19 resultados para Homology directed repair

em Brock University, Canada


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The purpose of this correlational study was to investigate the relationship between the degree of self-directed learning readiness and stress for level one nursing students and level two nursing students. One hundred female nursing students participated in the study who were attending an Ontario Community College. Data were collected from the main nursing campus and the satellite nursing campus using the random sample method. Instruments used were said to be valid and reliable for testing self-directed learning readiness and stress respectively. Data were analyzed using frequency response to each item, means and standard deviation, and the Pearson product correlation between selfdirected learning readiness and stress. The results of the study show that there is a difference in the relationship between the degree of self-directed learning readiness and stress between the level one nursing students and the level two nursing students. Such results will be of particular interest to nursing instructors and administrators when planning for delivery of programs to such students.

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Mitochondria have an important role in cell metabolism, being the major site of ATP production via oxidative phosphorylation (OXPHOS). Accumulation of mtDNA mutations have been linked to the development of respiratory dysfunction, apoptosis, and aging. Base excision repair (BER) is the major and the only certain repair pathway existing in mitochondria that is in responsible for removing and repairing various base modifications as well as abasic sites (AP sites). In this research, Saccharomyces cerevisiae (S. cerevisiae) BER gene knockout strains, including 3 single DNA glycosylase gene knockout strains and Ap endonuclease (Apn 1 p) knockout strain were used to examine the importance of this DNA repair pathway to the maintenance of respiratory function. Here, I show that individual DNA glycosylases are nonessential in maintenance of normal function in yeast mitochondria, corroborating with previous research in mammalian experimental models. The yeast strain lacking Apn 1 p activity exhibits respiratory deficits, including inefficient and significantly low intracellular ATP level, which maybe due to partial uncoupling of OXPHOS. Growth of this yeast strain on respiratory medium is inhibited, but no evidence was found for increased ROS level in Apn 1 p mitochondria. This strain also shows an increased cell size, and this observation combined with an uncoupled OXPHOS may indicate a premature aging in the Apnlp knockout strain, but more evidence is needed to support this hypothesis. However, the BER is necessary for maintenance of mitochondrial function in respiring S.cerevisiae.

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~ This study focuses on the process of self-directed learning that individuals go through as they adapt to new work situations. This is a study of how one critical incident, specifically the transition from a traditional office structure to a home office structure, affected employees and what their learning process was as they adapted to the new environment. This study has 3 educational foundations: adult learning, self-directed learning, and the social context from which the learning will occur. Six women and 2 men were interviewed approximately 1 year following the transition. Analysis of the data revealed 5 themes of: impacts of the self-directed environment on participants' personal lives, their roles, skill set, productivity, and the physical environment; support offered by the organization, family, and office administration; personal development, specific learning needs, and personal skills; boundaries as they relate to family and work; and skill set and orientation requirements of new home office employees. The findings revealed the learning processes of the 8 participants. The learning processes of these participants were discussed within a theoretical framework of the learners, their immediate surroundings, and the larger social environment. The results indicated that the transition from a directed work environment to a self directed work environment is a complex, interrelated process. An element found throughout the theoretical framework is that of control. A second critical element is the need for participants to have a clearly defined work role and an opportunity to engage in discussion with peers and the community. Further findings reinforced the importance of climate and found that the physical environment is a key factor in a successful selfdirected work environment. The findings of this study revealed that no one factor makes an individual function successfully in a self-directed work environment, but that it is a complex interplay among the leamer, their immediate surroundings, and the social environment that will have the greatest impact on success. Recommendations are made which can be used to guide organizational leaders in facilitating employees' transition from a directed to a self-directed work environment. Additionally, recommendations are made for further research in the area of self-directed work environments.

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Ontario Colleges of Applied Arts and Technology (CAATs) are currently in the process of restructuring to ensure quality, accountability, and accessibility of college education. References to learner involvement and self-directed learning are prevalent. "Alternative delivery" and "paradigm shift" are current buzzwords within the Ontario CAAT system as an environment is created supportive of change. Instability of funding has also dictated a need for change. Therefore, a focus has become quality of learning with less demand on public resources. This qualitative case study was conducted at an Ontario CAAT to gather descriptive, perceptual data from post-secondary community college educators who were identified as supportive of self-directed learning and from post-secondary, traditional-aged college students who were perceived by their educators to be selfdirected learners. This college was selected because of initiatives to modify its academic paradigm to encourage what was reputed in the Ontario CAAT system to be self-directed learning. The purpose of this study was to investigate how postsecondary, traditional-aged college students and their educators perceive self-directed learning as part of the teaching-learning experience within a community college setting. Educator participants of the study were selected based on the results of a teaching and learning survey intended to identify educators supportive of self-directed learning. A total of 317 surveys were distributed to every full-time educator at the sample college; 192 completed surveys were returned for a return rate of 61 %. Of these, 8% indicated instructional beliefs and values supportive of self-directed learning. A purposive sample of six educators was selected using a maximulp variation sampling strategy. A network selection sampling strategy was used to select a purposive sample of seven post-secondary students who were identified by the sample educators as selfdirected learners. The results of the study show that students and educators have similar perspectives and operating definitions of self-directed learning and all participants believe they either practice or facilitate self-directed learning. However, their perspectives and practices are not consistent with the literature which emphasizes learner autonomy or control in course structure and content. A central characteristic of the participants represented in this study is the service-oriented professions with which each is associated. Experientiallearning opportunities were highly valued for the options provided in increasing learner independence and competencies in reflective practice. Although there were discrepancies between espoused theory and theory in practice in terms of course structure, the process of self-directed learning was being practiced and supported outside the classroom structure in clinical settings, labs and related experiences.

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The concept of self-directed learning was examined from the nurse educators' perspective. One structured interview, lasting between one and one-and-a half hours, was conducted with each of 14 nurse educators in two community college continuing education programs in nursing. One community college program encourages selfdirected learning; the other encourages self-study and active participation in the teaching/learning process. All 14 interviews were tape-recorded and transcribed verbatim. Verbatim transcripts were analyzed for themes, patterns, and relationships utilizing analyst-constructed typologies. six prerequisites or necessary conditions for facilitating self-directed learning in a community college continuing education program in nursing were identified. ~he crucial issue in facilitating self-directed learning was found to be the issue of teacher-control.

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This study addressed the problem of instructor support for self-directed learning, specifically, learner-directed program planning, within a classroom setting in higher education. A combination of survey, interview, document analysis, and observation was used to assess and evaluate the attitudes and practices of a sample of full-time faculty at an Ontario university. Eighty-seven percent of the study sample reported instructional beliefs, values, and expectations that were not supportive of self-directed learning, especially in terms of student participation in program planning. Planning was seen as the responsibility of the instructor. Instructors were least open to student participation in the planning of the evaluation of learning. However, there was considerable stated support for other of the basic principles of adult education. The remaining 13% of the study sample reported instructional beliefs, values, and expectations that were fully supportive of self-directed learning. Instructional practices were analyzed in relation to the instructors' stated beliefs. Although practices reflected, in many instances, instructors' statements of support, there were some significant discrepancies between apparent support for the concept of self-directed learning and actual classroom practice. Both beliefs and practice were compared to a research model of self-directed learning. Most instructors did not have a concept of self-directed learning as comprehensive as that described in the research model. Instructor support for self-directed learning was profoundly influenced by the university setting. It was concluded that more strenuous attempts to research, enhance, and promote instructional and institutional support for self-directed learning in higher education are warranted.

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The neuropeptide Th1RFamide with the sequence Phe-Met-Arg-Phe-amide was originally isolated in the clam Macrocallista nimbosa (price and Greenberg, 1977). Since its discovery, a large family ofFl\1RFamide-related peptides termed FaRPs have been found to be present in all major animal phyla with functions ranging from modulation of neuronal activity to alteration of muscular contractions. However, little is known about the genetics encoding these peptides, especially in invertebrates. As FaRP-encoding genes have yet to be investigated in the invertebrate Malacostracean subphylum, the isolation and characterization ofFaRP-encoding DNA and mRNA was pursued in this project. The immediate aims of this thesis were: (1) to amplify mRNA sequences of Procambarus clarkii using a degenerate oligonucleotide primer deduced from the common amino acid sequence ofisolated Procambarus FaRPS, (2) to determine if these amplification products encode FaRP gene sequences, and (3) to create a selective cDNA library of sequences recognized by the degenerate oligonucleotide primer. The polymerase chain reaction - rapid amplification of cDNA ends (PCR-RACE) is a procedure in which a single gene-specific primer is used in conjunction with a generalized 3' or 5' primer to amplify copies ofthe region between a single point in the transcript and the 3' or 5' end of cDNA of interest (Frohman et aI., 1988). PCRRACE reactions were optimized with respect to primers used, buffer composition, cycle number, nature ofgenetic substrate to be amplified, annealing, extension and denaturation temperatures and times, and use of reamplification procedures. Amplification products were cloned into plasmid vectors and recombinant products were isolated, as were the recombinant plaques formed in the selective cDNA library. Labeled amplification products were hybridized to recombinant bacteriophage to determine ligated amplification product presence. When sequenced, the five isolated PCR-RACE amplification products were determined not to possess FaRP-encoding sequences. The 200bp, 450bp, and 1500bp sequences showed homology to the Caenorhabditis elegans cosmid K09A11, which encodes for cytochrome P450; transfer-RNA; transposase; and tRNA-Tyr, while the 500bp and 750bp sequences showed homology with the complete genome of the Vaccinia virus. Under the employed amplification conditions the degenerate oligonucleotide primer was observed to bind to and to amplify sequences with either 9 or 10bp of 17bp identity. The selective cDNA library was obselVed to be of extremely low titre. When library titre was increased, white. plaques were isolated. Amplification analysis of eight isolated Agt11 sequences from these plaques indicated an absence of an insertion sequence. The degenerate 17 base oligonucleotide primer synthesized from the common amino acid sequence ofisolated Procambarus FaRPs was thus determined to be non-specific in its binding under the conditions required for its use, and to be insufficient for the isolation and identification ofFaRP-encoding sequences. A more specific primer oflonger sequence, lower degeneracy, and higher melting temperature (TJ is recommended for further investigation into the FaRP-encoding genes of Procambarlls clarkii.

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This correlational study was designed to investigate the relationship between self-directed learning and personality type. A sample of 133 graduate and undergraduate education students completed the MBTI and the SDLRS. Two hypotheses were examined: (a) scores on the intuitive scale will account for a significant amount of the variance in the prediction of selfdirected learning readiness and, (b) scores on the introverted scale will account for a significant amount of the variance in self-directed learning readiness. Stepwise multiple regression analyses indicated that psychological type accounts for 28% of the variance in self-directed learning. Support for the first hypothesis was found with 15% of the variance in selfdirected learning accounted for by intuition. The second hypothesis was not supported. Introversion accounted for 13% of the variance but in a negative manner. Results of this study indicate that personality type does influence the ability of the learner to be self-directed in studies. These findings add another dimension for the adult educator to consider when attempting to develop self-directedness in learners.

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The study determined students' perceptions of self-directed learning in their courses. Tests to assess perceptions are not being used in many programs. Assessments such as the Self-Directed Readiness Scale (SDLRS) and the Oddi continuing Learning Inventory (OCLI) have weaknesses that may have affected the use of tests. In this study, the creation of the Self-Directed Learning Test (SDLT) monitored students' perceptions by addressing what students were told before registration, how much input students had in developing the structure of the course, how much input students have in determining the evaluation for the course, what style of learning is taking place, and the characteristics of learning found among students. Fifty-one students in the pre-service program at Brock University completed the SDLT. Results showed that 47.1% of the sample liked self-directed learning. Several students who stated that they did not like selfdirected learning did not know what self-directed learning was. Results supported Brookfield's (1986) claim for more education on what self-directed learning is. The study did not support Knowles' (1980) assumption that adult students know and want to follow self-directed approaches to learning. The SDLT is a good method for monitoring self-directed learning and how students perceive their courses.

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Surface proteinaceous fibrils, termed fimbriae, were first identified on gram negative bacteria in the 1940s. Fungal fimbriae, discovered some 25 years later, are found on members of all fungal classes. In the present study, polyclonal antiserum raised against the fimbrial proteins of U. vio/acea were used in order to identify antigenically related proteins from Coprinus cinereus and Schizophy//um commune. Two polypeptides with molecular masses of 37 and 39 kDa from C. cinereus were observed and confirm earlier results. A single previously unidentified 50 kDa polypeptide in S. commune crossreacted with the antiserum. The 50 kDa protein was found to consist of 3 isoforms with isoelectric points ranging from 5.6 to 5.8. A fimbrial cDNA derived from U. vio/acea was used to identify DNA restriction fragments from C. cinereus and S. commune showing homology to the fimbrial transcript of U. vio/acea. Heterologous hybridization with this cDNA was used in order to screen a C. cinereus genomic DNA library. A single clone, A2-3A, with a 14 kbp insert showed strong homology to the pfim3-1 cDNA. The region of homology, a 700 bp Xba I fragment, was subcloned into pUG19. This plasmid was refered to as pXX8. DNA sequence determinations of pXX8 and adjacent fragments from A2-3A suggested that the cloned DNA was a portion of the rONA repeat encoding the small subunit rRNA. DNA sequence analysis of pfim3-1 yielded an incomplete open reading frame. The predicted amino acid sequence codes for a 206 amino acid, 22 kDa polypeptide which contains a domain similar to a transmembrane domain from rat leukocyte antigen, GDS3. As well, an untranslated 576 nucleotide domain showed 81 % homology to pXX8 and 830/0 homology to the 188 rRNA sequence of Ustilago maydis. This sequence was found adjacent to a region of adenine-thymine base pairs presumed to represent the polyadenylation sequence of the fimbrial transcript. The size and extent of homology is sufficient to account for the hybridization of pfim3-1 to rDNA. It is suggested that this domain represents a completely novel regulatory domain within eukaryotes that may enable the observed rapid regeneration of fimbriae in U. violacea.

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This study examines the connection between leisure group participation and learning activities undertaken by participants in the Society for Creative Anachronism (SCA), a medieval recreationist group. The thesis of this connection was developed through the researcher's observations during SCA participation. The intent of this study is to understand adult learning from the self-directed learning, lifelong learning, and -transformative learning components derived from participant's SCA experiences. This qualitative study was conducted by interviewing eight active SCA participants, two in each participation theme of historical research, artistic representation, performance, and martial skills. Informants' responses demonstrated an integration of their leisure activity with learning. The contextualization of learning a s both a primary activity and a necessary support to participation, places learning a t the heart of participants' SCA related activities. The positive descriptions of learning activities, descriptive terms of ownership, and situating learning as an enjoyable activity engaged for the pleasure of the experience, provides adult educators with a fascinating glimpse of willing and engaged adult learners pursuing lifelong learning outside of the traditional educational structure. Two themes emerged during the interviews. First, bonding with others provided the motivation to continue their activities. Secondly, a feeling of commitment and helonging defined their enjoyment and satisfaction with SCA participation. The clear implications are that adult educators can create effective learning communities by developing educational structures that engage adult learners wi th meaningful social interaction.

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In animals, both stress resistance and longevity appear to be influenced by the insulin/insulin-like growth factor-l signaling (lIS) pathway, the basic organization of which is highly conserved from invertebrates to vertebrates. Reduced lIS or genetic disruption of the lIS pathway leads to the activation of forkhead box transcription factors, which is thought to upregulate the expression of genes involved in enhancing stress resistance, including perhaps key antioxidant enzymes as well as DNA repair enzymes. Enhanced antioxidant and DNA repair capacities may underlie the enhanced cellular stress resistance observed in long-lived animals, however little data is available that directly supports this idea. I used three. experimental approaches to test the association of intracellular antioxidant and DNA base excision repair (BER) capacities with stress resistance and longevity: (1) a comparison of multiple vertebrate endotherm species of varying body masses and longevities; (2) a comparison of long-lived Snell dwarf mice and their normallittermates; and (3) a comparison of hypometabolic animals undergoing hibernation or estivation with their active counterparts. The activities of the five major intracellular antioxidant enzymes as well as the two rate-limiting enzymes in the BER pathway, apurininc/apyrimidinic (AP) endonuclease and polymerase ~, were measured. These measurements were performed in one or more of the following: (1) cultured dermal fibroblasts; (2) brain tissue; (3) heart tissue; (4) liver tissue. My results indicate that antioxidant enzymes are not universally upregulated in association with enhanced stress resistance and longevity. I also did not find that BER enzyme activity was positively correlated with longevity, in an inter-species context, though there was evidence for enhanced BER in long-lived Snell dwarf mice. Thus, while there were instances in which enhanced antioxidant and BER enzyme activities were associated with increased stress resistance and/or longevity, this was not universally the case, indicating that other mechanisms must be involved. These results suggest the need to re-examine existing 'oxidative stress' hypotheses of longevity and probe further into the molecular physiology of longevity to discover its mechanistic basis.

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Photosynthesis in general is a key biological process on Earth and Photo system II (PSII) is an important component of this process. PSII is the only enzyme capable of oxidizing water and is largely responsible for the primordial build-up and present maintenance of the oxygen in the atmosphere. This thesis endeavoured to understand the link between structure and function in PSII with special focus on primary photochemistry, repair/photodamage and spectral characteristics. The deletion of the PsbU subunit ofPSII in cyanobacteria caused a decoupling of the Phycobilisomes (PBS) from PSII, likely as a result of increased rates of PSII photodamage with the PBS decoupling acting as a measure to protect PSII from further damage. Isolated fractions of spinach thylakoid membranes were utilized to characterize the heterogeneity present in the various compartments of the thylakoid membrane. It was found that the pooled PSIILHCII pigment populations were connected in the grana stack and there was also a progressive decrease in the reaction rates of primary photochemistry and antennae size of PSII as the sample origin moved from grana to stroma. The results were consistent with PSII complexes becoming damaged in the grana and being sent to the stroma for repair. The dramatic quenching of variable fluorescence and overall fluorescent yield of PSII in desiccated lichens was also studied in order to investigate the mechanism by which the quenching operated. It was determined that the source of the quenching was a novel long wavelength emitting external quencher. Point mutations to amino acids acting as ligands to chromophores of interest in PSII were utilized in cyanobacteria to determine the role of specific chromophores in energy transfer and primary photochemistry. These results indicated that the Hl14 ligated chlorophyll acts as the 'trap' chlorophyll in CP47 at low temperature and that the Q130E mutation imparts considerable changes to PSII electron transfer kinetics, essentially protecting the complex via increased non-radiative charge Photosynthesis in general is a key biological process on Earth and Photo system II (PSII) is an important component of this process. PSII is the only enzyme capable of oxidizing water and is largely responsible for the primordial build-up and present maintenance of the oxygen in the atmosphere. This thesis endeavoured to understand the link between structure and function in PSII with special focus on primary photochemistry, repair/photodamage and spectral characteristics. The deletion of the PsbU subunit ofPSII in cyanobacteria caused a decoupling of the Phycobilisomes (PBS) from PSII, likely as a result of increased rates of PSII photodamage with the PBS decoupling acting as a measure to protect PSII from further damage. Isolated fractions of spinach thylakoid membranes were utilized to characterize the heterogeneity present in the various compartments of the thylakoid membrane. It was found that the pooled PSIILHCII pigment populations were connected in the grana stack and there was also a progressive decrease in the reaction rates of primary photochemistry and antennae size of PSII as the sample origin moved from grana to stroma. The results were consistent with PSII complexes becoming damaged in the grana and being sent to the stroma for repair. The dramatic quenching of variable fluorescence and overall fluorescent yield of PSII in desiccated lichens was also studied in order to investigate the mechanism by which the quenching operated. It was determined that the source of the quenching was a novel long wavelength emitting external quencher. Point mutations to amino acids acting as ligands to chromophores of interest in PSII were utilized in cyanobacteria to determine the role of specific chromophores in energy transfer and primary photochemistry. These results indicated that the Hl14 ligated chlorophyll acts as the 'trap' chlorophyll in CP47 at low temperature and that the Q130E mutation imparts considerable changes to PSII electron transfer kinetics, essentially protecting the complex via increased non-radiative charge.

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Development of guanidine catalysts is explored through direct iminium chloride and amine coupling, alongside a 2-chloro-l,3-dimethyl-IH-imidazol-:-3-ium chloride (DMC) induced thiourea cyclization. Synthesized achiral catalyst N-(5Hdibenzo[ d,t][1,3]diazepin-6(7H)-ylidene)-3,5-bis(trifluoromethyl) aniline proved unsuccessful towards O-acyl migrations, however successfully catalyzed the vinylogous aldol reaction between dicbloro furanone and benzaldehyde. Incorporating chirality into the guanidine catalyst utilizing a (R)-phenylalaninol auxiliary, generating (R)-2-((5Hdibenzo[ d,t] [1,3 ]diazepin-6(7H)-ylidene ) amino )-3 -phenylpropan-l-ol, demonstrated enantioselectivity for a variety of adducts. Highest enantiomeric excess (ee) was afforded between dibromofuranone and p-chlorobenzaldehyde, affording the syn conformation in 96% ee and the anti in 54% ee, with an overall yield of30%. Attempts to increase asymmetric induction were focused on incorporation of axial chirality to the (R)phenylalaninol catalyst using binaphthyl diamine. Incorporation of (S)-binaphthyl exhibited destructive selectivity, whereas incorporation of (R)-binaphthyl demonstrated no effects on enantioselectivity. Current studies are being directed towards identifying the catalytic properties of asymmetric induction with further studies are being aimed towards increasing enantioselectivity by increasing backbone steric bulk.

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Scientists have been debating for decades the origin of life on earth. A number of hypotheses were proposed as to what emerged first RNA or DNA; with most scientists are in favour of the "RNA World" hypothesis. Assuming RNA emerged first, it fellow that the RNA polymerases would've appeared before DNA polymerases. Using recombinant DNA technology and bioinformatics we undertook this study to explore the relationship between RNA polymerases, reverse transcriptase and DNA polymerases. The working hypothesis is that DNA polymerases evolved from reverse transcriptase and the latter evolved from RNA polymerases. If this hypothesis is correct then one would expect to find various ancient DNA polymerases with varying level of reverse transcriptase activity. In the first phase of this research project multiple sequence alignments were made on the protein sequence of 32 prokaryotic DNA-directed DNA polymerases originating from 11 prokaryotic families against 3 viral reverse transcriptase. The data from such alignments was not very conclusive. DNA polymerases with higher level of reverse transcriptase activity were non-confined to ancient organisms, as one would've expected. The second phase of this project was focused on conditions that may alter the DNA polymerase activity. Various reaction conditions, such as temperature, using various ions (Ni2+, Mn2+, Mg2+) were tested. Interestingly, it was found that the DNA polymerase from the Thermos aquatics family can be made to copy RNA into DNA (i.e. reverse transcriptase activity). Thus it was shown that under appropriate conditions (ions and reactions temperatures) reverse transcriptase activity can be induced in DNA polymerase. In the third phase of this study recombinant DNA technology was used to generate a chimeric DNA polymerase; in attempts to identify the region(s) of the polymerase responsible for RNA-directed DNA polymerase activity. The two DNA polymerases employed were the Thermus aquatic us and Thermus thermophiles. As in the second phase various reaction conditions were investigated. Data indicated that the newly engineered chimeric DNA polymerase can be induced to copy RNA into DNA. Thus the intrinsic reverse transcriptase activity found in ancient DNA polymerases was localized into a domain and can be induced via appropriate reaction conditions.