Extraction of Bacterial Plasmid Dna and Analysis of Extracted Dna Samples
Experiment 2
Titles
Extraction of Bacterial Plasmid DNA and Analysis of Extracted DNA Samples
Objectives
* To learn the procedures needed in extracting the bacterial plasmid DNA * To determine the concentration of original DNA sample and purity of prepared DNA sample by using spectrophotometer * To analyze the extracted DNA sample by gel electrophoresis
Materials and methods
(Refer to UDBB2144 Laboratory 2A Manual Principles of biotechnology page 6-10)
Results
1 2 3 4 5 6
Lane | DNA | Amount (L) | Loaded by | 1 | VC 1kb DNA Ladder (Vivantis), 0.5 g | 5 | | 2 | | - | | 3 | Extracted Plasmid (Group 1) | 5 | | 4 | Extracted Plasmid (Group 2) | 5 | | 5 | Extracted Plasmid (Group 3) | 5 | | 6 | Extracted Plasmid (Group 4) | 5 | | Discussion
The A260to A280 ratio of OD reading obtained from the spectrophotometer can be used to determine the purity of DNA extracted. The DNA extracted can be contaminated by the proteins and RNA molecules that were not totally purified from the DNA solution. If the ratio is less than 1.8, then the DNA extracted might be contaminated by proteins. The ratio obtained from the calculation above, 1.8402 indicated the purity DNA extracted in experiment 2. Therefore the DNA extracted can undergo agarose gel electrophoresis with reliable result
However, the result obtained from the gel electrophoresis did not show clear discrete fragments of DNA, but smears, except for the blue DNA ladder marker. One of the possible causes of this result was the concentration of the DNA extracted in the experiment 2. From the calculation, the concentration of original DNA sample was5815µg/ml, which was might be too concentrated for the gel electrophoresis. The most suitable concentration of DNA sample for gel electrophoresis is below80µg/ml, but if the concentration is exceeds 200µg/ml, it will
Titles
Extraction of Bacterial Plasmid DNA and Analysis of Extracted DNA Samples
Objectives
* To learn the procedures needed in extracting the bacterial plasmid DNA * To determine the concentration of original DNA sample and purity of prepared DNA sample by using spectrophotometer * To analyze the extracted DNA sample by gel electrophoresis
Materials and methods
(Refer to UDBB2144 Laboratory 2A Manual Principles of biotechnology page 6-10)
Results
1 2 3 4 5 6
Lane | DNA | Amount (L) | Loaded by | 1 | VC 1kb DNA Ladder (Vivantis), 0.5 g | 5 | | 2 | | - | | 3 | Extracted Plasmid (Group 1) | 5 | | 4 | Extracted Plasmid (Group 2) | 5 | | 5 | Extracted Plasmid (Group 3) | 5 | | 6 | Extracted Plasmid (Group 4) | 5 | | Discussion
The A260to A280 ratio of OD reading obtained from the spectrophotometer can be used to determine the purity of DNA extracted. The DNA extracted can be contaminated by the proteins and RNA molecules that were not totally purified from the DNA solution. If the ratio is less than 1.8, then the DNA extracted might be contaminated by proteins. The ratio obtained from the calculation above, 1.8402 indicated the purity DNA extracted in experiment 2. Therefore the DNA extracted can undergo agarose gel electrophoresis with reliable result
However, the result obtained from the gel electrophoresis did not show clear discrete fragments of DNA, but smears, except for the blue DNA ladder marker. One of the possible causes of this result was the concentration of the DNA extracted in the experiment 2. From the calculation, the concentration of original DNA sample was5815µg/ml, which was might be too concentrated for the gel electrophoresis. The most suitable concentration of DNA sample for gel electrophoresis is below80µg/ml, but if the concentration is exceeds 200µg/ml, it will