965 resultados para internal transcribed spacer (ITS)


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The addition of a capped mini-exon [spliced leader (SL)] through trans-splicing is essential for the maturation of RNA polymerase (pol) II-transcribed polycistronic pre-mRNAs in all members of the Trypanosomatidae family. This process is an inter-molecular splicing reaction that follows the same basic rules of cis-splicing reactions. In this study, we demonstrated that mini-exons were added to precursor ribosomal RNA (pre-rRNA) are transcribed by RNA pol I, including the 5' external transcribed spacer (ETS) region. Additionally, we detected the SL-5' ETS molecule using three distinct methods and located the acceptor site between two known 5' ETS rRNA processing sites (A' and A1) in four different trypanosomatids. Moreover, we detected a polyadenylated 5' ETS upstream of the trans-splicing acceptor site, which also occurs in pre-mRNA trans-splicing. After treatment with an indirect trans-splicing inhibitor (sinefungin), we observed SL-5' ETS decay. However, treatment with 5-fluorouracil (a precursor of RNA synthesis that inhibits the degradation of pre-rRNA) led to the accumulation of SL-5' ETS, suggesting that the molecule may play a role in rRNA degradation. The detection of trans-splicing in these molecules may indicate broad RNA-joining properties, regardless of the polymerase used for transcription.

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Background: Parasites of the Leishmania genus alternate between the flagellated extracellular promastigote stage and intracellular amastigotes. Here we report the characterization of a Leishmania isolate, obtained from a cutaneous leishmaniasis patient, which presents peculiar morphological features. Methods: The parasite was cultured in vitro and characterized morphologically using optical and electron microscopy. Identification was performed based on monoclonal antibodies and internal ribosomal spacer typing. In vitro macrophage cultures, murine experimental models and sand fly infections were used to evaluate infectivity in vitro and in vivo. Results: The isolate was identified as Leishmania (Viannia) braziliensis. In the atypical promastigotes grown in culture, a short flagellum surrounded or interrupted by a protuberance of disorganized material was observed. A normal axoneme was present close to the basal body but without elongation much further outside the flagellar pocket. A disorganized swelling at the precocious end of the axoneme coincided with the lack of a paraflagellar rod structure. The isolate was able to infect macrophages in vitro, induce lesions in BALB/c mice and infect Lutzomyia longipalpis. Conclusions: Notwithstanding the lack of an extracellular flagellum, this isolate infects macrophages in vitro and produces lesions when inoculated into mice. Moreover, it is able to colonize phlebotomine sand flies. Considering the importance attributed to the flagellum in the successful infection and survival of Leishmania in the insect midgut and in the invasion of macrophages, these findings may bring new light into the infectious mechanisms of L. (V.) braziliensis.

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Various organisms have been characterized by molecular methods, including fungi of the genus Cryptococcus. The purposes of this study were: to determine the discriminatory potential of the RAPD (Random Amplified Polymorphic DNA) primers, the pattern of similarity of the Cryptococcus species, and discuss their useful application in epidemiological studies. We analyzed 10 isolates of each specie/group: C. albidus, C. laurentii complex, C. neoformans var. grubii, all from environmental source, and two ATCC strains, C. neoformans var. grubii ATCC 90112, and C. neoformans var. neoformans ATCC 28957 by RAPD-PCR using the primers CAV1, CAV2, ZAP19, ZAP20, OPB11 and SEQ6. The primers showed a good discriminatory power, revealing important differences between them and between species; the SEQ6 primer discriminated a larger number of isolates of three species. Isolates of C. laurentii showed greater genetic diversity than other species revealed by all six primers. Isolates of C. neoformans were more homogeneous. Only the primer CAV2 showed no amplification of DNA bands for C. albidus. It was concluded that the use of limited number of carefully selected primers allowed the discrimination of different isolates, and some primers (e. g., CAV2 for C. albidus) may not to be applied to some species.

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Under the Chinese National Antarctic Research Expedition program in 2006, the annual thermal mass balance of landfast ice in the vicinity of Zhongshan Station, Prydz Bay, east Antarctica, was investigated. Sea ice formed from mid-February onward, and maximum ice thickness occurred in late November. Snow cover remained thin, and blowing snow caused frequent redistribution of the snow. The vertical ice salinity showed a 'question-mark-shaped' profile for most of the ice growth season, which only turned into an 'I-shaped' profile after the onset of ice melt. The oceanic heat flux as estimated from a flux balance at ice-ocean interface using internal ice temperatures decreased from 11.8 (±3.5) W/m**2 in April to an annual minimum of 1.9 (±2.4) W/m**2 in September. It remained low through late November, in mid-December it increased sharply to about 20.0 W/m**2. Simulations applying the modified versions of Stefan's law, taking account the oceanic heat flux and ice-atmosphere coupling, compare well with observed ice growth. There was no obvious seasonal cycle for the thermal conductivity of snow cover, which was also derived from internal ice temperatures. Its annual mean was 0.20 (±0.04) W/m/°C.

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El cáncer de próstata es el tipo de cáncer con mayor prevalencia entre los hombres del mundo occidental y, pese a tener una alta tasa de supervivencia relativa, es la segunda mayor causa de muerte por cáncer en este sector de la población. El tratamiento de elección frente al cáncer de próstata es, en la mayoría de los casos, la radioterapia externa. Las técnicas más modernas de radioterapia externa, como la radioterapia modulada en intensidad, permiten incrementar la dosis en el tumor mientras se reduce la dosis en el tejido sano. Sin embargo, la localización del volumen objetivo varía con el día de tratamiento, y se requieren movimientos muy pequeños de los órganos para sacar partes del volumen objetivo fuera de la región terapéutica, o para introducir tejidos sanos críticos dentro. Para evitar esto se han desarrollado técnicas más avanzadas, como la radioterapia guiada por imagen, que se define por un manejo más preciso de los movimientos internos mediante una adaptación de la planificación del tratamiento basada en la información anatómica obtenida de imágenes de tomografía computarizada (TC) previas a la sesión terapéutica. Además, la radioterapia adaptativa añade la información dosimétrica de las fracciones previas a la información anatómica. Uno de los fundamentos de la radioterapia adaptativa es el registro deformable de imágenes, de gran utilidad a la hora de modelar los desplazamientos y deformaciones de los órganos internos. Sin embargo, su utilización conlleva nuevos retos científico-tecnológicos en el procesamiento de imágenes, principalmente asociados a la variabilidad de los órganos, tanto en localización como en apariencia. El objetivo de esta tesis doctoral es mejorar los procesos clínicos de delineación automática de contornos y de cálculo de dosis acumulada para la planificación y monitorización de tratamientos con radioterapia adaptativa, a partir de nuevos métodos de procesamiento de imágenes de TC (1) en presencia de contrastes variables, y (2) cambios de apariencia del recto. Además, se pretende (3) proveer de herramientas para la evaluación de la calidad de los contornos obtenidos en el caso del gross tumor volumen (GTV). Las principales contribuciones de esta tesis doctoral son las siguientes: _ 1. La adaptación, implementación y evaluación de un algoritmo de registro basado en el flujo óptico de la fase de la imagen como herramienta para el cálculo de transformaciones no-rígidas en presencia de cambios de intensidad, y su aplicabilidad a tratamientos de radioterapia adaptativa en cáncer de próstata con uso de agentes de contraste radiológico. Los resultados demuestran que el algoritmo seleccionado presenta mejores resultados cualitativos en presencia de contraste radiológico en la vejiga, y no distorsiona la imagen forzando deformaciones poco realistas. 2. La definición, desarrollo y validación de un nuevo método de enmascaramiento de los contenidos del recto (MER), y la evaluación de su influencia en el procedimiento de radioterapia adaptativa en cáncer de próstata. Las segmentaciones obtenidas mediante el MER para la creación de máscaras homogéneas en las imágenes de sesión permiten mejorar sensiblemente los resultados de los algoritmos de registro en la región rectal. Así, el uso de la metodología propuesta incrementa el índice de volumen solapado entre los contornos manuales y automáticos del recto hasta un valor del 89%, cercano a los resultados obtenidos usando máscaras manuales para el registro de las dos imágenes. De esta manera se pueden corregir tanto el cálculo de los nuevos contornos como el cálculo de la dosis acumulada. 3. La definición de una metodología de evaluación de la calidad de los contornos del GTV, que permite la representación de la distribución espacial del error, adaptándola a volúmenes no-convexos como el formado por la próstata y las vesículas seminales. Dicha metodología de evaluación, basada en un nuevo algoritmo de reconstrucción tridimensional y una nueva métrica de cuantificación, presenta resultados precisos con una gran resolución espacial en un tiempo despreciable frente al tiempo de registro. Esta nueva metodología puede ser una herramienta útil para la comparación de distintos algoritmos de registro deformable orientados a la radioterapia adaptativa en cáncer de próstata. En conclusión, el trabajo realizado en esta tesis doctoral corrobora las hipótesis de investigación postuladas, y pretende servir como cimiento de futuros avances en el procesamiento de imagen médica en los tratamientos de radioterapia adaptativa en cáncer de próstata. Asimismo, se siguen abriendo nuevas líneas de aplicación futura de métodos de procesamiento de imágenes médicas con el fin de mejorar los procesos de radioterapia adaptativa en presencia de cambios de apariencia de los órganos, e incrementar la seguridad del paciente. I.2 Inglés Prostate cancer is the most prevalent cancer amongst men in the Western world and, despite having a relatively high survival rate, is the second leading cause of cancer death in this sector of the population. The treatment of choice against prostate cancer is, in most cases, external beam radiation therapy. The most modern techniques of external radiotherapy, as intensity modulated radiotherapy, allow increasing the dose to the tumor whilst reducing the dose to healthy tissue. However, the location of the target volume varies with the day of treatment, and very small movements of the organs are required to pull out parts of the target volume outside the therapeutic region, or to introduce critical healthy tissues inside. Advanced techniques, such as the image-guided radiotherapy (IGRT), have been developed to avoid this. IGRT is defined by more precise handling of internal movements by adapting treatment planning based on the anatomical information obtained from computed tomography (CT) images prior to the therapy session. Moreover, the adaptive radiotherapy adds dosimetric information of previous fractions to the anatomical information. One of the fundamentals of adaptive radiotherapy is deformable image registration, very useful when modeling the displacements and deformations of the internal organs. However, its use brings new scientific and technological challenges in image processing, mainly associated to the variability of the organs, both in location and appearance. The aim of this thesis is to improve clinical processes of automatic contour delineation and cumulative dose calculation for planning and monitoring of adaptive radiotherapy treatments, based on new methods of CT image processing (1) in the presence of varying contrasts, and (2) rectum appearance changes. It also aims (3) to provide tools for assessing the quality of contours obtained in the case of gross tumor volume (GTV). The main contributions of this PhD thesis are as follows: 1. The adaptation, implementation and evaluation of a registration algorithm based on the optical flow of the image phase as a tool for the calculation of non-rigid transformations in the presence of intensity changes, and its applicability to adaptive radiotherapy treatment in prostate cancer with use of radiological contrast agents. The results demonstrate that the selected algorithm shows better qualitative results in the presence of radiological contrast agents in the urinary bladder, and does not distort the image forcing unrealistic deformations. 2. The definition, development and validation of a new method for masking the contents of the rectum (MER, Spanish acronym), and assessing their impact on the process of adaptive radiotherapy in prostate cancer. The segmentations obtained by the MER for the creation of homogenous masks in the session CT images can improve significantly the results of registration algorithms in the rectal region. Thus, the use of the proposed methodology increases the volume overlap index between manual and automatic contours of the rectum to a value of 89%, close to the results obtained using manual masks for both images. In this way, both the calculation of new contours and the calculation of the accumulated dose can be corrected. 3. The definition of a methodology for assessing the quality of the contours of the GTV, which allows the representation of the spatial distribution of the error, adapting it to non-convex volumes such as that formed by the prostate and seminal vesicles. Said evaluation methodology, based on a new three-dimensional reconstruction algorithm and a new quantification metric, presents accurate results with high spatial resolution in a time negligible compared to the registration time. This new approach may be a useful tool to compare different deformable registration algorithms oriented to adaptive radiotherapy in prostate cancer In conclusion, this PhD thesis corroborates the postulated research hypotheses, and is intended to serve as a foundation for future advances in medical image processing in adaptive radiotherapy treatment in prostate cancer. In addition, it opens new future applications for medical image processing methods aimed at improving the adaptive radiotherapy processes in the presence of organ’s appearance changes, and increase the patient safety.

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Human P-glycoprotein (Pgp) confers multidrug resistance to cancer cells by ATP-dependent extrusion of a great many structurally dissimilar hydrophobic compounds. The manner in which Pgp recognizes these different substrates is unknown. The protein shows internal homology between its N- and C-terminal halves, each comprised of six putative transmembrane helices and a consensus ATP binding/utilization site. Photoactive derivatives of certain Pgp substrates specifically label two regions, one on each half of the protein. In this study, using [125I]iodoarylazidoprazosin ([125I]IAAP), a photoactive analog of prazosin, we have demonstrated the presence of two nonidentical drug-interaction sites within Pgp. Taking advantage of a highly susceptible trypsin cleavage site in the linker region of Pgp, we characterized the [125I]IAAP binding to the N- and C-terminal halves. cis(Z)-Flupentixol, a modulator of Pgp function, preferentially increased the affinity of [125I]IAAP for the C-terminal half of the protein (C-site) by reducing the Kd from 20 to 6 nM without changing the labeling or affinity (Kd = 42–46 nM) of the N-terminal half (N-site). Also, the concentration of vinblastine (Pgp substrate) and cyclosporin A (Pgp modulator) required for 50% inhibition of [125I]IAAP binding to the C-site was increased 5- to 6-fold by cis(Z)-flupentixol without any effect on the N-site. In addition, [125I]IAAP binding to the N-site was less susceptible than to C-site to inhibition by vanadate which blocks ATP hydrolysis and drug transport. These data demonstrate the presence of at least two nonidentical substrate interaction sites in Pgp.

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Introduction. In 2003, Iraq was invaded by the US coalition forces that ousted Saddam Hussein’s regime from power before occupying the whole country. The intension, declared by the then American George W. Bush, was to ‘build a decent and democratic society at the centre of the Middle East’ that ‘will become a place of progress and peace.’1 In 2014, three years after the withdrawal of the last American soldier, however, it is difficult to overestimate or exaggerate what is at stake. National unity and territorial integrity have never been so seriously threatened since the country is experiencing the internal fighting in its modern history. Many parts of Iraq, including the northern oil city of Kirkuk, long claimed as an integral part of the semi-autonomous region of Kurdistan, are out of the control of the central government. Large areas in the north including the strategic city of Mosul were seized by the fighters of the Islamic State, an Al-Qaeda offshoot, formerly known as ISIS, who threatened to invade the Kurdistan region before being attacked by airstrikes by the US. They proclaimed a caliphate on both sides of the border with Syria, where they also control vast territory.

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In the EU circuit (especially the European Parliament, the Council and Coreper) as well as in national parliaments of the EU Member States, one observes a powerful tendency to regard 'subsidiarity' as a 'political' issue. Moreover, subsidiarity is frequently seen as a one-way street : powers going 'back to' Member States. Both interpretations are at least partly flawed and less than helpful when looking for practical ways to deal with subsidiarity at both EU and Member states' levels. The present paper shows that subsidiarity as a principle is profoundly 'functional' in nature and, hence, is and must be a two-way principle. A functional subsidiarity test is developed and its application is illustrated for a range of policy issues in the internal market in its widest sense, for equity and for macro-economic stabilisation questions in European integration. Misapplications of 'subsidiarity' are also demonstrated. For a good understanding, subsidiarity being a functional, two-way principle neither means that elected politicians should not have the final (political!) say (for which they are accountable), nor that subsidiarity tests, even if properly conducted, cannot and will not be politicised once the results enter the policy debate. Such politicisation forms a natural run-up to the decision-making by those elected for it. But the quality and reasoning of the test as well as structuring the information in a logical sequence ( in accordance with the current protocol and with the one in the constitutional treaty) is likely to be directly helpful for decisionmakers, confronted with complicated and often specialised proposals. EU debates and decision-making is therefore best served by separating the functional subsidiarity test (prepared by independent professionals) from the final political decision itself. If the test were accepted Union-wide, it would also assist national parliaments in conducting comparable tests in a relatively short period, as the basis for possible joint action (as suggested by the constitutional treaty). The core of the paper explains how the test is formulated and applied. A functional approach to subsidiarity in the framework of European representative democracy seeks to find the optimal assignment of regulatory or policy competences to the various tiers of government. In the final analysis, this is about structures facilitating the highest possible welfare in the Union, in the fundamental sense that preferences and needs are best satisfied. What is required for such an analysis is no less than a systematic cost/benefit framework to assess the (de)merits of (de)centralisation in the EU.

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Mutual recognition is one of the most appreciated innovations of the EU. The idea is that one can pursue market integration, indeed "deep' market integration, while respecting 'diversity' amongst the participating countries. Put differently, in pursuing 'free movement' for goods, mutual recognition facilitates free movement by disciplining the nature and scope of 'regulatory barriers', whilst allowing some degree of regulatory discretion for EU Member States. This BEER paper attempts to explain the rationale and logic of mutual recognition in the EU internal goods market, its working in actual practice for about three decades now, culminating in a qualitative cost/benefit analysis and its recent improvement in terms of 'governance' in the so-called New Legislative Framework (first denoted as the 2008 Goods package) thereby ameliorating the benefits/costs ratio. For new (in contrast to existing) national regulation, the intrusive EU procedure to impose mutual recognition is presented as well, with basic data so as to show its critical importance to keep the internal goods market free. All this is complemented by a short summary of the scant economic literature on mutual recognition. Subsequently, the analysis is extended to the internal market for services. This is done in two steps, first by reminding the debate on the origin principle (which goes further than mutual recognition EU-style) and how mutual recognition works under the horizontal services directive. This is followed by a short section on how mutual recognition works in vertical (i.e. sectoral) services markets.

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This study examined whether supervision characteristics impacted on mental health practice and morale, and developed a new Supervision Attitude Scale (SAS). Telephone surveys were conducted with a representative sample of 272 staff from public mental health services across Queensland. Although supervision was widely received and positively rated, it had low average intensity, and assessment and training of skills was rarely incorporated. Perceived impact on practice was associated with acquisition of skills and positive attitudes to supervisors, but extent of supervision was related to impact only if it was from within the profession. Intention to resign was unrelated to extent of supervision, but was associated with positive attitudes to supervisors, accessibility, high impact, and empathy or praise in supervision sessions. The SAS had high internal consistency, and its intercorrelations were consistent with it being a measure of relationship positivity. The study supported the role of supervision in retention and in improving practice. It also highlighted supervision characteristics that might be targeted in training, and provided preliminary data on a new measure.

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Em virtude da necessidade de inovação das empresas, gerada pelas transformações tecnológicas e pela velocidade da globalização, uma tendência mundial começa a surgir: o enfraquecimento de mão de obra no setor industrial e um considerável aumento no setor de serviços. Essa realidade pressupõe a criação de várias empresas, na sua maioria, de pequeno porte, o que dá especial importância a esse estudo, pois aliado à criação de novas empresas, o seu índice de mortalidade extremamente alto indica um problema a ser investigado. Portanto, a presente pesquisa procura sugerir estratégias de marketing eficientes como fatores amenizadores desse problema, e um caminho possível para a manutenção e o desenvolvimento das pequenas empresas. Criou-se, então, um modelo para a aplicação de estratégias de marketing, específico para empresas de pequeno porte. O modelo desenvolvido neste trabalho parte de uma revisão crítica da bibliografia referente às peculiaridades das pequenas empresas, como também das práticas de marketing, e busca correlacioná -las, através de uma pesquisa descritiva no formato de estudo de caso e determinando quais as condições para o uso do marketing nas pequenas empresas, quais as estratégias compatíveis e quais as mudanças necessárias à sua prática.

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This research focuses on two groups of local companies; namely, high-growth local companies and other local companies, to examine and compare the influence of utilising governmental initiatives, servicing foreign MNCs and internationalisation on their strategic planning process. The theme of this thesis argues that the approach of an organisation towards strategic planning is not only determined by the internal influences; namely, its firm size and the planning behaviour and attitude of an entrepreneur, as revealed in the literature, but it can also be affected by external influences. The theoretical contribution of this research determines this unique situation in Singapore, and tests the robustness of the conventional models of planning in smaller companies. As a result of the external influences, this study reveals that local companies are more likely to undertake a much more formal strategic planning than the conventional Western literature and models would indicate. High-growth local companies, in comparison, however, had undertaken a more formal and rigorous strategic planning process than other local companies.

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This dissertation uncovers the path of Pernambuco’s independent cinema of the Retomada, of its production in the second half of the 1990s until mid 2012 when the sector manages to secure a funding edict (Funcultura) for its films and at the same time establishes a few symbolic mechanisms which contributed to the consolidation of a new cycle, the post-Retomada, namely: cinephilia and brodagem. Because it is a cinematography (pernambucana) outside the Rio-São Paulo axis, that is, outside the great center of the brazillian culture industry, the cinema produced in Pernambuco built a very specific modus operandi, in which a material basis, the consolidation of Funcultura, intercrossed with two symbolic practices feeding on each other, cinephilia and brodagem. In this regard, we seek not to establish the material perspective as the primary determination as opposed to the symbolic one. Futhermore, we followed the path of cinephilia in its direct commucation with the brodagem, and how both these logics pressured authorities to institute an incentive funding separated from other artforms in the state. From this point of view, we attempted to trace the trajectory of this cinematography in its external nuances, but also taking into account the internal nuances of its works of art (films). Following this, we created categories to define and distinguish the nuances of the Retomada and post-Retomada productions regarding ethical and aesthetical choices made by directors from both generations.

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This dissertation uncovers the path of Pernambuco’s independent cinema of the Retomada, of its production in the second half of the 1990s until mid 2012 when the sector manages to secure a funding edict (Funcultura) for its films and at the same time establishes a few symbolic mechanisms which contributed to the consolidation of a new cycle, the post-Retomada, namely: cinephilia and brodagem. Because it is a cinematography (pernambucana) outside the Rio-São Paulo axis, that is, outside the great center of the brazillian culture industry, the cinema produced in Pernambuco built a very specific modus operandi, in which a material basis, the consolidation of Funcultura, intercrossed with two symbolic practices feeding on each other, cinephilia and brodagem. In this regard, we seek not to establish the material perspective as the primary determination as opposed to the symbolic one. Futhermore, we followed the path of cinephilia in its direct commucation with the brodagem, and how both these logics pressured authorities to institute an incentive funding separated from other artforms in the state. From this point of view, we attempted to trace the trajectory of this cinematography in its external nuances, but also taking into account the internal nuances of its works of art (films). Following this, we created categories to define and distinguish the nuances of the Retomada and post-Retomada productions regarding ethical and aesthetical choices made by directors from both generations.

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Este artículo se enfoca en el estudio del período que corre entre 1935 y 1951 para destacar una de las empresas más importantes del PC a nivel mundial, con substanciales réditos en Argentina: la organización de las mujeres. En particular, nuestro interrogante es de qué modo un partido tradicionalmente considerado de estructuración directa y unitaria, profesional, eminentemente concentrado en la línea de formación ideológica por sobre la electoral, caracterizado por un perfil obrero y fuerte en términos de organización administrativa interna -particularmente, debido a su característica celular-, encuentra límites a esa propia caracterización organizacional en tanto delinea una política de intervención coyuntural que lo coloca en alianzas con otros actores y modifica, con ello, sus propios caracteres. El corpus documental de esta investigación incluye documentos y prensa partidaria (comunista y de otras agrupaciones políticas o sociales); archivos personales, autobiografías y entrevistas. La codificación permitió un ordenamiento de los datos extraídos de los documentos relevados, agrupándolos, según similitudes o diferencias relevantes en categorías que hicieron posible su análisis