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Transformation Of Escherichia Bacteria And Dna

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Transformation of Escherichia coli in different concentrations of Plasmid DNA

Introduction
This report discusses an experiment which students have to transform and plate competent Escherichia coli in different concentrations of plasmid DNA. This experiment uses four concentrations of plasmid DNA to perform four transformations. These concentrations are namely, 5 µl of distilled water, which acts as the control in this experiment; 2.5 µl of undiluted plasmid DNA; 1.0µl of undiluted plasmid DNA; and lastly, 1.0µl of plasmid DNA diluted 1 in 10. Transformation is an important aspect in genetic engineering as it allows for a particular DNA to enter into another cell. The aim of this experiment is to be able to perform a cell transformation
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Then, we placed 100 µl of the E. coli cells into four Eppendorf tubes each and labelled them A, B, C and D. After filling them up, we placed them into an ice box. After this, we needed to perform the transformation process by mixing the E. coli cells with different concentrations of plasmid DNA. In test tube A, we added 5 µl of distilled water to the E. coli cells. In test tube B, we added 2.5 µl of undiluted plasmid DNA to the E. coli cells. In test tube C, we added 1.0 µl of undiluted plasmid DNA to the E. coli cells. Lastly, in test tube D, we added 1.0 µl of plasmid DNA diluted 1 in 10 to the E. coli cells. Then, we used a P100 pipette and mixed all the liquids in all the Eppendorf tubes by pipetting the solution up and down. Then, we left the Eppendorf tubes along with its contents in the ice box for twenty minutes. After twenty minutes, we labelled the base of the four agar plates to match with the appropriate plasmid DNA concentration about to be plated. We also wrote our initials, the date, and the table number on the base of the agar plate. We then gently lifted up the cover of the agar plate with just enough space to allow us to plate each tube of cells onto four different agar plates – one agar plate to one plasmid DNA concentration. We used a spreader to spread the cells evenly over the surface of the agar, being careful not let the spreader touch
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