Browsing by Subject "transformation"
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Item Acculturation and Transformation among Female Immigrant Military Spouses in an ESL Learning Program at a Community College(2012-07-16) Darnell, PatriciaThis study was designed to explore the acculturation experiences of military-related immigrant wives enrolled in an ESL program in a selected community college. More specifically, the purpose of the study was to understand the personal and structural forces that facilitated or hindered their acculturation process into their community of residence and whether their participation and retention in ESL classes contributed to their acculturation. Using a qualitative design with the basic interpretive paradigm, data collection consisted of face-to-face interviews with 14 immigrant military wives from 10 differing countries who were either enrolled or had been enrolled in a community college ESL program. The site chosen served a multicultural population of military spouses who enrolled in educational programs that offered English language development. The nearest ESL program, located at a community college near the military base, became an information-rich site for the study. The findings from the study highlighted the role of English language as an essential element to adjustment into the society of the United States for military immigrant wives, leading to acculturation and subsequently personal transformation. The data revealed both external and internal forces that influenced the acculturation process. External (structural) forces included community, workforce, the military, and an educational institution. The secondary forces included racial discrimination, cultural differences, and social networks within the community. Internal forces included love and care and self-efficacy. Self-efficacy was manifested through their persistence, patience, and resilience.Item Biolistic and agrobacterium-mediated genetic transformation of immature and mature embryos of spring wheat cultivar Saratovskaya-29(Texas A&M University, 2005-08-29) Kopbayev, Arman A.Plant transformation provides a promising methodology of introducing new genes that encode desirable traits to a wide range of crop plants. Success in genetic transformation has been achieved in many of the important crop species, such as soybean, cotton, rice, corn. However, wheat, one of the major crops of the world, has been considered to be difficult to transform via either Agrobacterium or biolistic bombardment (Rakszegi et al., 2001). There have been limited studies on A. tumefaciens-mediated transformation of cereals, including wheat, because of the overall refractory character of host-pathogen interactions between Agrobacterium and the cereal plants (Gould et al., 1991; Hiei et al., 1994; Cheng et al., 1997). While the genetic transformation of rice using Agrobacterium has become routine, only a few successful studies of Agrobacterium- mediated transformation of wheat have been reported, and these involved a model spring wheat, Triticum aestivum cultivar Bobwhite (Cheng et al., 1997). Model genotypes are developed for ease of plant regeneration in tissue culture and both Agrobacterium and biolistic mediated transformation methods require regeneration of plants in tissue culture. More success has been achieved in obtaining fertile transgenic wheat plants by particle bombardment, or biolistics method (Vasil et al., 1992; Weeks et al., 1993; Becker et al., 1994; Zhou et al., 1995; Altpeter et al., 1996). Wheat plants of the model system cultivar Bobwhite were used in most of these studies as well. The primary objective of this study was to use the callus-based transformation procedures mentioned above with a non-model cultivar of hexaploid spring wheat Saratovskaya-29, widely grown in Kazakhstan, to test the genotype dependence of the previously developed transformation protocols with respect to stable transfer of DNA and regeneration of transgenic plants. The spring wheat cultivar Saratovskaya-29 (Albidum-24/ Lutescens-55-11) was chosen for the study as being one of the most widely grown wheat cultivars both in Russia and Kazakhstan. It was bred in early 50??s in the Research Institute of the South-East, Saratov. Because of its drought resistance and good baking quality traits, Saratovskaya-29 reached a peak of nearly 21.2 mln ha in the former USSR in 1996 (Martynov and Dobrotvorskaya, 1996). Economical importance of this cultivar makes it an appropriate candidate for further improvement of economically significant traits. Another objective of the study described was to compare the transformation efficiencies and inheritance in the transgenic plants produced.Item Characterization of the Meq oncoproteins of Marek's disease virus vaccine strain CVI988/Rispens(2010-07-14) Ajithdoss, Dharani K.Marek?s disease virus serotype-1 (MDV-1) causes T cell lymphomas in chickens. Vaccines prepared from attenuated CVI988/Rispens MDV-1 strain currently offer the best protection. Although attenuated CVI988 is non-oncogenic, it codes for two forms of the MDV-1 oncoprotein Meq (CVI-Meq and CVI-L Meq). In this study, both CVI-Meq proteins, like the Meq protein of Md5 (a very virulent oncogenic strain), transformed Rat-2 and NIH3T3 cells. Both CVI-Meq and CVI-L Meq proteins activated the meq promoter only in the presence of chicken c-Jun (CK-Jun) whereas Md5-Meq activated the same promoter irrespective of CK-Jun co-expression. However, all three Meq proteins bound the meq promoter regardless of whether CK-Jun was co-expressed. We constructed three chimeric Meq proteins, namely, Md5-CVI-Meq, CVI-Md5-Meq, and Md5-CVI-L by exchanging domains between Md5 meq and CVI meq genes. Although these chimeric Meq proteins transactivated the meq promoter, the activation was significantly less than Md5-Meq. The current study indicated amino acid residues at positions 71 and 320 were important for Md5-Meq increase transcription of its own promoter. All three Meq proteins activated the MDV gB, MMP-3 and Bcl-2 promoters and suppressed transcription from the MDV pp38/pp14 bidirectional promoter. CVI-Meq protein in the context of other Md5 genes caused tumors only in 6% of chickens when compared to parental rMd5 (a very virulent strain), which induced lymphomas in 100% of chickens, (Reddy and Lupiani, unpublished data). Taking advantage of these two different phenotypes, we constructed two chimeric Meq proteins, Md5/CVI-Meq and CVI/Md5-Meq, by exchanging DNA binding and transactivation domains between Md5-Meq and CVI-Meq to understand the role of the DNA binding and the transactivation domains of Meq in transformation. rMd5-Md5/CVI-Meq virus caused 100% mortality in chickens and T lymphomas were found at high frequency in the peripheral nerves and various organs such as the heart, spleen, kidney, and gonads. On the other hand, rMd5-CVI/Md5-Meq induced disease in 36% of chickens on average and lesions were primarily in the nerves. Very rarely, lesions were present in the spleen and heart and no tumors were present in the kidney or gonads. Our results suggest that both the DNA binding domain and transactivation domain of Meq could cooperatively determine the nature of lymphomas in chickens.Item Genetic Engineering of Beta-Carotene Production in Honeydew Melons (Cucumis melo L. inodorus)(2012-02-14) Ren, YanGenetic transformation is a useful tool to incorporate novel genes, potentially allowing sexual incompatibility and interspecific barriers to be circumvented. The purpose of this study was to improve beta-carotene levels in melon fruits by transferring a phytoene synthase (PSY) gene. At present, there are not sufficient regeneration and transformation studies reported on two commercially important melon types - western shipper cantaloupe and honeydew. To establish a high efficiency shoot regeneration system, we evaluated three types of explants in our elite breeding lines. A shoot tip with a hypocotyl and cotyledon fragments, regenerated shoots whereas a shoot tip with a hypocotyl without cotyledon, did not produce regenerants. Murashige & Skoog (MS) basal medium with 1 mg 1?? benzyladenine (BA), 0.26 mg 1?? abscisic acid (ABA) and 0.8 mg 1?? indole-3-acetic acid (IAA) was the best for regeneration from cotyledon explants in cantaloupe 'F39'. MS basal medium with 1 mg 1?? BA and 0.26 mg 1?? ABA was chosen for honeydew '150' to solve a curving-up problem of explants. Fifty to sixty percent of regenerants were found to be polyploids. To establish a reliable Agrobacterium-mediated transformation protocol, kanamycin sensitivity as well as Timentin[trademark] and Clavamox? were evaluated. Kanamycin 200 and 150 mg 1?? were chosen as the threshold levels for 'F39' and '150' respectively. No significant differences were found between Timentin[trademark] and Clavamox? in 'F39'; however, Clavamox? reduced the incidence of vitrification and increased the frequency of shoot elongation in '150'. A. tumefaciens strain EHA105, harboring pCNL56 carrying nptII and gusA genes, was used to establish a transformation protocol. The transformation efficiency was 0.3% from 'F39' and 0.5% from '150'. We introduced a watermelon PSY-C gene under the control of a fruit-specific promoter of a polygalacturonase gene into '150'. All the transgenic plants were tetraploids based on flow cytometry assays. Up to 32-fold of beta-carotene was elevated in the rind tissue of transgenic honeydew including phytoene increase. This is a very promising result for a further investigation to increase beta-carotene level in flesh tissue using the PSY-C gene with an appropriate promoter.Item Overexpression of Tobacco Osmotin Protein in Carrot (Daucus carota L.) to Enhance Drought Tolerance(2012-10-31) Annon, Ali Hani HamzaLack of water is one of the most significant issues that already threaten world agriculture as many countries are unable to meet the demand for water to grow the crops. To make matters worse, the water availability is expected to fall by half by 2050, thus severely restricting agriculture production. Genetic engineering of crops to enhance their tolerance to such unfavorable environment represents one of the few approaches that can help us address this problem. Osmotin and osmotin-like proteins are stress proteins, belonging to the plant PR-5 group of proteins, which induced in response to various types of biotic and abiotic stresses in several plant species. Carrot plants were transformed with tobacco osmotin gene that encodes a protein lacking 20 amino-acid sequence at the C terminal end under the control of CaMV 35S promoter using the Agrobacterium-mediated transformation method. The gene integration and expression were confirmed by Southern and Western blot analyses and the transgenic plants were evaluated for their ability to tolerate drought stress. Under drought conditions, transformants exhibited slower rates of wilting compared to the wild-type and gained the ability to recover faster than their untransformed counterparts when the drought stress was alleviated. Under water stress, transformants showed lower levels of H2O2 accumulation, reduced lipid peroxidation and electrolyte leakage, and higher leaf water content. Taken together with some earlier reports, our results provide additional evidence for the protective ability of tobacco osmotin protein against drought stress and suggest a possible means to achieve tolerance against a serious type of abiotic stress.Item Role of the Leucine Zipper of Marek's Disease Virus Oncoprotein Meq in Pathogenesis(2010-07-14) Suchodolski, Paulette F.Marek's disease virus (MDV), the etiologic agent of Marek's disease, is a potent oncogenic herpesvirus. MDV is highly contagious and elicits a rapid onset of malignant T-cell lymphomas in chickens within several weeks after infection. The MDV genome encodes an oncoprotein, Meq, which shares resemblance with the Jun/Fos family of bZIP transcription factors. Similar to c-Jun, the leucine zipper region of Meq allows the formation of homo- and heterodimers. Meq homo- and heterodimers have different DNA binding affinities and transcriptional activity; therefore, they may differentially regulate transcription of viral and cellular genes. Previous in vitro data has suggested that Meq homodimers may be involved in regulating viral latency/reactivation, while Meq/c-Jun heterodimers are involved in transformation. Therefore, this research investigates the role of Meq homodimers and Meq-Jun heterodimers in the pathogenicity of MDV, by generating chimeric meq genes, which contain the leucine zipper region of the yeast transcription factor GCN4 (meqGCN) or leucine zipper region of c-Fos (meqFos). Thus producing Meq proteins that exclusively homodimerize (MeqGCN) or heterodimerize with Jun family members (MeqFos). Recombinant viruses (rMd5-MeqGCN and rMd5- MeqFos) containing the chimeric genes meqGCN or meqFos, respectively, in place of parental meq were generated with overlapping cosmid clones of Md5, a very virulent MDV strain. The chimeric genes were evaluated in vitro and retained DNA binding and transactivation/repressive functions, however, selected cells expressing MeqGCN and MeqFos had reduced colony formation in soft agar. Both the rMd5-MeqGCN and rMd5- MeqFos viruses replicated in vitro and in vivo, but rMd5-MeqGCN was unable to transform T-cells in infected chickens, while rMd5-MeqFos induced preneoplastic nerve lesions in 50% of infected birds. However, a third virus rMd5-MeqFos/GCN, which contains one copy of each meqGCN and meqFos, induced preneoplastic nerve lesions in 60% of infected chickens and neoplastic lesions in 20% of infected chickens. These data provide the first in vivo evidence that both Meq homodimers and heterodimers are necessary for MDV induced transformation.