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DNA Lab - Atinuke Omolara Biology Lab 2112 5:10-8:00pm...

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ectrophoresis and visualization projected no results, which can be due to a number of experimental errors 2 3 4 5 6 7 8 Lanes rRNA tRNA DNA used in gel electrophoresis (http://www.genscript.com/) Atinuke Omolara Biology Lab 2112 5:10-8:00pm Instructor: Erin Walsh Due Date: December 3, 2008 Title: The Chromosomal DNA Extraction and Analysis of Escherichia coli Introduction: Life involves complex chemical interactions that take place among the many kinds of molecules found within cells. DNA or deoxyribonucleic acid is a molecule that is used to encode genetic information in cells. DNA is found in the nucleus, which is surrounded by a membrane, found in a cell. DNA also has a complex structure which constitutes of two chains running in opposite directions. Also, the chemical make-up of DNA permits it to form a double helix, coiled structure(Bowen). This experiment was designed to practice DNA extraction and analyze if the DNA that was isolated was contaminated with RNA. The DNA was extracted from a bacterium called Escherichia coli (E.coli). DNA extraction is a procedure used to isolate DNA from a cell for further analysis. This process is begun by breaking the cell wall, inhibiting DNAse enzymes, washing the DNA of any lipids or proteins and then precipitating it by adding an alcohol. The DNA extracted will be analyzed by setting up positive and negative controls and then visualized by running it through gel electrophoresis. Gel electrophoresis is a method for analyzing nucleic acid molecules in DNA. The gel is placed in an electrophoresis chamber containing an electrolyte buffer solution. The DNA samples are loaded into the wells made in the gel during casting. A direct current is then applied from a power supply and the nucleic acids migrate through the gels. The pores should separate the linear fragments of nucleic acid according to their size, meaning, the smaller the fragment, the faster it will migrate. In the end, a photograph of the gel is taken under UV light conditions and each lane shows the separation of the components of each sample.
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Methods and Materials: The methods and materials of this experiment can be referred to in the protocol attached to the lab report, with the following exceptions: A list of reactions and controls were given during lab as follows: Tube Samples 1 2 3 4 5 6 7 H 2 O 8 μL 5 μL 8 μL 5 μL 5 μL 5 μL 8 μL Nucleic Acid 10 μL (DNA) 10 μL (DNA) 10 μL (DNA) 10 μL (DNA) 10 μL (DNA) 10 μL (tRNA) 10 μL (tRNA) Buffer 2 μL 2 μL 2 μL 2 μL 2 μL 2 μL 2 μL DNAse ---- 3 μL --- 3 μL 3 μL 3 μL --- Stop Solution 2 μL 2 μL 2 μL 2 μL 2 μL 2 μL 2 μL Incubation period None (Kept on Ice) 15 minutes @37 C ˚ 15 minutes @37 C ˚ 1) 10 [email protected] 60 C before DNA ˚ is added 2) 15 minutes @37 C ˚ 15 minutes @37 C ˚ 15 minutes @37 C ˚ 15 minutes @37 C ˚
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Results: Figure 1 – Bench 1 gel electrophoresis results
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Figure 2 –“Test” gel results
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1. E. coli DNA prep (no incubation)
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