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Comparative Analysis of Silk Proteins and Discovery of Novel Sericin Gene in Lepidopteran Moths
WU, Bulah Chia-Hsiang
This thesis focuses on the silk components of the Mediterranean moth, Ephestia kuehniella, and the discovery of a novel silk gene, P150/ser6, in the silkworm, Bombyx mori. We analyzed and described the cocoon silk components in both species. In the first publication, we combined transcriptomic, genomic, and proteomic approaches to identify silk proteins in E. kuehniella. In the second publication, we described the discovery of gene P150/sericin6 in B. mori based on microsynteny analysis.
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Cytogenetická analýza vybraných mutantních linií bource morušového
CRHONKOVÁ, Adriana
Bombyx mori is the main model organism in order Lepidoptera, the biggest group with female heterogamety (ZZ/ZW). In past 5000 years of its breeding arose many mutant strains, kept in stocks in Japan, China or other countries. This study is focused on sex-limited pB (black larval marking) Bombyx mori (TWPB) strain. The aim is to confirm the presence of neo-W chromosome.
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Threads and Nets Made By Invertebrates
Šuláková, Eva ; Říhová, Dagmar (advisor) ; Hanel, Lubomír (referee)
This diploma thesis is focused on the silk production of invertebrates. The aim of the work is to systematically introduce important species and describe the purpose of their silk production. The ability to produce silk is widespread in many insect families. The production of proteinaceous fibers also occurs in arachnids and (in the form of byssus threads) also in mollusks. The Domestic Silk Moth (Bombyx mori) is an economically important insect, being a primary producer of silk. Hence, an integral part of the thesis focuses on Domestic Silk Moth. In this thesis all developmental stages of Silk Moth (from an egg to an adult) are described. In the past, efforts to breed silkworms also appeared in the Czech lands. Therefore, a short chapter of the thesis is devoted to the history of sericulture. Furthermore, this thesis describes methods of rearing silkworm larvae. For students of Natural Sciences and Biology, it is crucial to be in touch with live objects. Because the silkworm rearing is quite simple, it is possible to incorporate this topic to the school projects about holometabolous insects. The practical part of this thesis contains schedule of practical exercise about silk moths. And in final section of practical part, worksheets for lower-secondary students are presented. Keywords: silkworm...
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Nosema bombycis as a model organism for polar tube study.
Satrapová, Alexandra ; Petrů, Markéta (advisor) ; Trdá, Lucie (referee)
Microsporidia is a group of obligatory intracellular parasites with a unique invasion mechanism. One of the highly specialized invasion structures is the polar tube which is an example of a biological nanotube the study of which could bring usable knowledge in biotechnology. The thesis is a literary overeview which deals with the structure of microsporidian spores and presents N. bombycis as a suitable representative for the study of the infectious apparatus. In more detail, it summarizes the background to the structure of the polar tube and wall of N. bombycis spore.
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Gene manupulations in invertebrates
Čermáková, Eliška ; Schierová, Michaela (advisor) ; Konopásek, Ivo (referee)
Gene manipulations in invertebrates are based on the same approches used in vertebrates. The are applied for the development of new genotypes in model species, convenient as model systems of human hereditary diseases etc. Gene manipulations are important as well for practical purposes, which is shown by the example of trangenic mosquitoes. Recently, it has been proved that programmable nucleases can be successfully used in invertebrates. Key words: Gene manipulations, invertebrates, methods, applications, Drosophila melanogaster, Anopheles, Bombyx mori, malaria
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Embryonic development of the unfertilized silkworm eggs
VRCHOTOVÁ, Markéta
Transgenesis of silkworms has great potential for the development of silk with new properties as well as for the preparation of recombinant proteins for the use in biomedicine. Zabelina et al. (2015a) showed that transgenesis of parthenogenetic silkworms facilitates the selection and maintenance of transgenic homozygotes with stabile transgene insertions. However, the efficiency of transgenesis was less than 2 % compared to 60 % in the standard, non-parthenogenetic silkworms. The purpose of the present research was to explore the cause of this difference. Since transgenesis is normally performed at 25 °C but in the parthenogenetic silkworms at 15 °C (3 days incubation at this temperature is part of the protocol for the induction of parthenogenetic development), we assumed that the eggs incubated at 15 °C might have been injected with the DNA construct at unsuitable time. The work was therefore focused on the rate of embryogenesis at 15 °C in the eggs treated in different ways. Intensive cleavage of the control eggs (strain K23) was observed at 12 h after oviposition at 25 °C and between 24 and 36 h at 15 °C. The transgenesis of parthenogenetic silkworms is also complicated by the embryonic diapause. In the current work, diapause was suppressed by implanting PK1 ovaries into the non-parthenogenetic male hosts K23. Parthenogenetic development was activated by the heat shock in the chorionated eggs dissected from the implants. No cleavage was detected at 12 h after the acti-vation and nearly complete blastoderm was found at 48 h. In respect to the course of cleavage at 15 °C, transgene injection 24 h after the activating heat shock can be recommended. The eggs from endogenous ovaries of the K23 females, which also contained the implants of the PK1 ovaries, acquired partial capacity of parthenogenesis. Low rate of embryogenesis was also found in the transgenic clone VTG1. Current results suggest that more research is needed to understand and possibly explore differences in the rate of silkworms exposed to different treatments.
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Studies on the sericin 3 of \kur{Bombyx mori} and cloned sericin into \kur{Escherichia coli}
KRŮČEK, Tomáš
The spun-out silk fiber consists of two fibroin filaments that are cemented together by sericin coating. The serine-rich sericins, which make 20-30% of the cocoon silk proteins in Bombyx mori, are dissolved in hot water during silk fiber reeling from the cocoon. The sericin extract is usually discarded. Only small amounts are currently used in cosmetics and lately also as replacement of bovine serum products in the cell culture media. The use in culture media is hindered by poor standardization of the extracts. To overcome this problem, we attempted preparation of several recombinant proteins based on partial sequences of one out of the three sericin genes of Bombyx mori, sericin 3. Translation product of the Ser3 gene is extremely serine-rich with a relatively high representation of the aspartate, asparagin, glutamate, and glutamine. Using specific primers we have prepared a cDNAs of 267, 279, 525, 672, and 528 bp, respectively, derived from the Ser3 gene. The cDNAs were cloned and expressed as fusion proteins with hexahistidine in Escherichia coli. The proteins were purified by affinity chromatography and analyzed by acrylamide electrophoresis. Ser3 gene contain repetitive motifs rich in serine and including some charged amino acids.
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