CaB Research Leaders Challenge Nathan Battersby, Gemma Mancuso, Murad Jabarov, Viktoria Weisz and Ramya Selladurai

How can we improve Canada’s Natural Resource sector? Athabasca Oil Sands, Alberta Tailing Ponds http://goo.gl/ORUzVo

Alcanivorax borkumensis Active Genes: ALK B1 ALK B2 Cytochrome P450 [1] Teimoori, A., et al. (2012). Biotechnology and Applied Biochemistry 59(6): 457-464. http://goo.gl/T7cMeS

Research Gene Knockout Techniques Replacement of each gene Isolate the enzyme encoding genes to test hydrocarbon-degrading capabilities

Canada meets high global demand for oil by extracting and refining Bitumen http://goo.gl/7P75xB

Tailing Ponds Water used in the hot water filtration process must be stored in tailing ponds due to its toxicity [1] Contents of water in tailing ponds: - sand - clay - residual bitumen [2] Tailing ponds are located along the Athabasca River in Alberta http://goo.gl/z96Cks [1] Frank et al. Environmental Science & Technology, 2009 43, 266-71. [2] Canadian Association of Petroleum Producers. Oil Sands Today, 2015.

Rainbow Trout Rainbow trout exposed to water influenced by tailings for short periods of time experience negative health effects [3]: -Fin erosion -Compromised immune systems -Decrease in size of spleens http://goo.gl/6WZoSu [3] McNeill et al. Ecotoxicology and Environmental Safety, 2012 84, 254-261

Hypothesis The genes that encode for enzymes that degrade hydrocarbons found in our bacteria A. borkumensis and their success rate in degrading hydrocarbons found in tailing ponds

Control! Alkanivorax borkumensis has 3 genes Alk B1, Alk B2, Cyt-P450 which will be tested individually, all together and in combinations http://goo.gl/NpstbR

Homologous Recombination = The Gene Switch

The Products Αlk Β1, Αlk Β2 & Δ Cyt-P450 Αlk Β1 Αlk Β2 Cyt - P450 the control

Biological Photocopier PCR Polymerase Chain Reaction http://goo.gl/7f7HEe http://goo.gl/n9pc8f

Amplification! Replicate, replicate and replicate! 30 Cycles = 1,073,741,842 Copies!! http://goo.gl/MS1rEX

Nucleic Acid Electrophoresis http://goo.gl/8GAaie http://goo.gl/YDbUov

DNA sequencing http://goo.gl/vEhhjq http://goo.gl/F8n4nd

Conclusion We aim to provide an ecologically friendly method to deal with the hydrocarbon waste products of the Athabasca Oil Sands operations The bacterium A. borkumensis will be able to perform the task of hydrocarbon degradation Laboratory techniques used: Gene knockout, homologous recombination, gene knockout, polymerase chain reaction, nucleic acid electrophoresis

Developing Biofilm composed of A. borkumensis Future Research: Developing Biofilm composed of A. borkumensis Determining exact byproducts of A. borkumensis metabolism Bioengineering: Replacement of Alk B1, Alk B2 and Cyc P450 with kanamycin resistance gene Our Overall Goal: Bioremediation of tailing ponds, in order to clean up the environment http://goo.gl/nIUofF [9] Reva et al. Global features of the Alcanivorax borkumensis SK2 genome, 2007 [10] Sabriova et al. Functional genome analysis of Alcanivorax borkumensis strain SK2: Alkane metabolism , environmental adaptations and biotechnological potential.

Acknowledgements Alex Somerville Rehman Ata Daniel Prete Crystal Gadishaw - Lue