DNA laddering is a feature that can be observed when DNA fragments, resulting from Apoptosis DNA fragmentation are visualized after separation by gel electrophoresis the first described in 1980 by Andrew Wyllie at the University Edinburgh medical school DNA fragments can also be detected in cells that underwent necrosis, but when these DNA fragments after separation are subjected to gel electrophoresis no clear "ladder" pattern is apparent.

DNA degradation

DNA laddering is a distinctive feature of DNA degraded by caspase-activated DNase (CAD), which is a key event during

apoptosis Apoptosis (from ) is a form of programmed cell death that occurs in multicellular organisms and in some eukaryotic, single-celled microorganisms such as yeast. Biochemistry, Biochemical events lead to characteristic cell changes (Morphology (biol ...

. CAD cleaves genomic DNA at internucleosomal linker regions, resulting in DNA fragments that are multiples of 180–185

base-pair A base pair (bp) is a fundamental unit of double-stranded nucleic acids consisting of two nucleobases bound to each other by hydrogen bonds. They form the building blocks of the DNA double helix and contribute to the folded structure of both DNA ...

s in length. Separation of the fragments by agarose gel electrophoresis and subsequent visualization, for example by

ethidium bromide Ethidium bromide (or homidium bromide, chloride salt homidium chloride) is an intercalating agent commonly used as a fluorescent tag (nucleic acid stain) in molecular biology laboratories for techniques such as agarose gel electrophoresis. It ...

staining, results in a characteristic "ladder" pattern. A simple method of selective extraction of fragmented DNA from apoptotic cells without the presence of high molecular weight DNA sections, generating the laddering pattern, utilizes pretreatment of cells in

ethanol Ethanol (also called ethyl alcohol, grain alcohol, drinking alcohol, or simply alcohol) is an organic compound with the chemical formula . It is an Alcohol (chemistry), alcohol, with its formula also written as , or EtOH, where Et is the ps ...

Apoptosis and necrosis

While most of the morphological features of apoptotic cells are short-lived, DNA laddering can be used as final state read-out method and has therefore become a reliable method to distinguish

from

necrosis Necrosis () is a form of cell injury which results in the premature death of cells in living tissue by autolysis. The term "necrosis" came about in the mid-19th century and is commonly attributed to German pathologist Rudolf Virchow, who i ...

. DNA laddering can also be used to see if cells underwent apoptosis in the presence of a virus. This is useful because it can help determine the effects a virus has on a cell. DNA laddering can only be used to detect apoptosis during the later stages of apoptosis. This is due to DNA fragmentation taking place in a later stage of the apoptosis process. DNA laddering is used to test for apoptosis of many cells, and is not accurate at testing for only a few cells that committed apoptosis. To enhance the accuracy in testing for apoptosis, other assays are used along with DNA laddering such as TEM and TUNEL. With recent improvements to DNA laddering, DNA laddering has become a more reliable, and reasonable technique to use when detecting apoptosis. It is also important to note that DNA laddering occurs differently depending on the type of cell, so there may be slight changes in the process of DNA laddering depending on the cell that is being investigated.

References

{{reflist Apoptosis Biological techniques and tools Cell biology Electrophoresis Laboratory techniques Programmed cell death

DNA degradation

Apoptosis and necrosis

See also

References