American River Diversity

With the number of bacteria in our universe exceedingly numerous compared to the human population, it could seem quite difficult or maybe even impossible for scientists to analyze each and every single one of these microbes. So that is why we would be using a simple approach to looking at diversity in bacterial communities. In this experiment we went out to collect samples from the American River in order to analyze the various diversity in Bacteria species in the community. We focus our experiment by looking at a portion of the gene called the 16S rRNA which is found in all microbial species. Instead of using a shotgun metagenomics approach to DNA sequencing in which the total genomic DNA is sheared into tiny fragments for sequencing.
We will be using a method called Polymerase Chain Reaction (PCR) to amplify the 16S rRNA gene present in all prokaryotic cellular life and also culturing samples onto an agar plate for visual identification. This will allow for the accessibility of comparing the similarity or differences in the DNA sequence, which can reveal close/distant connection between various bacterial species in the American River community based on the amplified 16S rRNA gene. The 16S rRNA subunit gene contains hypervariable regions based on which further analysis will be conducted in Bio 184 to see the diversity of various bacterial species. Furthermore, we will conduct a gel electrophoresis by allowing the DNA which has a slightly negative charge from its phosphate backbone to travel to the positive end. This would allow us to see if our genomic DNA extraction and PCR were successful from the experiment conducted. Additionally, we would also quantify the cultured bacterial species present in our plates by calculating the total number of colony forming units (CFU) in each dilution. By examining the number and types of colonies present, this can lead us to indicate the diverse ecological conditions that might be responsible for the various distinctive results that might
Using the Powersoil DNA extraction kits, one gram of soil will be used with six individual reagents to purify the sample of any impurities that might interfere with our results. The six reagents used are labeled as C1, C2, C3, C4, C5, and C6, each with their individual properties to unpurified unwanted material in our soil sample. From here, perform a serial dilution from the soil and water sample. This will be used for plating bacteria on agar to see potential colonies that are present in the samples. Colonies that arises from this sample can help us estimate the culturable bacteria (CFU's) found in the American