Dna Report

4127 Words Aug 26th, 2015 17 Pages
Name: Sabina Shrestha Name partner: Marieke RoosStudent Numbers: 1215671/4228073 Practicum assistants: Brijith Thomas,Room: 1 Joanna Pawlak, Lara van der Woude, Date: 12/12/2014 Valerie SelsEmail: saburo_aikini441@yahoo.com |

Lab Report DNA: Plasmids and Nucleases

1. Abstract
The goal of this practicum was to isolate plasmid DNA from Escherichia coli (E. coli), to identify it, to prove that the plasmid is circular and double-stranded and to give bacterial cells new genetic properties via transformation. An unknown plasmid S was isolated from the
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23-24)
The calculation of stock solutions can be found in the appendices.

3.3. Plasmid yield & Purification
3.3.A. Agarose-gel electrophoresis
This experiment was carried out by following the instructions given in the handbook (De Smit & Vijgenboom, 2014, p. 25-26).
3.3.B. Purity A260/A280
This experiment was carried out by following the instructions given in the handbook (De Smit & Vijgenboom, 2014, p. 27).
The calculation of purity can be found in the appendices.
3.4. Identification of plasmid
This experiment was carried out with the aim of identifying unknown plasmid S while comparing it with reference plasmids indicated in the handbook (De Smit & Vijgenboom, 2014, p. 27-28). The amount of DNA used for digesting was 3µl, determined by comparing the band pattern seen in the photo of isolated plasmid in gel (see section 3.3 A). Then a DNA stock solution was prepared by following the scheme shown in Table 2 (see appendices).

3.5. Transformation of E. COLI The transformation began with the plating of bacteria on agar plates, one without antibiotic and the other with a suitable antibiotic. After one night incubation of bacteria at 37 °C, the number of colonies multiplied were counted and the amount of plasmid transformed was estimated. The details of the procedures can be found in the handbook ( De Smit &

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