The establishment of two novel bovine cell lines by transfection with the putative transforming region of bovine adenovirus type 3

Human adenoviruses (Ads), members of the family adenoviridae, are medium-sized DNA viruses which have been used as valuable research tools for the study of RNA processing, oncogenic transformation, and for the development of viral vectors for use in gene delivery and immunization technology. The left 12% of the linear Ad genollle codes for products which are necessary for the efficient replication of the virus, as well as being responsible for the forlllation of tumors in animallllodels. The establishlllent of the 293 cell line, by immortalization of human embryonic kidney cells with th~ E1 region of Ad type S (AdS), has facilitated extensive manipulation of the Ads and the development of recombinant Ad vectors. The study of bovine adenoviruses (BAVs), which cause mild respiratory and gastrointestinal infections in cattle has, on the other hand, been limited primarily to that of infectivity, immunology and clinicallllanifestations. As a result, any potential as gene delivery vehicles has not yet been realized. Continued research into the molecular biolo~gy of BAVs and the development of recolllbinant vectors would benefit from the development of a cell line analogous to that of the 293 cells. In an attelllpt to establish such a cell line, the recombinant plaslllid pKC-neo was constructed, containing the left 0-19.7% of the BAV type 3 (BAV3) genome, and the selectable marker for resistance to the aminoglycoside G418, a neomycin derivative. The plasmid construct was then used to transfect both the Madin-Darby bovine kidney (MDBK) -iicell line and primary bovine lung cells, after which G418-resistant foci were selected for analysis. Two cell lines, E61 (MDBK) and E24 (primary lung), were subsequently selected and analysed for DNA content, revealing the presence of the pKC-neo sequences in their respective genomes. In addition, BAV3 RNA transcripts were detected in the E61 cells. Although the presence of E1 products has yet to be confirmed in both cell lines, the E24 cells exhibit a phenotype characteristic of partial transformation by E1. The apparent immortalization of the primary lung cells will permit exploitation of their ability to take up exogenous DNA at high efficiency.

The wonders of Niagara : a visit to America's greatest cataract with a description of the points of interest in a region of scenic grandeur and beauty.

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Understanding the Cultural Health Beliefs in Diabetes Education Amongst the Aboriginal Population within a City in the Southern Region of Ontario

This study examined the cultural health beliefs in diabetes education amongst the Aboriginal population within a city in Southern Ontario. The purpose was to contribute to the development of a culturally relevant diabetes handbook as well as to delivery styles within current diabetes education programs. To this end, a focus group was conducted with Aboriginal men and women between the ages of 18-70 years with type 2 diabetes. Participants were recruited from 2 Aboriginal community centres and an Aboriginal health centre in a city in Southern Ontario. Themes were drawn from the analysis of the focus group transcripts and combined with the findings from the research literature. The major themes that merged were drawn from Eurocentric and Aboriginal theories. The results were a set of recommendations on the type of format for diabetes educational programs such as traditional group activities, variety of electronic format, and culture specific educational resources. The emergent results appear to provide some important insights into program planning for diabetes education centres within Aboriginal communities.

Petrography, Petrochemistry and petrogenesis of Huronian volcanic rocks of the Elliot Lake region, Ontario

In the Elliot lake region of northern Ontario, Yolcanlc lava piles represent the lowermost units of the Huronian SUpergroup. These rocks general1y trend east-west and belong to the Elliot lake Group. They are s1tuated on the north and south limbs or the QuIrke lake Syncline. The volcanIc rocks of this study contain a secondary minerai assemblage consisting of actinolite, biotite, chlorIte, eptdote/cllnozoislte tttanomagnettte and calcite characteristic of greenschist metamorphism. Compilation of data suggests that metamorphism of the volcanic rocks proceeded between 325- and 425-C and between 2.4 and 4.7 kb. Geochemtcally these lavas represent tholeiitic and calc-alkaline assemblages. The tholeiites are character1sttcally enriched tn Fe and Tt and consist mainly of basalts, basaltic andesites and andesites. These rocks are believed to have formed by the partIal melting of a peridottte source at low P-T. In contrast, the calc-alkaline rocks are depleted in Fe and TI, but show a signIficant enrichment In 51 and Zr; andesIte Is the major rock type for thIs assemblage. I·t Is postUlated that the calc-alkalIne sU1te of rocks was the result of eIther the partIal meltIng of abasaltic·magma at shallow depth, or the melttng of s1al1c crustal materIal due to the added we1ght of tholeiitIc material on an unstable crust and to downwarplng processes Inttlated by convection cells.

The cloning and reconstitution of a bovine adenovirus type 2 E3 deletion mutant and the sequencing and analysis of the early 4 region

Recombinant Adenoviruses (Ads) have been shown to have potential applications in three areas: gene therapy, high level protein expression and recombinant vaccines.' At least three different locations within the Ad genome can be deleted and subsequently used for the insertion of foreign sequences. These include the Early 3 (E3), Early 1 (E1) and Early 4 (E4) regions. Viral vectors of this type have been well studied in Human Ads 2 and 5, however one has not yet been constructed for Bovine Adenovirus Type 2 (BAV2). The E3 region is located between 76.6 and 86 m.u. on the r-strand and is transcribed in a rightward direction. The gene products of the Early 3 region (E3) have been shown to be non-essential for viral replication, in vitro, but are required for host immunosurveillance. This study represents the cloning and reconstitution of a BAV2 E3 deletion mutant. A deletion of 1800bp was made within the E3 region of BAV2 and the thymidine kinase gene was subsequently inserted in the deleted area . . The plasmid pdlE3-4tk1 (23.4Kbp) was constructed and used to to facilitate homologous recombination with the wild type BAV2 to produce a mutant. Southern Blotting and Hybridization results suggest the presence of a BAV2 E3 deletion mutant with thymidine kinase sequences present. The E4 region of Human Adenovirus types 2 and 5 is located at the extreme right end of the genome (91.3 map units - 99.1 map units) and is transcribed in a leftward direction giving rise to a complicated set of differentially spliced mRNAs. Essentially there are 7 open reading frames (ORFs) encoding for at least 7 polypeptides. The gene products encoded by the E4 region have been shown to be essential for the expression of late viral genes, host cell shutoff and normal viral growth. We have cloned and sequenced the right end segment between 90.5 map units and 100 map units of the BAV2 genome. The results show several open reading frames which encode polypeptides exhibiting homology to three polypeptides encoded by the E4 region of human adenovirus type 2. These include the 14kDa protein encoded by ORF1, the 34kDa protein encoded by ORF6 and the 13kDa protein encoded by ORF3. The nucleotide sequence, restriction enzyme map, and ORF map of the E4 region could be very useful in future molecular manipulation of this region and could possibly explain the slow growth rate of BAV2 in MDBK cells.