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Characterization of fungal community in spruce (Picea abies) litter using cultivation and T-RFLP
Kolářová, Zuzana ; Koukol, Ondřej (advisor) ; Brabcová, Vendula (referee)
Fungi have a key role in the decomposition of coniferous litter and affect nutrient cycling in forest ecosystems. Therefore, great emphasis is placed on exploring the diversity of these organisms. The aim of this thesis was to describe fungal diversity in spruce litter and revealed temporal development of this community in a forest regenerating after bark beetle outbreak. Another objective was to compare sites with different length of bark beetle damage. The study area was located in the Bohemian Forest mountain range. Litter bags with spruce needles were placed on the forest soil and several samplings were performed in the course of three years. Diversity of fungi and changes in the fungal community were assessed by two methods: cultivation of needles on 2ř MEA and fingerprinting method T-RFLP. In total 71 fungal species were obtained from needles during a three-year succession by cultivation approach. Using T-RFLP 122 different fragments were generally recorded. The dominant species were Scleroconidioma sphagnicola, Thysanophora penicillioides, Hormonema dematiodes, Ceuthospora pinastri, species of genus Chalara, Trichoderma polysporum, Mycena galopus and unknown species Helotiales sp. 1. Primary saprotrophs occured in the community mainly in first 8 months and then were replaced by basidiomycetes....
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Určení pohlaví ptáků analýzou DNA
Rádsetoulalová, Iva
In my bachelor thesis -- Determining the sex of birds by DNA analysis, I dealt with covering information about ways of determining sex of birds. I focused on molecular methods, especially sexing birds by analyzing DNA by PCR method, which is based on amplification of CHD1 gen. CHD1 gen is located on both sex chromosomes of almost all bird's species. DNA is getting out of blood samples, feathers or intact membranes. I described avian reproductive system, sex chromosomes, the most common methods of determining sex, PCR method and sex determining genes and genes usable for determining sex of birds.
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Srovnání metod RFLP, RAPD, SSR a AFLP používaných při studiu genomu Prunus persica (L.)
Vlašínová, Martina
The Peachtree (Prunus persica (L.) is a considerable agricultural breed which is planted more than 4.000 years. It has nutritional and garnish utilization. The molecular methods spread out still very fast and they expand the findings about the research of the Prunus breed. These methods are the fundamental part of the molecular genetics, the breeding, the fingerprinting and the mapping of the plant genome. Within the framework of the breeding of the new kinds the molecular markers SSR, RFLP, AFLP and RAPD help to improve the Peachtree quality and to increase the resistance to diseases and pathogens. These molecular markers are used during the research of the genetic variability and of the structure of the Prunus breed.
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Microbial profile of model cheese
Lavičková, Ivana ; Veselá, Mária (referee) ; Vítová, Eva (advisor)
This Diploma thesis deals with the identification of microorganisms in samples of experimentally produced cheese. Model samples of cheese were produced in association with a private manufacturer. Raw organic milk was used to make the cheese. The theoretical part provides an overview of the issues such as the characteristics of the cheese, its properties and compounds. A special chapter is dedicated to molecular diagnostic methods, which serve for identification of microorganisms. In the experimental part of the thesis were identified some microorganisms in the samples. A polymerase chain reaction was used. The DNA was isolated from coarse lysates using phenol extraction; it was amplified using specific primers and demonstrated by gel electrophoresis. DNA of lactic acid bacteria (Lactobacillus spp., Lactococcus spp.) and yeast was ascertained in the cheese. The samples do not contain DNA of pathogenic genera Bacillus and Salmonella.
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Isolation, Identification and Characterisation of Microbial Communities of Wine and Selected Foods
Šuranská, Hana ; Španová, Alena (referee) ; Jarošová, Alžběta (referee) ; Omelková, Jiřina (advisor)
Proposed dissertation thesis deals with wine and artisanal cheeses microbiology. The first part is focused on identification of yeasts isolated from grapes and musts during production of white and red wines. The grape varieties were grown under the integrated and organic farming on Moravian vineyard. Yeasts were identified by ITS-PCR-RFLP method (amplifying internal transcribed spacer ITS: ITS1, ITS2 and 5.8S rDNA) and unknown species were subjected to partial sequencing of ITS rDNA region. In total, 524 isolates were divided into 14 different species belonging to six genus were identified from. The first stages of fermentation process were characterised by predominance of non-saccharomyces species especially H. uvarum. Due to increased ethanol concentration strains of S. cerevisiae prevailed in the later phases of the process. Further, partial aim of this study was to isolate and to apply selected autochthonous S. cerevisiae strains as starter culture during controlled industrial wine fermentation process. Genus Saccharomyces was distinguished from other non-saccharomyces species by ITS-PCR-RFLP. Further, in order to distinguish Saccharomyces genus at the species and the strain level, several molecular methods were applied including PCR-fingerprinting (rep- and RAPD-PCR), species-specific primers (multiplex and touchdown PCR), LSU-DGGE and interdelta PCR. Species-specific primers enabled us to distinguish some species of the Saccharomyces sensu stricto complex. Furthermore, interdelta PCR seems to be useful tool for S. cerevisiae strains identification. Among 120 isolated autochthonous strains belonging to Saccharomyces genus, 45 different strains were identified. Based on its sufficient technological properties (osmo- and ethanol tolerance, low H2S production etc.), S. cerevisiae 1-09 strain isolated from grape berries coming from moravian vineyard was chosen. Strain S. cerevisiae 1-09 was tested in small amount of must and after that also during industrial fermentation of red and white wine production. Based on the results of chemical and sensorial analysis, the strain seems to be suitable for application as the starter culture for winemaking process. The final part of this thesis is focused on quantification and identification of the yeasts isolated from artisanal cheeses and their by-products coming from Western Balkan Countries. Isolated species were identified by ITS-PCR-RFLP, partial sequencing and by physiological tests. Among the 20 yeast species found, D. hansenii, C. zeylanoides and Y. lipolytica were found to be predominant. Moreover, we developed culture-independent, semi-quantitative technique based on construction of ITS-clone library from metagenomic DNA to investigate complex fungal communities associated with artisanal cheeses and their by-products. Novel technique is based on direct extraction of total DNA from the sample. This was compared with culture-dependent ITS-PCR-RFLP and culture-independent LSU-DGGE methods. The results highlighted the discrepancies among these methods. Finally, the divergences among applied methods were confirmed by correlation analysis and by indices of general biodiversity and dominance of species. ITS-clone library approach combines the advantages of cultivation-based analysis and LSU-DGGE with semi-quantification of fungal species without the requirement of their cultivation. This study might open new perspectives in direct and complex analysis of yeasts and moulds in food matrices.
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Blastocystis subtypes in pigs
LAKATOSOVÁ, Lucie
Blastocystis is an anaerobic single-celled protozoan that commonly occurs in the intestinal tract of animals and humans. It is a genetically very variable organism. Blastocystis can be found both in healthy specimens and in patients with gastrointestinal disorders. In 2011/2012, I examinated already isolated DNA samples from domestic and wild pigs by molecular methods in the laboratory of the ASCR Parasitology Institute. In total, 110 samples were examined. The domestic pigs were positive in 91 % (51/56) and wild pigs in 67 % (36/54). The overall prevalence of domestic pigs was higher by 24 %. Several sequences of SSU rRNA gene obtained during the work suggest that pigs may serve as host for atypical Blastocystis subtypes.
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Population biology of \kur{Rhytidoponera} ants (Hymenoptera:Formicidae)
BIZOS, Jiří
Ants from the genus Rhytidoponera are known for specific type of social organization in which some workers can reproduce. Species of this ants are commonly occuring in lowland rainforests of New Guinea but their biology is unknown to great extent. This thesis reviews available molecular methods suitable for study of population structure and phylogeography of Rhytidpoponera ants in rainforest environment.
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Review of molecular phylogeography of \kur{Crematogaster polita} (Hymenoptera: Formicidae)
DRAGOVÁ, Klára
Crematogaster polita group Smith F. 1865 is tropical species, where it forms dominant elements of the arboreal fauna. This species has polydomous large colonies characterized by territorial behaviour. Its morphology is very variable and therefore identification of this species is very difficult. This thesis review molecular methods that are used for study of population and can be alternatively used for identification of cryptic species.
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Construction of expression vector for cyanobacterium \kur{Synechocystis} sp. PCC 6803
RICHTOVÁ, Jitka
The cyanobacterium Synechocystis sp. PCC 6803 is an important model organism used in the study of photosynthesis and related processes. The only disadvantage of Synechocystis for molecular biology studies is that it lacks a cloning/expression system. The aim of this work was to evaluate whether it is possible to use a mutated high copy version of the native Synechocystis plasmid for expression of a chosen gene in this organism.
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