The main difference between genomic DNA and plasmid DNA isolation is that genomic DNA isolation uses strong lysis including the enzymatic or mechanical breakdown of the cell membranes to release the genomic DNA into the solution, while plasmid DNA isolation uses mild alkaline lysis to get plasmid DNA into the solution along with the genomic DNA. Furthermore, in the genomic DNA isolation, following the cell lysis, genomic DNA can be purified from lipid membrane and proteins while in plasmid DNA isolation, neutralization with potassium acetate will separate plasmid DNA from genomic DNA.
Genomic DNA and plasmid DNA isolation are two extraction methods important for DNA isolation to use in different techniques in biotechnology.
Key Areas Covered
1. What is Genomic DNA Isolation
– Definition, Process, Importance
2. What is Plasmid DNA Isolation
– Definition, Process, Importance
3. What are the Similarities Between Genomic DNA and Plasmid DNA Isolation
– Outline of Common Features
4. What is the Difference Between Genomic DNA and Plasmid DNA Isolation
– Comparison of Key Differences
Key Terms
Cell Lysis, Genomic DNA Isolation, Plasmid DNA Isolation
What is Genomic DNA Isolation
Genomic DNA isolation is the extraction procedure of genomic DNA from cells. One of the main features of genomic DNA isolation is that it requires a strong lysis step to release genomic DNA into the solution. Generally, the cell wall can be broken down enzymatically by lysozymes and proteinase K. In contrast, mechanical breakdown of the cell wall is the more universal process done by bead beating on a vortex. Following cell lysis, DNA purification either by phenol-chloroform extraction or other methods is essential for cleaning lipid bilayer, proteins, and other cell debris.
Figure 1: Genomic DNA Isolation
Furthermore, degradation of genomic DNA is one of the main drawbacks that occur during genomic DNA isolation. Therefore, it is essential to keep low temperatures or to use chemical inhibitors to prevent DNase activity while preventing chromosome breakage during extraction.
What is Plasmid DNA Isolation
Plasmid DNA isolation is the extraction process of plasmid DNA from cells. It also keeps plasmid DNA out from genomic DNA of these cells. However, one of the key points of plasmid DNA isolation is performing mild cell lysis procedure such as alkaline lysis. Basically, the buffer of alkaline lysis contains sodium hydroxide and SDS, which completely denature both plasmid DNA and genomic DNA. However, it is essential to prevent excessive denaturation, which may irreversibly denature plasmid DNA.
Figure 2: Plasmid DNA Isolation
The step that follows cell lysis is the neutralization in plasmid DNA isolation. Generally, potassium acetate is the reagent for neutralization. Due to the small size of plasmid DNA compared to genomic DNA, plasmid DNA easily undergoes renaturation while genomic DNA tends to tangle. After that, centrifugation can form a pellet containing genomic DNA bound to proteins. Therefore, plasmid DNA remains in the supernatant. Ultimately, this plasmid DNA can be precipitated with ethanol.
Similarities Between Genomic DNA and Plasmid DNA Isolation
- These are two methods of DNA extraction used in biotechnology.
- The two main steps of both types of DNA isolation are the cell lysis and purification of DNA.
Difference Between Genomic DNA and Plasmid DNA Isolation
Definition
Genomic DNA isolation refers to the technique used to isolate DNA in biological samples, while plasmid DNA isolation refers to the isolation of plasmid DNA from biological samples.
Samples
Genomic DNA can be isolated from different biological samples, while plasmid DNA isolation is mainly from cell cultures of bacteria or fungi.
Cell Lysis
Moreover, cell lysis is the main difference between genomic DNA and plasmid DNA isolation. The genomic DNA isolation uses strong lysis including an enzymatic or mechanical breakdown of the cell membrane to release genomic DNA into the solution while plasmid DNA isolation uses mild alkaline lysis to get plasmid DNA into the solution along with the genomic DNA.
Denaturation
Furthermore, the pH of the cell lysis buffer for genomic DNA isolation is 7-9, which does not denature DNA while the pH of the cell lysis buffer of plasmid DNA isolation is 12.1-12.3, which denatures both plasmid DNA and genomic DNA.
Second Step
Besides, following the cell lysis, genomic DNA can be purified from lipid membrane and proteins while in plasmid DNA isolation, the second step is to neutralize with potassium acetate to separate plasmid DNA from genomic DNA.
During Centrifugation
During centrifugation, genomic DNA appears in the pellet while plasmid DNA appears in the supernatant.
Significance
Also, genomic DNA isolation is a pretty straightforward method, while plasmid DNA isolation is a more sensitive method.
Downstream Applications
The downstream applications of genomic DNA isolation can be PCR or sequencing, while the downstream applications of plasmid DNA isolation can be cloning transfection or gene therapy.
Conclusion
Genomic DNA isolation is a method of extracting DNA from different types of biological samples. Generally, it is a more straightforward method, which involves the harsh breakdown of cell membranes followed by genomic DNA purification from lipid bilayer, proteins, and other cell debris. On the other hand, plasmid DNA isolation is a more sensitive method, which uses mild alkaline lysis to break open cell membranes. Moreover, it contains a neutralization step to separate plasmid DNA from genomic DNA. Finally, plasmid DNA occurs in the supernatant. Therefore, the main difference between genomic DNA and plasmid DNA isolation is the methods of cell lysis and purification of DNA.
References:
1. Kennedy, Suzanne. “Plasmid vs. Genomic DNA Extraction: The Difference.” Bitesize Bio, 20 June 2019, Available Here.
Image Courtesy:
1. “Figure 17 01 02” By CNX OpenStax (CC BY 4.0) via Commons Wikimedia
2. “Preparation-of-plasmid-dna-by-mwaqas-noman-hafeez-khosa-6-638” By Noman-Hafeez khosa (CC BY-SA 4.0) via Commons Wikimedia
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