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Department: Microbiology More Information A Day in the Life Working in the Hay LabMy work in the laboratory of Dr. Anthony Hay focuses on the interactions of defective phages, and how it appears that the bacterium Escherichia coli has domesticated some the phage genome for its own benefit in switching from the planktonic to biofilm lifestyle. In order to study how these phage genes may be used I genetically manipulate the genome of the bacterium or introduce exogenous DNA via plasmid vectors, and then I analyze the manifested effects using a myriad of quantitative and qualitative techniques.Genetic manipulation is the heart of molecular biology, and has rapidly evolved past the reliance upon endonuclease restriction and ligation used heavily over the past fifty years. The mainstay of our research focuses on “knocking out” or deleting specific genes we feel are of relevant interest. The procedure that we use was developed by Wanner and Detsenko known as the Lambda Red system, is the gold standard of knockout protocols across nearly all strains of E. coli and even some strains of Salmonella. This protocol requires me to have knowledge of the polymerase chain reaction developed by Karry Mullins, the ability to induce plasmid borne genes, as well as the ability to transform genetic material into the host via electroporation. In addition to this relatively new protocol I also clone many genes of interests, and sometimes their promoters, into plasmid vectors. Between the Lambda Red protocol and the more mainstay molecular cloning, during any given day I am very busy utilizing a wide variety of genetic techniques to silence or express the remnant phage genes of E. coli. Though the genetic manipulation of E. coli is interesting and rewarding when successful, the most enjoyable part of research in the Hay laboratory is getting to explore the consequences of your genetic tampering. One of the beauties of researching at Cornell is that there is never a lack of very expensive equipment to be played with, some of which I use to inspect the biofilm forming capabilities of bacteria, or some times to look at the cells themselves. To image biofilms I use Confocal Scanning Laser Microscopy (CSLM), in which the microscope will scan an object layer by layer and create a “stack” of these images as it moves up or down the object. Software can then take these image stacks and convert them into 3-D images. CSLM allows us to visualize the biofilm forming capabilities of both our wild type and mutant strains in very intimate detail. To visualize cell surface structures that may be affected by mutagenesis we employ Transmission Electron Microscopy (TEM). This type of microscopy relies on a high energy beam of electrons being directed at a sample stained with a heavy metal such as tungsten or molybdenum, which scatters some electrons and allows others through, in doing so generating an image. Though I find the different types of microscopy employed in my work to be of particular interest there is other equipment that I have become familiarized with during my two years working for Dr. Hay. Spectrophotometery, luminosity, and fluorescence are all measured by different pieces of equipment for assays that run. While being less flashy than some of the higher end optics, these machines are invaluable in assessing cellular fitness and genomic expression. The work I have done in Dr. Hay’s lab has greatly increased my understanding of many biological concepts, especially those concepts surrounding DNA replication and expression. More importantly work in the laboratory has given me a crash course in patience and perseverance, as there is a “re” in research for a reason. |