A reprogramação nuclear de uma célula somática a um estado embrionário tem diversas aplicações, como pesquisas básicas na biologia do desenvolvimento, terapia celular, melhoramento genético em animais de produção e conservação de espécies. As principais técnicas utilizadas para a reprogramação nuclear são a transferência nuclear de células somáticas (TNCS) e a geração de células tronco pluripotente induzidas (iPS). Muitos trabalhos têm mostrado uma baixa eficiência no processo de reprogramação nuclear nas duas técnicas, além disso, modificações epigenéticas tem sido apontada como a principal barreira para uma reprogramação nuclear eficiente. Por esse motivo, medidas como a utilização de células menos diferenciadas e/ou alteração do perfil epigenético das células somáticas podem aumentar a eficiência destas técnicas. Por isso, o objetivo deste trabalho foi investigar a influência de marcas epigenéticas em células bovinas utilizadas na reprogramação nuclear mediada por TNCS ou superexpressão de genes relacionados a pluripotêcia (iPS). Para isso, utilizamos 3 abordagens. Primeiro, analisamos marcações epigenéticas relacionadas ao desenvolvimento embrionário e pluripotência (H3K9me2, H3K9me3, H3K9ac, 5mC e 5hmC) em diferentes tipos celulares, analisamos a expressão gênica de genes responsáveis por essas marcações em células de diferentes tecidos (ex. células tronco mesenquimais (MSC) e fibroblastos) e as utilizamos como doadoras de núcleo na TNCS. Na segunda e a terceira abordagem, utilizamos células com menores níveis de H3K9me2 para a geração de iPS e na TNCS, respectivamente. Além disso, por se mostrar eficiente na TNCS, analisamos o efeito da sincronização do ciclo celular por privação de soro fetal bovino (SFB) na geração de células iPS. Com o intuito de diminuir os níveis de H3K9me2, as células foram tratadas com UNC0638, um inibidor especifico das metiltransferases de histona G9a/GLP. Nossos resultados do primeiro experimento mostraram que as MSC podem ser utilizadas como doadoras de núcleo na TNCS, no entanto, mesmo com algumas diferenças na expressão gênica em relação aos fibroblastos, a produção de blastocistos não foi diferente entre as duas células. No segundo experimento, as células privadas de SFB geraram mais colônias que as células controle, enquanto que as células tratadas não apresentaram diferença. Por último, as células tratadas com o UNC0638 apresentaram um menor nível de metilação no DNA em zigotos em relação às células controle. Os resultados encontrados neste trabalho podem contribuir para o melhor entendimento dos mecanismos epigenéticos envolvidos na reprogramação nuclear de bovinos / Nuclear reprogramming of somatic cells to embryonic state has several aplications, such as basic research on developmental biology, cell therapy, genetic improvement in livestock animals and preservation of endangered species. The principal techniques utilized to achieve nuclear reprogramming are Somatic Cell Nuclear Transfer (SCNT) and induced pluripotency. Several works has reported low efficiency rates of nuclear reprogramming when these techniques are used to reprogram somatic cells. Moreover, epigenetic modifications acquired during development act as epigenetic barrier to the complete reprogramming process. For this reason, strategies such as use of less differentiated cells and/or modification of epigenetic profile of somatic cells might increase the efficiency these techniques. The objective of this work was investigate the influence of epigenetic marks in bovine cells utilized on nuclear reprogramming experiments mediated by SCNT or induced pluripotency. To investigate it, we used three approaches. First, we analyzed the epigenetic marks related to the embryonic development and pluripotency (e.g H3K9me2, H3K9me3, H3K9ac, 5mC and 5hmC), gene expression of genes involved in these epigenetic marks in different tissues (i.e. mesenchymal stem cells (MSC) and fibroblasts) and their use as nuclear donor cells on SCNT procedure. Regarding the second and the third approach, we utilized cells with reduced levels of H3K9me2 to generate iPS cells and cloned embryos, respectively. Furthermore, since serum starvation has been demonstrated increase SCNT developmental rates, we assessed the effect of cell cycle synchronization mediated by serum starvation on nuclear reprogramming using iPS cells. Aiming decrease the levels of H3K9me2, cells were treated with UNC0638, a chemical probe that works as a specific inhibitor of the histone methyltransferases G9a and its counterpartner GLP. Our results showed that MSC are suitable to be used as nuclear donors on SCNT procedures, however, in spite of differences on gene expression comparing with fibroblasts, the embryonic developmental rates were not improved. On the second experiment, cells privated of fetal calf serum produced more iPS cells colonies than control cells, whereas cells treated with UNC did not show differences when compared with untreated cells. Lastly, UNC treated donor cells treated produced cloned zygotes with lower levels of DNA methylation compared to zygotes derivated from untreated cells. The results presented here will contribute to the better understanding of the epigenetic mechanisms involved on bovine nuclear reprogramming

Transportation-network companies such as Uber and Lyft have permeated cities around the world. While they have been lauded for introducing a new mobility option, questions of equity have been raised, including in relation to people with disabilities. Numerous lawsuits have been filed against Uber and Lyft for lack of disabled accessibility of transportation service they facilitate, with some of the lawsuit focusing on wheelchair accessibility. These have been reported in the media while work on the topic is lacking in the academic literature. This doctoral research explores wheelchair accessibility in transportation hailed through Uber and Lyft in the U.S., with a partial focus on the phenomenon in Washington, DC. The study sought to gain insight into wheelchair-accessibility matters with respect to four main stakeholders - riders, companies, drivers, and regulators and other government agencies. The purpose was a) to deepen understanding on the topic based on a holistic approach to core issues and produce evidence-based knowledge and b) to provide policy recommendations where needed. A mixed-methods approach was used to investigate the research problem. The quantitative strand surveyed 341 wheelchair users in the U.S., while the qualitative strand consisted of three sets of 16 qualitative interviews with drivers on the Uber and Lyft apps, Uber and Lyft company representatives, and representatives of relevant government agencies in Washington, DC. The study finds the following: 1) The service might be filling a transportation gap for some wheelchair users even though it is a two-tiered system that disadvantages motorized-wheelchair users. 2) The challenges surrounding wheelchair accessibility are perceived differently by the stakeholders and as a result there are differing views of how to solve them. 3) Overcoming inaccessibility challenge needs to involve changes on both Uber and Lyft and government agencies. Uber and Lyft and government agencies will find this study of interest to help them appreciate other stakeholders' perspective and use the findings to support policy decisions. / Doctor of Philosophy / Transportation-network companies connect drivers who are interested in providing a ride to riders who are looking for transportation through smartphone apps. Uber and Lyft are the most common of these companies. Both companies operate in many cities around the world. They are praised for creating a transportation alternative, but there is a concern that they may not serve everyone equally. For example, people with disabilities, especially wheelchair users have complained that they are not receiving the same service as those who do not use wheelchair. In the US, there are numerous lawsuits have been filed against Uber and Lyft for lack of disabled accessibility. Some of the lawsuit focus on wheelchair accessibility. These issues are reported in the media mostly. This doctoral research looks at wheelchair accessibility in transportation hailed through Uber and Lyft in the U.S., with a partial focus on the phenomenon in Washington, DC. The study sought to gain insight into wheelchair-accessibility matters with respect to four main stakeholders - riders, companies, drivers, and regulators and other government agencies. The purpose was a) to deepen understanding on the topic based on a holistic approach to core issues and produce evidence-based knowledge and b) to provide policy recommendations where needed. A survey and a set of interviews were used to investigate the research problem. The study finds the following: 1) The service might be filling a transportation gap for some wheelchair users even though it is a two-tiered system that disadvantages motorized-wheelchair users. 2) The challenges surrounding wheelchair accessibility are perceived differently by the stakeholders and as a result there are differing views of how to solve them. 3) Overcoming inaccessibility challenge needs to involve changes on both Uber and Lyft and government agencies. Uber and Lyft and government agencies will find this study of interest to help them appreciate other stakeholders' perspective and use the findings to support policy decisions.

A tranferência nuclear de células somáticas (TNCS) está sendo utilizada para produzir cavalos de elite. No entanto, durante este procedimento pode ocorrer a perfuração da zona pelúcida, levando, ocasionalmente, à secção da massa celular interna, e conseqüente derivação de gêmeos monozigóticos. Além de serem relatadas alterações no processo de imprinting genômico, que conduzem ao desenvolvimento de doenças. Com a descoberta da possibilidade de reprogramar as células somáticas a um estado de pluripotência (iPSCs), estas células passaram a ser muito utilizadas em pesquisas de neurociência. Contudo, também ocorrem modificações epigenéticas durante esta reprogramação celular. Portanto, nossas hipóteses são que os gêmeos eqüinos gerados pela TNCS podem levar às irregularidades no desenvolvimento do sistema nervoso. O padrão de metilação do SNRPN nas estruturas dos fetos muares clonados, e as células iPSCs são diferentes dos padrões encontrados nos muares analisados. A expressão dos genes SNRPN, Necdin e UBE3A são maiores no cérebro, enquanto a expressão do H19 é maior nas membranas extra-embrionárias. Em nosso estudo, obtivemos duas gestações gemelares equinas derivadas da TNCS, que foram interrompidas com 40 e 60 dias de gestação, e comparados com gestações eqüinas únicas de idade similar. Diferenças no comprimento entre os embriões gêmeos foram observadas aos 40 (2.0 e 2.2 cm 10%) e aos 60 (6,5 e 8,5 cm 24%) dias de gestação. Somente o plexo coróide do quarto ventrículo apresentou-se mais desenvolvido nos fetos com maior comprimento. Ao analisarmos fetos muares clonados em diferentes idades gestacionais e compará-los com muares, nos períodos embrionário, fetal e adulto, não foi observada diferença no padrão de metilação do gene SNRPN. No entanto, na décima passagem das células iPSC o padrão de metilação alterou, em relação aos muares estudados e ao padrão observado nos fibroblastos. Ao analisarmos os fetos clonados nas diferentes idades gestacionais observou-se no cérebro menor expressão dos gene H19 e UBE3A, e maior expressão do gene SNRPN. Contudo, a expressão do gene Necdin variou entre as estruturas estudadas. Em conclusão, apesar dos gêmeos eqüinos provenientes de TNCS diferirem quanto ao tamanho, morfologicamente são iguais. Dentre as estruturas cerebrais o plexo coróide se apresentou mais desenvolvido nos fetos de maior comprimento. Os fetos muares clonados não apresentaram diferença no padrão de metilação do gene SNRPN. No entanto, as iPSCs apresentaram alteração no padrão de metilação deste gene na décima passagem. Embora os genes SNRPN, Necdin e UBE3A sejam expressos no cérebro, o SNRPN apresentou-se prevalente nessa estrutura / The nuclear transfer of somatic cells (SCNT) is being used to produce elite horses. However, during this procedure can occur drilling of the zona pellucida, leading occasionally to the section of the inner cell mass, and subsequent derivation of monozygotic twins. Besides being related changes in genomic imprinting process, leading to the development of diseases. With the discovery of the possibility to reprogram somatic cells to a pluripotent state (iPSCs), these cells have become widely used in neuroscience research. However, also occur epigenetic changes during this cellular reprogramming. Therefore, our hypothesis is that equine twins caused by equine ART could lead to developmental irregularities of the nervous system. The patterns of SNRPN methylation in the structures of cloned mule fetuses and in iPSCs are different from the patterns found in the analyzed mules. And the expression of SNRPN, Necdin and UBE3A genes are higher in the brain, while the higher expression of H19 gene occurs in the extraembryonic membranes. In our study we derived two equine twin SCNT pregnancies that were interrupted at 40 and 60 days of gestation and compared to singleton fetuses of similar age. Differences in lengths between twin embryos were observed at both 40 (2.0 and 2.2 cm 10%) and 60 (6.5 and 8.5 cm 24%) days of gestation. Only the choroid plexus in the fourth ventricle more developed in the twins with the greatest length. Analyzing mules cloned fetuses at different gestational ages, and compare them with mules at embryonic, fetal and adult period; there was no difference in the pattern of methylation in SNRPN gene. However, in the tenth passage of the iPSCs the methylation pattern was altered in relation to the studied mules and the pattern observed in fibroblasts. When the cloned fetuses at different gestational ages were analyzed, the brain presented lower expression of H19 and UBE3A genes, and higher expression of SNRPN gene. However, the expression of Necdin gene varied among the structures studied. In conclusion, despite the twin horses from SCNT differ in size, they are morphologically identical. Among the brain structures the choroid plexus performed more developed in the fetuses of greater length. Cloned mules fetuses showed no difference in the pattern of methylation SNRPN gene. However, iPSCs have changes in the pattern of methylation of this gene in the tenth passage. Although SNRPN, Necdin and Ube3A genes are expressed in the brain, SNRPN is prevalent in this structure

Small and Medium Enterprises (SMEs) are one of the principal driving forces in the development of an economy because of its significant contribution in terms of number of enterprises, employment, output and exports in most developing as well as developed countries. But SMEs, particularly in developing countries like India, face constraints in the functional areas of technology, finance, marketing and human resources. Moreover these SMEs have been exposed to intense international competition since early 1990s because of globalisation. However, globalisation, the process of continuing integration of the countries in the world has opened up new opportunities for SMEs of developing countries to cater to wider international markets which bring out the need for these SMEs to develop competitiveness for their survival as well as growth. Subcontracting relationship of SMEs with Large Enterprises (LEs) is an important source of access to technology and other infrastructural resources for SMEs of developing countries enabling them to develop their capabilities and become competitive. In the era of globalisation, Trans National Corporations (TNCs) are expanding their production facilities to developing countries for availing the advantages of productivity and distribution more than ever and India is no exception. These TNCs which concentrate on core operations and outsource non core activities offer better scope for local SMEs to have subcontracting relationship with them. Superior work culture of these TNCs would be reflected in their subcontracting relationships with local SME suppliers since TNCs place much emphasis on their value chain to be organised according to international standards. The key issue is whether Indian SMEs have entered into subcontracting relationship with TNCs and if so what is the nature of these subcontracting relationships? Does this relationship with the TNCs offer more scope for receiving assistance of various kinds for subcontracting SMEs? What is the extent and diversity of this TNC assistance? Given the quantum of assistance, does it facilitate the innovations and economic performance of SMEs? These questions have been addressed in the study with reference to subcontracting SMEs of three major TNCs in the Indian automobile industrya Japanese TNC, a German TNC and a Swedish TNCall the three being located in Bangalore, India. A theoretical framework for subcontracting relationship between TNCs and SMEs is proposed focusing on TNC assistance, SME innovations and economic performance, based on literature review. The study is based on an Ex Post Facto Exploratory research using primary data collected from a sample of 81 SME subcontractors selected from the list of first level suppliers provided by the three TNCs. The relevant data were collected using a structured questionnaire by visiting the firms and having personal interviews with entrepreneurs/senior managers of the firms. An assessment of nature of the SME subcontractors and their subcontracting relationships with TNCs revealed that these SME subcontractors of the three TNCs are in different stages of TNCSME subcontracting relationship. The Swedish TNCSME subcontracting relationships are in the initial stage, the Japanese TNCSME subcontracting relationships are in the growing stage and German TNCSME subcontracting relationships are in an enduring stage of the relationship. Therefore they are alternatively referred to as initial stage SME subcontractors, growing stage SME subcontractors and enduring stage SME subcontractors. The assistance a subcontracting SME received from its TNC customer was measured using 20 item variables under 7 dimensions of assistance related to product, production process, managerial know-how, marketing, human resource, financial and purchase process. A model to measure the degree of assistance was formulated with these 20 item variables for measuring the seven dimensions of assistance. Confirmatory factor analysis was used to establish the validity of the model. The degree of assistance was calculated as a single summated score for each case by summing up the composite scores of the seven dimensions of assistance calculated using weighted average method. The assistance provided to SME subcontractors was more in the case of German TNC than those of Japanese TNC and Swedish TNC. Even though there was evidence of assistance from TNCs to SME subcontractors, these SMEs received more of product related and purchase process assistance whereas assistance for their production process, managerial know-how, marketing, human resource, financial requirements was not very high. This implies that subcontracting relationship of Indian SMEs with TNCs is confined more to a mere purchase supply relationship where both the parties are concerned about the basic requirements of purchase supply relationship like detailed specifications, proper feedback on product performance, advance information about future orders, preferential pricing and proper payment. Innovation of the subcontracting SMEs was measured based on six dimensions of innovation, namely, new product developments, product modifications, process improvements, informal R&D/NPD expenditure, informal R&D/NPD employees and raw material selection. Firms were classified into two groups as high innovators and low innovators based on the innovation score calculated using the six dimensions. To probe the role of TNC assistance in the innovations of subcontracting SMEs, a logistic regression analysis was performed using the equation with degree of assistance, capital and labour as explanatory variables and innovation level as criterion variable. Our analysis revealed the positive role of assistance in the innovations of SMEs. However, the positive role of this assistance was found to be decreasing as focus of our analysis shifted from the initial stage SME subcontractors to the growing stage SME subcontractors and then to the enduring stage SME subcontractors even when the assistance they received increased from one stage to the other. SME subcontractors operating in the initial stage of the relationships, who in general lack technological competence, rely more and make more use of the assistance, even when the assistance itself is low, for their innovative performance. As the relationship gets older, the SMEs may be able to enhance their inhouse competence using the assistance from the TNC and may make more use of their own resources for innovations along with the assistance from TNCs. Therefore, as SME firms begin to use more of their inhouse resources for their innovations, it is likely that the role of TNC assistance for SME innovations would decrease. Increasing positive role of labour for SME innovations as the relationship shifted from initial to enduring stages substantiate the argument of increasing utilisation of inhouse resources for innovations as relationship prolongs over a period of time. A comparative analysis of the economic performance of SME subcontractors revealed that the German subcontractors were having more labour productivity and value added to value of output and low capital productivity whereas Swedish subcontractors were having high capital productivity and low labour productivity and value of added to value of output compared to other two. In order to probe the role of degree of assistance on the economic performance of SMEs, a regression analysis was performed using the equation with degree of assistance, capital and labour as explanatory variables and value addition as criterion variable. The results revealed the contributory role of TNC assistance in the economic performance of subcontracting SMEs but this contributory role of assistance was found to be decreasing even when the assistance itself is increasing whereas the contribution of labour increased, as our analysis shifted from the initial stage to the growing and then to the enduring stage of relationships. The extent of assistance might be low at the initial stage of the relationship but the small and young SME subcontractors who in general, lack technological competence might be able to make more use of this assistance since the assistance from TNCs would be one of their major sources for technical upgradation and growth. As the relationship prolongs both contracting and subcontracting firms may rely on trust and build up close relationship which would enable the SMEs to enhance their sales to the TNC and receive more assistance. At the same time, some of the SMEs might even strengthen their inhouse resources due to TNC assistance which in turn could be further used for their economic performance. This could be the reason for the decreasing contributory role of assistance and increasing contributory role of labour for the economic performance of subcontracting SMEs as our analysis moved to more enduring relationships. Further analysis using other regression models revealed that the firms, which received more assistance from the TNC customers utilised labour more efficiently than firms which received lesser assistance. TNC assistance was found to be an important factor for the enhancement of labour productivity of subcontracting SMEs, especially for young and small firms operating in the initial stage of the relationship. The subcontracting SMEs were able to make use of the innovations which they could carry out with TNC assistance for their economic performance. Quantile regression analysis, performed to have a more comprehensive picture of the effect of degree of assistance on economic performance, revealed that that for SMEs having average factor productivities, the assistance from TNCs contributed less to their economic performance relative to those SMEs which had low and high factor productivities, respectively. Moreover, SMEs which had high factor productivities were able to make more use of their innovations for their economic performance. These findings clearly show that the TNC assistance enables subcontracting SMEs to enhance their innovations and economic performance. Initially, the SME subcontractors might receive less assistance but these SMEs who in general, have limited resources may make more use of this assistance for better performance and enhancing their internal capabilities. As the subcontracting relationship endures over a period of time, subcontracting SMEs would be able to enhance their performance and develop their internal resources which in turn could be further used for their performance. If that is so, even when the SMEs are able to receive more assistance from TNCs and deliver higher performance, the contribution of this assistance for their performance would decrease. Thus this diminishing role of assistance from TNCs in the economic performance of subcontracting SMEs supports the fact that SME subcontractors are able to improve their internal capabilities and competitiveness through long term subcontracting relationships with TNCs. An assessment of the relative importance of SME factors which encourage subcontracting of the SMEs with TNCs indicated that inhouse R&D efforts and technological capability, frequent and proper communication, financial stability, skilled manpower, reputation of the subcontracting firm etc. are the most important factors, indicating the importance of inhouse/internal resources of the subcontracting firms. An assessment of the factors with respect to improvement needed for these SMEs revealed that the entrepreneurs/managers of the SMEs had realised the need to improve their inhouse resources and develop their technological capabilities with enhanced skilled manpower and better machinery. Given the importance of internal resources of SMEs for forging subcontracting relationships with TNCs, Indian SMEs need to place utmost priority for enhancing their own technical and manpower resources. Results of our study underlines the need for a policy thrust to expand the coverage of subcontracting involving local SMEs with domestic economy based global TNCs. The promotion of linkages of SMEs with TNCs depends on two factors: (i) the availability of local SMEs who have the prerequisite capabilities in terms of quality, delivery and cost and (ii) availability of information of such SMEs and their capabilities for TNCs. This context calls for the simultaneous strategy of competitiveness enhancement of local SMEs to the required level and providing information about the available capabilities of SMEs to TNCs.

A tranferência nuclear de células somáticas (TNCS) está sendo utilizada para produzir cavalos de elite. No entanto, durante este procedimento pode ocorrer a perfuração da zona pelúcida, levando, ocasionalmente, à secção da massa celular interna, e conseqüente derivação de gêmeos monozigóticos. Além de serem relatadas alterações no processo de imprinting genômico, que conduzem ao desenvolvimento de doenças. Com a descoberta da possibilidade de reprogramar as células somáticas a um estado de pluripotência (iPSCs), estas células passaram a ser muito utilizadas em pesquisas de neurociência. Contudo, também ocorrem modificações epigenéticas durante esta reprogramação celular. Portanto, nossas hipóteses são que os gêmeos eqüinos gerados pela TNCS podem levar às irregularidades no desenvolvimento do sistema nervoso. O padrão de metilação do SNRPN nas estruturas dos fetos muares clonados, e as células iPSCs são diferentes dos padrões encontrados nos muares analisados. A expressão dos genes SNRPN, Necdin e UBE3A são maiores no cérebro, enquanto a expressão do H19 é maior nas membranas extra-embrionárias. Em nosso estudo, obtivemos duas gestações gemelares equinas derivadas da TNCS, que foram interrompidas com 40 e 60 dias de gestação, e comparados com gestações eqüinas únicas de idade similar. Diferenças no comprimento entre os embriões gêmeos foram observadas aos 40 (2.0 e 2.2 cm 10%) e aos 60 (6,5 e 8,5 cm 24%) dias de gestação. Somente o plexo coróide do quarto ventrículo apresentou-se mais desenvolvido nos fetos com maior comprimento. Ao analisarmos fetos muares clonados em diferentes idades gestacionais e compará-los com muares, nos períodos embrionário, fetal e adulto, não foi observada diferença no padrão de metilação do gene SNRPN. No entanto, na décima passagem das células iPSC o padrão de metilação alterou, em relação aos muares estudados e ao padrão observado nos fibroblastos. Ao analisarmos os fetos clonados nas diferentes idades gestacionais observou-se no cérebro menor expressão dos gene H19 e UBE3A, e maior expressão do gene SNRPN. Contudo, a expressão do gene Necdin variou entre as estruturas estudadas. Em conclusão, apesar dos gêmeos eqüinos provenientes de TNCS diferirem quanto ao tamanho, morfologicamente são iguais. Dentre as estruturas cerebrais o plexo coróide se apresentou mais desenvolvido nos fetos de maior comprimento. Os fetos muares clonados não apresentaram diferença no padrão de metilação do gene SNRPN. No entanto, as iPSCs apresentaram alteração no padrão de metilação deste gene na décima passagem. Embora os genes SNRPN, Necdin e UBE3A sejam expressos no cérebro, o SNRPN apresentou-se prevalente nessa estrutura / The nuclear transfer of somatic cells (SCNT) is being used to produce elite horses. However, during this procedure can occur drilling of the zona pellucida, leading occasionally to the section of the inner cell mass, and subsequent derivation of monozygotic twins. Besides being related changes in genomic imprinting process, leading to the development of diseases. With the discovery of the possibility to reprogram somatic cells to a pluripotent state (iPSCs), these cells have become widely used in neuroscience research. However, also occur epigenetic changes during this cellular reprogramming. Therefore, our hypothesis is that equine twins caused by equine ART could lead to developmental irregularities of the nervous system. The patterns of SNRPN methylation in the structures of cloned mule fetuses and in iPSCs are different from the patterns found in the analyzed mules. And the expression of SNRPN, Necdin and UBE3A genes are higher in the brain, while the higher expression of H19 gene occurs in the extraembryonic membranes. In our study we derived two equine twin SCNT pregnancies that were interrupted at 40 and 60 days of gestation and compared to singleton fetuses of similar age. Differences in lengths between twin embryos were observed at both 40 (2.0 and 2.2 cm 10%) and 60 (6.5 and 8.5 cm 24%) days of gestation. Only the choroid plexus in the fourth ventricle more developed in the twins with the greatest length. Analyzing mules cloned fetuses at different gestational ages, and compare them with mules at embryonic, fetal and adult period; there was no difference in the pattern of methylation in SNRPN gene. However, in the tenth passage of the iPSCs the methylation pattern was altered in relation to the studied mules and the pattern observed in fibroblasts. When the cloned fetuses at different gestational ages were analyzed, the brain presented lower expression of H19 and UBE3A genes, and higher expression of SNRPN gene. However, the expression of Necdin gene varied among the structures studied. In conclusion, despite the twin horses from SCNT differ in size, they are morphologically identical. Among the brain structures the choroid plexus performed more developed in the fetuses of greater length. Cloned mules fetuses showed no difference in the pattern of methylation SNRPN gene. However, iPSCs have changes in the pattern of methylation of this gene in the tenth passage. Although SNRPN, Necdin and Ube3A genes are expressed in the brain, SNRPN is prevalent in this structure

代表國際合作的國際組織對於國家的行為有什麼影響？趨同或趨異？這個問題國關學界主要是新現實主義與新自由制度主義兩個主流理論在進行辯論；前者的研究顯示，國際組織或其它國際合作類型，只是主權國家（特別是強權國家）的工具，本身沒有獨立性，更遑論能制約國家行為或導引國家行為的協調一致。而後者從賽局理論推演出合作的可能與必要性，並認為制度（或建制）是有效維繫合作關係與調和各國行為，進而穩定國際秩序的方法。 然而，在世界貿易組織的架構下，國際貿易合作中食品衛生安全的相關衝突一直存在著。而在國際貨幣基金或世界銀行的架構下，糧食援助的合作似乎造成許多受援國境內的糧食危機未減反增。這樣的結果與新自由制度主義的論點不符，同時暗示著維繫國際合作的制度（或建制）可能具有不當性或偏重對特定法益的保護。另一方面，這樣的情況也無法透過新現實主義得到周延的解釋，因為合作中的權力爭奪並不是只單單發生在體系層次，並且最後影響或主導結果的行為者也非該理論向來強調的主權國家。 當前國際食品貿易的進/出口規範與具有結構調整性質的糧食援助機制究竟是過度保護了貿易自由，還是能為每一個個人可能遭遇的食品衛生風險或糧食危機提供必要之保障？這是本研究的首要顧念。透過個案研究與過程追蹤的方法，本文認為在無政府狀態2.0的環境下，國際食品貿易與糧食援助的制度（或建制）存在著法益失衡與不當性；是一種無法公允保障進口國消費者食品安全或受援國農民糧食主權的合作模式。這樣的結果在既有的國關理論得不到全面性的分析和解釋，因此文中提出「寧靜政治」的論述對之進行補充，說明擁有貿易出口利益的國家，其政府在國內層次如何因為寧靜政治現象而流失權力（退位），成為跨國公司的代理人，而非最高的主權享有者與治權行使者。同時，在體系層次，這些國家還為跨國公司爭取利益最大化，假國際合作之名攻佔食品進口國或糧食受援國的國內市場，釀成公共衛生或糧食主權上的危機。 研究結果顯示，跨國公司是造成國際制度（或建制）成為失衡天秤與喪失正當性的幕後原兇。也是在國際經貿這個議題領域中導致國家退位的關鍵行為者。從強調資本主義和貿易自由化的經濟學角度來看，這或許是市場機制正常的發揮，即使有人不樂見，也毋需過於擔心。然而，任何「自由」都不能是毫無限制地，也不該是無所節度的。全球經貿自由化與市場資本主義的發展若是繼續以目前的方式在運作與擴張，跨國公司累積的財富與權力就會在更多的議題領域超越或擊退主權國家，屆時除了食品衛生安全與糧食安全外，恐怕還會有更多攸關人類安全的法益在商品化的國際市場上漸漸淪為類奢侈財。 從理性選擇的角度來看，此種結果或許是國家在參與經貿合作前本能預見之事，但基於理性選擇，認為這是可以忍受的代價。然而，本文質疑國家（特別是小國）所付出的代價真能讓其在合作過程中換取到大於損失的利益。就像個案中台灣在進口美國牛肉的問題上，或墨西哥在糧食援助的計畫裡，這些國家並沒有在國際合作中得到預期的好處，反而失去了原本可以享有的衛生安全與糧食自主性。退一步言，假設這些國家真的從合作中得到大於損失之利益，那麼這些利益的分配在其國內層次上往往也不否符合公平與正義。毋寧，利益最後只是由特定少數的行為者所享有，但衛生安全風險或糧食基本權的不利益卻轉嫁由進口國或受援國的全體人民買單。

Transnational corporations are playing an important role in the global economy of today. Many of these corporations have great economic resources and have the possibility of contributing to the development of societies in developing states. At the same time, in their search for profit, the activities of TNCs have proven fatal to some of the individuals employed by them, or otherwise in contact with their activities. Within the international legal framework, corporations are not traditionally treated as subjects and if a TNC allocates its production to a state with lax human rights protection, no binding international standards exist to regulate the conduct of the corporation.  In my thesis I will assess the position of TNCs under the present core human rights instruments and soft law initiatives. I will also analyze a draft treaty text produced by the Intergovernmental Working Group on Business and Human Rights, released in July 2018, to reach a conclusion on whether such an instrument would affect the international legal status of TNCs and provide a more robust protection of international human rights.