Using silkworms as the potential host to spin spider silk-like fibers is an area of intense research world-wide. The conventional methods used to create transgenic silkworms hosting spider silk-like gene limits the incorporation of spider silk-like protein and do not improve the mechanical performance of the composite silkworm/spider silk fibers. In this dissertation, synthetic spider ampullate genes were incorporated into the precise site of the fibroin heavy chain or light chain using the latest genome editing technology CRISPR/cas9 guided non-homologous end joining as opposed to conventional random integration using transposon-based piggyBac system. These protocols, with extensive applicability to other silkworm researches, improved the content of spider silk-like protein in the transgenic silkworm/spider silk fibers, increases genetic stability in offspring, and improves the mechanical performance of the transgenic fibers compared to traditional methods. In addition, an enhanced green fluorescence protein (eGFP) was successfully incorporated into the fibroin light chain of silkworms using CRISPR/C as 9 initiated homologous recombination. The transgenic silkworm/spider fibers emitted strong green fluorescence under excitation. These results demonstrate that the we successfully developed a protocol to make silkworm as a host to spin spider silk-like fibers.

Spider silks are incredible natural materials that have a wide variety of properties that can rival or outperform even common synthetic materials like Nylon and Kevlar. As nature’s architects, orb-weaving spiders spin seven different silks that are used for very specific roles throughout the spider’s lifecycle. These silks are comprised of proteins called spidroins. Each of these spidroins has evolved to have properties such as strength and/or stretch that make these silks successful and highly adapted in their designated roles in web construction, prey capture and reproduction. This study involves the production of minor ampullate silk by genetically modifying the bacteria Escherichia coli. Minor ampullate is a lesser studied silk that is used for the first spiral of the orb web. This spiral is a template that the spider uses to finish the web and provides stability during the web construction. Minor ampullate silk is strong, however it does not stretch so it may be well-suited for certain applications such as ballistic materials. By producing and purifying different arrangements of minor ampullate silk protein, it is possible to learn how this protein can be expressed without using the spider itself. This investigation sheds light on how deviations in the protein sequence and motif arrangement can produce different properties, which can potentially be used to make new materials.

Due to its strength, flexibility, and biocompatibility, spider silk is a highly appealing material for applications in the medical field. Unfortunately, natural spider silk is difficult to obtain in large quantities because spiders are territorial and cannibalistic, making them impractical to farm. Synthetic spider silk proteins produced by transgenic hosts such as bacteria and goats have made it possible to obtain the quantities of spider silk needed to study it more fully and to investigate its potential uses. The spider silk proteins produced in our laboratory do not have an optimal purification method to remove all of the non-biocompatible contaminants and have not previously been tested for their biocompatibility. The first focus of this dissertation was to create goat cells that can be used to create new goats. These new goats will produce proteins that can be purified more efficiently and more completely. The second focus of this dissertation was to perform biocompatibility tests on goat-derived spider silk proteins. Prior to performing any biocompatibility tests, a method was established for removing endotoxins – an impurity that causes an immune response in the body – from the proteins. This work has shed light on areas for improvement in the silk protein purification process and laid groundwork for the production of new goat-derived proteins. These steps will help make it possible for synthetic spider silk to progress further toward becoming a viable biomaterial.

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Orientador: João Sinézio de Carvalho Campos / Dissertação (mestrado) - Universidade Estadual de Campinas, Faculdade de Engenharia Química / Made available in DSpace on 2018-08-26T11:45:54Z (GMT). No. of bitstreams: 1 Bergamasco_DaivesArakem_M.pdf: 2473917 bytes, checksum: f246b9e27a6e46a969245f8ff3666b15 (MD5) Previous issue date: 2015 / Resumo: A indústria têxtil vem constantemente investindo tanto no processamento quanto na melhoria de seus produtos. Dentre a gama de materiais se destaca a seda, porém há a necessidade de adequá-la (beneficiamento) no sentido de se obter seu máximo rendimento. O presente trabalho visa apresentar resultados dos processos de degomagem e tratamento corona sobre tecidos de seda por analisar destacadas propriedades físicas tais como hidrofilidade, resistência à tração e perda de massa. Submeteram-se amostras de tecido de seda ao tradicional processo de degomagem com sabão de Marsellha e também pelo processo de tratamento corona. Posteriormente realizaram-se ensaios de hidrofilidade (medidas de ângulo de contato), resistência à tração e perda de massa. Os resultados mostram que: (i) tanto a degomagem quanto o tratamento corona promovem excelente hidrofilidade; no entanto o tempo para completa hidrofilidade é de 45 s quando tratado por corona e de 60 minutos quando por degomagem; (ii) a resistência a tração é maior com o processo corona e (iii) a perda de massa com a degomagem é em média 26% e com o processo corona é de 0,3%. Cabe ressaltar que com o processo corona praticamente não há geração de efluente em comparação com o de degomagem, o que significa outra grande vantagem para a indústria. De acordo com os resultados observam-se grandes vantagens na utilização do processo corona no que tange ao beneficiamento em tecido de seda / Abstract: The textile industry has been constantly investing in the processing and in the improvement of its products. Among the range of materials stands out silk, but there is a need to adjust it (improvement) in order to obtain their maximum efficiency. This paper presents results of degumming processes and corona treatment on silk fabrics by analyzing important physical properties such as hydrophility, tensile strength and weight loss. Underwent silk tissue samples to traditional degumming process Marsellha soap and also for the corona treating process. Subsequently took place hydrophility tests (contact angle measurements), tensile strength and weight loss. The results show that: (i) degumming and corona treatment promote excellent hydrophility; however, the time to complete hydrophility is 45 s when treated by corona and 60 minutes when a degumming; (ii) the tensile strength is higher with the corona process, and (iii) the mass loss is degumming with an average of 26% and the corona process is 0.3%. Note that with the corona process there is no generation of effluent compared to degumming, which means another advantage to the industry. According to the results, there are large advantages in using the corona process with regard to the processing of silk fabric / Mestrado / Ciencia e Tecnologia de Materiais / Mestra em Engenharia Química

The diploma thesis focuses on create a marketing mix in the company New Brand, which deals with the adjustment of silk accessories. Specifies the various components of the marketing mix and business environment analysis. Proposes to extend the product range and focus on individual customers.

Spiders are capable of producing a variety of silk types, each with their own unique protein composition and function. Dragline silk in particular, has been of great interest due to its high tensile strength and extensibility. In the past, synthetically produced dragline fibers have not been able to match the superior properties of natural silk. A recent discovery in the western black widow spider, Latrodectus hesperus, might be the missing link between the current state of synthetic silks and naturally produced fibers. Our research is centered around the discovery a new family of five low-molecular-weight cysteine-rich proteins (CRPs) and their potential function within dragline silk. This study focuses on the characterization of recombinantly expressed CRP1, CRP2, and CRP4. Through structural analysis using circular dichroism, it has been determined that the CRP family members have mostly alpha-helical secondary structure and exhibit small differences in their ability to maintain their structure in the presence of changing environmental conditions. The study also covers the effects of temperature and pH on the folding and unfolding of the CRPs. It appears that pH is the dominant influence on protein unfolding within the major ampullate gland.

The Tomioka Silk Mill, an extraordinary industrial heritage in the Prefecture of Gunma on the Honshu island, is waiting to be included to the List of Unesco (nowadays is only on the List Tentative). Former factory, documenting the integration of French technology and Japonese architecture, was realised on the principle of classical factory zoning. The conversion of the mill, aiming at multiple-municipal functions, is reflected in a new urbanistic conception of the area in the municipal context. The project proposes and checks functional zones for users and sightseeing routes for turists. The part of that is evaluating the potential of the mill objects, of the landscape of the mill and of the functional municipal relations.

Both existing buildings of designed Training Center are situated on the southern edge of the Tomioka Silk Mill above the steep river side. It is former dormitory for female workers and a house of the first manager of the factory Paul Brunat. Preparation of registration of the factory as UNESCO World Heritage Site and its subsequent functional transformation raises the need for trained professionals. Lot of organizations from surrounding area are now studying topic of the Tomioka Silk Mill. Thematic lectures about silk industry, history of the Tomioka Silk Mill and related sites, seminars, workshops and training of guides and other voluntaries, preparation and discussion of propagation of the mill, international conferences of experts of industrial heritage and silk production from all around the world and experts from Gunma prefecture, exhibitions and other events could take place in the new Training Center. Dormitory Rooms for seminars, lectures and offices for temporary teachers are situated in the southern block of former dormitory. In the other block, rooms for short-term accommodation stay in original condition of traditional Japanese style. Connection of waterfront and training center has been achieved through increase of entrances into the training rooms and restroom from the south of the building. This narrow strip of ground is settled by elevated terraces and green areas planted with bamboo. Building of the training center expanded to a new hygienic block on the west side. Part of it slightly exceeds above the river and creates observation terrace and it also makes the walking passage wider. Green atrium is situated between two blocks of dormitory. Brunat House Brunat house is designed for conferences and administration. The first manager has used this object as residence only a few years and then its use was changed to a school for workers of the mill. In addition to the main hall there is entrance foyer, dressing room, sanitary facilities and cafe bar. Administrative body of the Tomioka Silk Mill – the Tomioka Silk Mill Division is situated in the western block of Brunat house. Other activities of this office are: area and building maintenance, care of visitors, guides training, research and works related to world heritage registration. They also collaborate with Gunma government office World Herritage Registration Promotion Division and other experts. Waterfront Waterfront with southern sunlight is ideal for creating place for relax connected to surrounding functions of museum and training centre. Relax waterfront should offer calm corners, opportunity for sitting or lying on the southern sunlight and also nice viewpoint on the mountains surrounding the Tomioka city. It should connect Japanese nature with industrial environment and western style, as it was in the time of establishment of the factory. Training Centre should meet the needs of local organization and also experts from all around the world. Therefore it should be mix of Japanese and western culture.