Alkaline lysis
Encyclopedia
Alkaline lysis is a method used in molecular biology
to break cells open to isolate plasmid
DNA
or other cell components such as protein
s. Bacteria containing the plasmid of interest are first grown, then lysed with a strong alkaline buffer consisting of a detergent sodium dodecyl sulfate
(SDS) and a strong base sodium hydroxide. The detergent breaks the membrane's phospholipid
bilayer and the alkali denatures proteins involved in maintaining the structure of the cell membrane. Through a series of steps involving agitation, precipitation, centrifugation, and the removal of supernatant, cellular debris is removed and the plasmid is isolated and purified. The protocol may vary slightly from lab to lab.
Also, alkaline lysis is sometimes used to extract plant genetic material. The plant cells are subjected to a strongly alkaline solution containing a detergent (usually a zwitterionic or nonionic detergent
such as Tween 20), and the mixture is incubated at high temperature. This method is not used as often due to the sodium hydroxide's tendency to damage genetic material, reducing DNA fragment size.
Molecular biology
Molecular biology is the branch of biology that deals with the molecular basis of biological activity. This field overlaps with other areas of biology and chemistry, particularly genetics and biochemistry...
to break cells open to isolate plasmid
Plasmid
In microbiology and genetics, a plasmid is a DNA molecule that is separate from, and can replicate independently of, the chromosomal DNA. They are double-stranded and, in many cases, circular...
DNA
DNA
Deoxyribonucleic acid is a nucleic acid that contains the genetic instructions used in the development and functioning of all known living organisms . The DNA segments that carry this genetic information are called genes, but other DNA sequences have structural purposes, or are involved in...
or other cell components such as protein
Protein
Proteins are biochemical compounds consisting of one or more polypeptides typically folded into a globular or fibrous form, facilitating a biological function. A polypeptide is a single linear polymer chain of amino acids bonded together by peptide bonds between the carboxyl and amino groups of...
s. Bacteria containing the plasmid of interest are first grown, then lysed with a strong alkaline buffer consisting of a detergent sodium dodecyl sulfate
Sodium dodecyl sulfate
Sodium dodecyl sulfate , sodium laurilsulfate or sodium lauryl sulfate is an organic compound with the formula CH311OSO3Na). It is an anionic surfactant used in many cleaning and hygiene products...
(SDS) and a strong base sodium hydroxide. The detergent breaks the membrane's phospholipid
Phospholipid
Phospholipids are a class of lipids that are a major component of all cell membranes as they can form lipid bilayers. Most phospholipids contain a diglyceride, a phosphate group, and a simple organic molecule such as choline; one exception to this rule is sphingomyelin, which is derived from...
bilayer and the alkali denatures proteins involved in maintaining the structure of the cell membrane. Through a series of steps involving agitation, precipitation, centrifugation, and the removal of supernatant, cellular debris is removed and the plasmid is isolated and purified. The protocol may vary slightly from lab to lab.
Also, alkaline lysis is sometimes used to extract plant genetic material. The plant cells are subjected to a strongly alkaline solution containing a detergent (usually a zwitterionic or nonionic detergent
Detergent
A detergent is a surfactant or a mixture of surfactants with "cleaning properties in dilute solutions." In common usage, "detergent" refers to alkylbenzenesulfonates, a family of compounds that are similar to soap but are less affected by hard water...
such as Tween 20), and the mixture is incubated at high temperature. This method is not used as often due to the sodium hydroxide's tendency to damage genetic material, reducing DNA fragment size.