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Plasmid preparation - Info
Plasmid preparation
Overview
What is a plasmid preparation?
In addition to the bacterial chromosome, bacteria possess so-called extrachromosomal plasmids on which genes are contained. A characteristic feature of plasmids is their ring shape. They are double-stranded DNA molecules that are able to replicate themselves. For this purpose, a plasmid has a sequence called the origin of replication (ORI).
In modern science, plasmids are used to duplicate genetic material within bacteria and to transfer foreign genes to other organisms.
The process by which plasmid DNA is isolated from transformed bacteria is called plasmid preparation.
Why do you do a plasmid preparation?
Such a plasmid preparation allows scientists to work with the plasmid DNA in vitro, i.e. outside the cell.
The entire process of plasmid preparation can be divided into two sub-steps. First, the cell content is released by the destruction of the cell walls. This step is also known as "unlocking" the cell. The cell extract obtained is then treated in such a way that all components except for the desired plasmid DNA are removed.
What types of plasmid preparation kits exist?
Plasmid preparation kits provide the scientist with all necessary tools to perform such a plasmid preparation. Based on the amount of plasmid DNA that can be isolated, these kits are divided into four groups.
A miniprep kit allows the isolation of up to 25 µg plasmid DNA.
Midiprep kits are used for quantities up to 0.5 mg.
Larger amounts up to 2 mg are isolated using a maxiprep kit.
If these plasmid DNA amounts are not sufficient, a gigaprep kit can be used, which can isolate up to 2 mg plasmid DNA.
A selection of our plasmid preparation kits
| Product | Manufacturer | Item No. | Amount |
|---|---|---|---|
| QuickClean II Plasmid Miniprep Kit |
50 rxns |
||
| Plasmid DNA Extraction Mini Kit (100prep) |
100 preps |
||
| QuickPure Plasmid Mini kit |
200 rxns |
||
| EZ-10 Spin Column Endotoxin-Free Plasmid DNA Mini-Preps Kit |
50 preps |
||
| FavorFilter Plasmid Midi SP Kit (25prep)-spin column |
25 preps |
||
| GoldHi EndoFree Plasmid Midi Kit |
10 rxns |
||
| Plasmid DNA Maxi Endotoxin Removal kit(10prep)-spin column |
10 preps |
||
| GoldHi EndoFree Plasmid Maxi Kit |
10 rxns |
||
| EZ-500 Spin Column Endotoxin-Free Plasmid DNA Maxi-Preps Kit |
20 preps |
What are the differences in a plasmid preparation?
The plasmid preparation can be carried out by means of alkaline lysis or Koch lysis. As the name suggests, the two procedures differ in the methodology for lysis of the bacterial cell wall.
In alkaline lysis, a detergent called sodium dodecyl sulfate (SDS) is used to dissolve the bacterial cell wall and thus make the contents accessible. The addition of sodium hydroxide then causes the pH to rise to alkaline. This has the consequence that the hydrogen bridge bonds between the DNA strands are released. In contrast to chromosomal DNA, however, plasmid DNA is capable of being completely renatured afterwards. The remaining components are then simply centrifuged so that the plasmid DNA remains in the supernatant.
In the case of cooking lysis, the bacterial cell wall is dissolved with the help of the enzyme lysozyme. This is followed by denaturation of the bacterial proteins with isopropyl alcohol. In this procedure, the chromosomal DNA and denatured protein are also centrifuged and the plasmid DNA is extracted from the supernatant.
