Isolate Cells Containing a New-found Structure
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For each of the SLP assignments, you will be provided with a hypothetical experimental scenario or data. These assignments are more opened-ended than the case assignments. You will speculate about possible explanations and the ways they might be tested, but be sure to that your hypotheses are grounded in accepted biological science. In doing this, you will mimic the action of scientists who are continuously collecting new data, formulating hypotheses, and testing their ideas.
In a newly-explored deep-sea environment, several potentially new species of apparent bacteria have been discovered. In some of these single-celled life forms, a novel structure has been observed. The structure is apparently bound to the interior surface of the cell membrane and so does not float freely in the cytosol. Preliminary investigation indicates that this structure typically appears circular or ovoid in shape. It apparently consists of a long chain of nucleic acid wrapped around a tube of an unknown protein.
What approach would you use to isolate only cells that contain this new structure? What techniques could be used to characterize the structure and composition of the structure? How might you identify the type of protein and nucleic acid? What would you hypothesize to be the function of this structure? To what eukaryotic organelle might this structure be analogous and why?
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Solution Summary
Cell fractionation is an important enrichment method for isolation and identification of low abundance proteins in the new structure of the bacteria. The structure may contain integral membrane proteins so using this method is ideal. Using Triton X-114, which is a detergent used to solubilize and separate membrane proteins via phase partitioning. Triton X can separate the aqueous and detergent phases at above 20C. As the temperature increases, the phase separate into both hydrophilic and hydrophobic phases. The membrane proteins are in the hydrophobic fractions so we can enriched the protein using the hydrophobic phases. (1) For the extraction of membrane protein, we can use the membrane protein extraction kit.
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Isolation of cells containing this structure-
The first step to isolate the cells that contain this new structure is to find out if the bacteria containing the new structure is gram positive and gram negative.
First, characterize the bacteria to find out if it is gram positive or gram negative by doing a gram-staining procedure. If it is gram positive, then screen the gram positive bacteria on selective media.
If it is gram negative, then screen the gram negative bacteria on gram negative selective media.
Once you know that the new structure is in the gram negative or gram positive bacteria, then you can screen either of them to isolate the cells containing the new structure.
Since the new structure is composed of nucleic acid ( DNA and RNA) and some protein. If the new species is culturable as in you can get gram positive or gram negative colonies, then I would cultured the new single cell species and isolate total nucleic acid (DNA or RNA) by doing a nucleic acid extraction using a lysis detergent (Trizol or SDS). After nucleic acid is isolated, I can run a gel electrophoresis to separate ...
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