Week 2 Research Proposal

Research Proposal

1. Why research Staphylococcus Aureus DNA extraction?

In general, experimental research is important because it allows us to reinforce what we already know about organisms, but it can also open the door to new discoveries. For example, defining the best technique to extract DNA material from an organism can either confirm a hypothesis made on existing knowledge or it can lead us to the discovery of new information. In addition, determining the best method to extract DNA from a specific organism can be found valuable to an institution, as it can serve as a basis for the development of further investigations made on that subject. I will be conducting several experiments to extract DNA material from the Staphylococcus Aureus bacteria cells. Staphylococcus Aureus or S. aureus for short, is a bacterium that can be part of the normal skin flora of human beings and never cause a problem; it is estimated that 33% of Americans carry this microbe without illness (MRSA Tracking). However, if given the opportunity S. aureus will attack the body and cause illness; and what’s worse, is that many strains have developed resistance to widely used antibiotics such as Vancomycin and Methicillin making this seemingly harmless organism deadly under certain conditions (Gram-positive bacteria). Analyzing the DNA of this bacterium to understand how it developed this drug resistance is another important reason to figure out what is the best way extract its DNA as best as possible.

2. Research tactics.

There will be a total of 6 DNA extraction protocols (Determining an), one of which will be a commercial kit guaranteed to render high yields of quality DNA material (Ready-LyseTM lysozyme). Then I will analyze the extracted material using a microvolume UV-Vis spectrophotometer to quantify the results. The results that I will be observing are the concentration of DNA material and also the quality of the material extracted (260/280 and).

3. Variable table

Name	I/D/C	Symbol	Units	Description
Nucleic Acid Concentration	D		ng/uL	This figure will depend on the protocol used to extract DNA. The higher the value the more DNA material there was extracted.
Nucleic Acid Purity	D	260/230	unit-less ratio	This figure will help determine the ratio of pure DNA material there is through the extraction in contrast to organic contaminants such as proteins. The ideal 260/280 value is ~1.8, arbitrarily.
Method of Extraction	I	Protocol no.		There will six different methods used to extract the bacterium’s DNA and will identified as “Protocol…” followed by number 1, 2, 3, 4, or 5; for practical reasons.
Bacteria Sample	C			The bacteria used to determine the effectiveness of each extraction method will all come from the same isolated colonies (petri dish).
Equipment	C			Equipment used for centrifuging, heating, cooling, and measuring will all be the same throughout the different tests and trials thereafter.
Chemicals	C			Chemicals, such as alcohols and buffers will all come from the same source.

4. Overview of findings

Research question- 

     When comparing the quantity and quality of the DNA material extracted with the following methods: Boiling Protocol, TE Boiling Protocol, TE Freeze and Thaw Protocol, Lysis Protocol, and Meat Tenderizer Protocol, which DNA extraction protocol worked better? Which one extracted higher yields? Which one extracted better quality DNA?

Hypothesis-

      I think that Meat Tenderizer Protocol will extract better quality DNA in higher yields.

References

Developing an effective E-coli protocol. (n.d.). Raw data in preparation.

Gram-positive bacteria. (2011). In K. Rogers (Ed.), Bacteria and viruses. Retrieved from Gale Virtual Reference Library database. (Accession No. GALE|CX4040900042)

MRSA tracking. (2016, march 3). In Centers for disease control and prevention. Retrieved December 16, 2016, from https://www.cdc.gov/mrsa/tracking/

Ready-LyseTM lysozyme solution [Pamphlet]. (2012). Retrieved from http://www.epibio.com/docs/default-source/protocols/ready-lyse-lysozyme-solution.pdf?sfvrsn=6

260/280 and 260/230 ratios [Pamphlet]. (2008). Retrieved from http://www.nanodrop.com/Library/T009-NanoDrop%201000-&-NanoDrop%208000-Nucleic-Acid-Purity-Ratios.pdf