Alibrio Ficheri Lab Report

A. fischeriThe overall purpose of this study was to create a genomic library of Aliivibrio fischeri (A. fischeri) thus aiding in creating a restriction map of the lux operon. It also employs typical molecular techniques important for biologists to understand. In this portion of the lab, the chromosomal DNA (chDNA) will be isolated. Its purity will be measured using spectrophotometric analysis. Lastly, the DNA will be digested and verified via gel electrophoresis.

A. fischeri is a gram negative rod shaped marine bacteria that exhibits bioluminescence and is both found free-living or in symbiosis. It has a variety of hosts such as the bobtail squid and is named after the German scientist Bernhard Fischer (Garrity, 2005). Bioluminescence in
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fischeri chDNA. Lysing the pellet of A. fischeri required the hydrogen bonds, hydrophobic interactions, and Van der Waals forces to be disrupted. This was done by the detergent sodium dodecyl sulfate (SDS), which was present in the re-suspension buffer added and created protein-lipid-detergent complexes. Proteinase K is added to digest proteins and any nucleases that might damage the DNA (Peant and LaPointe, 2004). RNase is added to avoid impurities from RNA. The addition of the chaotropic salt guanidinium chloride present in the lysis buffer helps the DNA adhere to the silica surface of the DNeasy Mini spin column by forming a cationic bridge that connects the DNA and silica gel (Yang et. al., 1998). It also denatures lipids and proteins, preventing them from adhering by weakening their hydrophobic interactions, causing them to be simply washed away. Ethanol is added to further ease DNA binding while continuing to eliminate the lipids and proteins by creating a hydrophobic environment. Wash buffer 1 washes away proteins while Wash buffer 2 is used to remove salts. DNA grade water is added in the end allow the DNA to renature and detach itself from the silica spin column (Qiagen,

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