers for PCR searching of NRPS A-domains was proposed and the conditions of PCR reaction were optimized. In the first step a collection approximately 800 isolates of soil actinomycetes will serve as a source of genetic information for search of interesting NRPS A-domains, applicable for the construction of hybrid biosynthetic clusters. The isolates of this collection have been also characterized taxonomically...
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The directed search of genes for secondary metabolites in streptomycetes.
Bakal, Tomáš ; Najmanová, Lucie (advisor) ; Janderová, Blanka (referee)
Discoveries of new natural antibiotics are now relatively rare, therefore the construction of strains producing hybrid substances seems to be a very promising opportunity to gain new interesting biologically active compounds. This work is part of a larger project focused on the preparation of new biologically active substances derived from the antibiotic lincomycin. Lincomycin is composed of saccharide (MTL) and amino acid (propylhygric acid) moieties condensed by amide bond. Various modifications of amino acid moiety, especially of the side alkyl chain, are known to improve the antibiotic properties of final molecule. The bottleneck of biosynthesis of such modified compounds is the condensing enzyme NDL-synthetase, and especially its A-domain, which, similarly to nonribosomal peptide synthetases (NRPS), specifically recognizes and activates the amino acid precursor. In this work a set of degenerate primers for PCR searching of NRPS A-domains was proposed and the conditions of PCR reaction were optimized. In the first step a collection approximately 800 isolates of soil actinomycetes will serve as a source of genetic information for search of interesting NRPS A-domains, applicable for the construction of hybrid biosynthetic clusters. The isolates of this collection have been also characterized taxonomically...
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Isolation and characterisation of biologically active substances
Kettnerová, Eliška ; Nesměrák, Karel (advisor) ; Musilová, Adéla (referee)
This Bachelor thesis aims at isolation and partial identification of biologically active substances which are produced by actinomycetes and can be potentially applied in medicine. Cultivation broths of actinomycetes containing their metabolites were purified and pre-concentrated by solid phase extraction. Then, the bioassay of the extracts by Kirby-Bauer test using the sensitive strain Kocuria rhizophila was performed. Biologically active metabolites were analyzed and isolated by ultra- performance liquid chromatography with photo diode array detector. Isolated substances were assayed by mass spectrometry, which yielded relative molecular mass values of the unknown compounds. The values were compared with relative molecular masses of compounds listed in a chemical database, which involves natural products including antibiotics. We revealed that the unknown biologically active substances do not refer to any already discovered compound present in the database suggesting that the unknown compounds may be novel. More mass spectrometry and nuclear resonance experiments have to be carried out in order to elucidate their structure. Key words: actinomycetes, antibiotics, SPE, UPLC, HPLC Subject heading: analysis of secondary metabolites, bioassay test, isolation of biologically active compounds,...
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Potravní zdroje bezobratlých v Ardovské jeskyni a v jeskyni Domica - předběžné výsledky
Nováková, Alena ; Elhottová, Dana ; Krištůfek, Václav ; Lukešová, Alena ; Hill, P. ; Kováč, L. ; Mock, A. ; Luptáčik, P.
The chemistry (organic matter content and pH) of different substrates from the Ardovská and Domica Caves in the Slovak Karst National Park (Slovakia) and the composition of their microbial communities were studied. Differences in organic matter content and microbial communities were found between the substrates from the Domica and Ardovská Caves. A broad spectrum of photoautotrophic microorganisms (algae and cyanobacteria) was found in different cave microhabitats (cave sediment, rotten wood, earthworms casts and isopods faeces), but no algae were detected in the fresh bat droppings and in the intestinal tract of the isopod .i.M. graniger./i.. Microfungi and bacterial colonies play the primary role in transforming bat guano into a food source for saprophagous and microphytophagous invertebrates.
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Použití specifických aktinomycetových PCR primerů k charakterizaci bakteriálních společenstev exkrementů žížal
Hill, P. ; Krištůfek, Václav ; Feijoo, A. M. ; Gallego, G.
Tropical geophageous earthworms stimulate soil bacterial activity in soil that they ingest; it is uncertain if they affect the whole bacterial population or only subgroups. We sampled .i.Martiodrillus heterostichon, Polypheretima elongata./i. and bulk soil from a Bamboo plantation near Cali, Colombia. Pots of 1500 g of an adjacent tobacco field soil were brought to field capacity and two worms of each species placed in each pot. .i.Martiodrillus./i. gut contents and wormcasts from the soil surface and within tunnels were sampled after three months. The soil was maintained at field capacity for a further five months during which the worms continued to produce casts. Worm cast Amplified ribosomal DNA restriction analysis ARDRA (using Tag I) with actinomycete specific primers gave two strong bands at approximately 750 and 370 bp. ARDRA patterns from the three bamboo grove and four field soils had several additional bands notably at 140 and 126 bp.
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