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Molecular Identification of DMS-Producing Bacteria Isolated from Marine Algae - Dissertation Example

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Dimethylsulfide is recognized as the predominant form of sulfur liberated from the oceans to the atmosphere, representing a fundamental compound to the global sulfur cycle. Several studies have indicated that DMS emission demonstrates a significant effect on the magnitude of the earth’s cloud cover, influencing the planet’s climate in the process…
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Molecular Identification of DMS-Producing Bacteria Isolated from Marine Algae
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"Molecular Identification of DMS-Producing Bacteria Isolated from Marine Algae"

This study proposes to determine the identity of DMS-producing bacteria isolated from phytoplankton cultures and grown on minimal medium with DMSP as a carbon source. Specifically, the study will seek to amplify the 16s rRNA gene through polymerase chain reaction (PCR), sequence the amplified gene, and compare the obtained sequence against existing sequences deposited in GenBank. Objectives The main objective of the study is to identify unknown DMS-producing bacteria using 16s rRNA gene sequence. The specific objectives are: 1. Extract genomic DNA from cultures of DMS-producing bacteria 2. Amplify the 16s rRNA gene of the obtained bacterial DNA using generic primers 3. Sequence the 16s rRNA gene of the bacteria 4. Compare the obtained 16s rRNA sequence with the sequences in the National Center for Biotechnology Information GenBank database to determine the taxonomic position and/or identity of the unknown organism. 5. Construct a phylogenetic tree of the unknown bacteria based on the 16s rRNA sequence Methods DNA extraction DMS-producing bacteria have already been isolated from selected phytoplankton cultures and will be re-isolated in minimal medium with DMSP. Then, selected clonal isolates will be grown in marine broth for 18-24 hours and subcultured into 50 mL of fresh medium. Then, the bacterial suspension will be incubated at 37 °C for 5 h in a shaker incubator (Yoch et al., 2001b). The cells will be harvested by centrifugation and genomic DNA will be extracted from the pellets using QIAamp® DNA Mini Kit (Qiagen Inc., Valencia, Calif.) Procedures for the DNA extraction will be adapted from the protocol provided by the manufacturer. Agarose Gel Electrophoresis Electrophoresis will be done after DNA...
Molecular Identification of DMS-Producing Bacteria Isolated from Marine Algae

The main objective of the study is to identify unknown DMS-producing bacteria using 16s rRNA gene sequence. The specific objectives are: extract genomic DNA from cultures of DMS-producing bacteria, amplify the 16s rRNA gene of the obtained bacterial DNA using generic primers, sequence the 16s rRNA gene of the bacteria, compare the obtained 16s rRNA sequence with the sequences in the National Center for Biotechnology Information GenBank database to determine the taxonomic position and/or identity of the unknown organism, construct a phylogenetic tree of the unknown bacteria based on the 16s rRNA sequence. DMS-producing bacteria have already been isolated from selected phytoplankton cultures and will be re-isolated in minimal medium with DMSP. Then, selected clonal isolates will be grown in marine broth for 18-24 hours and subcultured into 50 mL of fresh medium.Then, the bacterial suspension will be incubated at 37 °C for 5 h in a shaker incubator (Yoch et al., 2001b). The cells will be harvested by centrifugation and genomic DNA will be extracted from the pellets using QIAamp® DNA MINI KIT (Qiagen Inc., Valencia, Calif.) Procedures for the DNA extraction will be adapted from the protocol provided by the manufacturer.

Electrophoresis will be done after DNA extraction to verify the presence and quality of DNA. Agarose gel electrophoresis (AGE) will be done according to the procedure suggested by Voytas (2000). Read More
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