Development of techniques for testing the effect of the E1 region on adenovirus integration /

Adenoviral vectors are currently the most widely used gene therapeutic vectors, but their inability to integrate into host chromosomal DNA shortened their transgene expression and limited their use in clinical trials. In this project, we initially planned to develop a technique to test the effect of the early region 1 (E1) on adenovirus integration by comparing the integration efficiencies between an E1-deleted adenoviral vector (SubE1) and an Elcontaining vector (SubE3). However, we did not harvest any SubE3 virus, even if we repeated the transfection and successfully rescued the SubE1 virus (2/4 transfections generated viruses) and positive control virus (6/6). The failure of rescuing SubE3 could be caused by the instability of the genomic plasmid pFG173, as it had frequent intemal deletions when we were purifying It. Therefore, we developed techniques to test the effect of E1 on homologous recombination (HR) since literature suggested that adenovirus integration is initiated by HR. We attempted to silence the E1 in 293 cells by transfecting E1A/B-specific small interfering RNA (siRNA). However, no silenced phenotype was observed, even if we varied the concentrations of E1A/B siRNA (from 30 nM to 270 nM) and checked the silencing effects at different time points (48, 72, 96 h). One possible explanation would be that the E1A/B siRNA sequences are not potent enough to Induce the silenced phenotype. For evaluating HR efficiencies, an HR assay system based on bacterial transfonmatJon was designed. We constmcted two plasmids ( designated as pUC19-dl1 and pUC19-dl2) containing different defective lacZa cassettes (forming white colonies after transformation) that can generate a functional lacZa cassette (forming blue colonies) through HR after transfecting into 293 cells. The HR efficiencies would be expressed as the percentages of the blue colonies among all the colonies. Unfortunately, after transfonnation of plasmid isolated from 293 cells, no colony was found, even at a transformation efficiency of 1.8x10^ colonies/pg pUC19, suggesting the sensitivity of this system was low. To enhance the sensitivity, PCR was used. We designed a set of primers that can only amplify the recombinant plasmid fomied through HR. Therefore, the HR efficiencies among different treatments can be evaluated by the amplification results, and this system could be used to test the effect of E1 region on adenovirus integration. In addition, to our knowledge there was no previous studies using PCR/ Realtime PCR to evaluate HR efficiency, so this system also provides a PCR-based method to carry out the HR assays.

Vegetational history and geochronology of several sites in South Southwestern Ontario with discussion on mastodon extinction in Southern Ontario /

This study has three purposes: to establish a chronologically controlled vegetational history for a number of sites in south Southwestern Ontario; to utilize the resulting data to support and/or add to the current understanding of Quaternary geology and stratigraphy, and the glacial and postglacial history of the Great Lakes in south Southwestern Ontario; and to attempt to propose a possible explanation for the extinction of the mastodon in Southern Ontario. Palynological and geochronological analyses were conducted on material collected from eleven sites (east to west): Verbeke Mastodon Site, Woloshko Mastodon Site, Walker Pond II, Pond Mills I, Lake Hunger Bog, Bouckaert Site. Mabee Site, Cornell Bog. Colles Lake I, Folden Mastodon Site and Forest Pond. Individual geochronologically controlled (where possible) vegetational histories were reconstructed for each of the sites investigated. The results of the individual studies, when considered in overview. indicated the existance of an established closed boreal forest throughout south Southwestern Ontario by 10,000 years B.P. This evidence for a significant climatic change coincident throughout south Southwestern Ontario supports the proposed age of 10,000 years B.P. for the Pleistocene/Holocene Boundary (Terasmae, 1972). Remnant patches of 'open spruce parkland' persisted in small local 'wet' areas. It was in these areas that the mastodon was restricted during early Holocene time. With continued encroachment by the surrounding boreal forest, possibly speeded up by this browser's destructive feeding habits, the spruce enclaves shrank and the mastodon became extinct in south Southwestern Ontario. The results of this thesis basically support Dreimanis' (1967, 1968) proposed 'Environmental-Climatic' theory for mastodon extinction. It is suggested that increased dryness during the present interglacial compared to the climate of earlier interglacials may be the key to unravelling the problem of mastodon extinction in eastern North America.

Burk's guide of Niagara Falls : eighty illustrations of valued interest to any visitor, guide map on pages 32 and 33, directions as suggested by a resident ... /

Cover title.

Familiar Science of The Scientific Explanation of Common Things

There is a signature on this book that reads "Julia Cleveland 22 East-8 Street, Erie Penna."

Drawing and Explanation of the Cleveland Family Coat of Arms

Drawing and explanation of the Cleveland Family coat of arms. Pro Deo et Patria means For God and Country. The name is Saxon in origin and in 1403 the “de” was dropped from the name. The drawing and text are said to be from Cleveland Genealogy by J.B. Cleveland, 1881. It can also be found in An Account of the Lineage of General Moses Cleaveland (founder of the city of Cleveland, Ohio) of Canterbury, compiled by H.G. Cleveland, n.d.

Explanation of the Land Surrendered to the Crown by the Six Nations Indians for Mr. Dickson

Explanation of the Land Surrendered to the Crown by the Six Nations Indians for Mr. Dickson: 1 handwritten page explaining the Six Nations surrender of land to the Crown in order to allow the Crown to transfer the land to Mr. Dickson to compensate him for legal fees. [This is not an older document. It was possibly written by the Niagara Historical Society], n.d.