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Existing methods to systematically characterize sequence-intrinsic activity of promoters are limited by relatively low throughput and the length of sequences that could be tested. Here we present Survey of Regulatory Elements (SuRE), a method to assay more than a billion DNA fragments in parallel for their ability to drive transcription autonomously. In SuRE, a plasmid library is constructed of random genomic fragments upstream of a barcode and decoded by paired-end sequencing. This library is transfected into cells and transcribed barcodes are quantified in the RNA by high-throughput sequencing. By computationally analyzing the resulting data using generalized linear models, we succeed in delineating subregions within promoters that are relevant for their activity on a genomic scale, and making accurate predictions of expression levels that can be used to inform minimal promoter reporter construct design. We also show how our approach can be extended to analyze the differential impact of single-nucleotide polymorphisms (SNPs) on gene expression.

Thesis (M. Phil.)--University of Hong Kong, 2007. / Also available online.

Thesis (M. Phil.)--University of Hong Kong, 2007. / Title proper from title frame. Also available in printed format.

Thesis (M. Phil.)--University of Hong Kong, 2005. / Title proper from title frame. Also available in printed format.

Sequence-specific PCR markers were derived from one new and eight previously identified random amplified polymorphic DNA (RAPD) markers for the purpose of identifying grape (Vitis) rootstocks. The markers were developed because the RAPD assay was found to be inconsistent and the original RAPD markers unreliable. Southern hybridization analysis of the RAPD gels with cloned RAPD bands as probes revealed deficiencies of scoring RAPD bands based solely on ethidium bromide staining. In one case, bands of the same size generated by the same primer in different rootstocks -- normally scored as the same marker -- failed to cross-hybridize, implying a lack of homology between the bands. In another case, a band scored as absent based on ethidium bromide staining was detected by hybridization. One of nine RAPD bands was cloned in the present study. All nine were partially sequenced and sequence-specific pairs of primers were synthesized for each for use under stringent PCR conditions. Three of the primer pairs generated products only from the rootstocks from which the original RAPD bands had been cloned; three others produced products from additional rootstocks while still generating polymorphisms; two others generated apparent length variants from some accessions; and one primer pair resulted in a loss of polymorphism. Based on the eight polymorphic markers, five of nine rootstocks could be unambiguously distinguished. / Graduation date: 1996

DNA repair mechanisms counteract the formation of deleterious DNA lesions and maintain genomic integrity. Nucleotide excision repair (NER) is an important DNA repair pathway because of its extraordinarily large substrate specificity. P53 protein regulates NER pathway in a transcription-dependent or transcription-independent manner. Inherited polymorphisms of NER pathway genes (XPC, HR23B, XPA, DDB2, XPB, XPD, ERCC1, XPF, and XPG) and TP53 gene may contribute to individual variations in genetic susceptibility to OSCC and correlate with the prognosis of 204 OSCC patients. We carried out a hospital-based case-control study to investigate the association of 25 various polymorphisms of nine NER pathway genes and TP53 gene with the risk for OSCC. There were 34 newly diagnosed OSCC patients and 135 frequency-matched controls without BQ chewing and smoking habit as well as 313 newly diagnosed OSCC patients with BQ chewing or smoking habit and 312 frequency-matched controls being recruited between November 2003 and July 2007 at Kaoshiung Veterans General Hospital. Genotyping was performed using the PCR-RFLP techniques or TaqMan real-time PCR method. The significant association between polymorphisms of NER pathway genes and OSCC risk was mainly found among subjects with BQ chewing or smoking habit. In the single locus analysis, GA and AA genotypes of ERCC1 G-641A (AOR, 0.64; 95% CI, 0.45-0.93 and AOR, 0.48; 95% CI, 0.29-0.79, respectively; p for trend, 0.002), CT genotype of XPF C-850T (AOR, 1.53; 95% CI, 1.08-2.18; p for trend, 0.014), as well as GG genotype of XPB A-1039G (AOR, 0.51; 95% CI, 0.26-0.98; p for trend 0.034) were significantly associated with the risk of OSCC. Furthermore, -641G/ -425T or -641G/ -425C haplotype of ERCC1 (AORs, 1.34; 95% CI, 1.02-1.77 and AOR, 1.56; 95% CI, 1.18-2.07, respectively; p for trend 0.002) as well as -850T/ -247T and -850T/ -247C haplotype of XPF (AOR, 1.45; 95% CI, 1.09-1.94 for; p for trend 0.016) were strongly associated with the risk of OSCC. A trend toward increased risk of OSCC was observed when people with the increasing number of at risk genotypes in the combined analyses of nine NER pathway genes with (p for trend, <0.001) or without (p for trend 0.001) TP53 gene. Finally, in the stratification analysis, the combined effects of nine NER pathway genes had a significantly increased risk of OSCC among younger group (¡Ø50 years old), Fukienece population, BQ chewers, light smokers, or light drinkers. Besides, in the prognosis analysis of 204 OSCC patients, HR23B A-823C, polymorphisms of XPA gene, XPD C-643G, XPG C787G, and the number of at risk genotypes of NER pathway genes were associated with pathologic stage, T classification, or N classification. The association between NER genetic polymorphisms and survival of patients was only found in XPA C-1778T polymorphism. These results suggested that the single polymorphism of XPB A-1039G, ERCC1 G-641A and XPF C-850T, the joint effect of genetic polymorphisms of NER pathway genes, and gene-environment combined effect were associated with the risk of OSCC. Furthermore, in the analysis of NER genetic polymorphisms and prognosis of OSCC, we found polymorphisms of XPA gene might be a prognostic factor for OSCC.

Thesis (Ph. D.)--York University, 2000. Graduate Programme in Biology. / Typescript. Includes bibliographical references. Also available on the Internet. MODE OF ACCESS via web browser by entering the following URL: http://wwwlib.umi.com/cr/yorku/fullcit?pNQ56253.

published_or_final_version / Paediatrics and Adolescent Medicine / Master / Master of Research in Medicine

published_or_final_version / Medical Sciences / Master / Master of Medical Sciences