Eukaryotic transcription factors by David S Latchman

By David S Latchman

Transcription, or the method during which DNA produces RNA, is a crucial point of gene expression. Transcription components keep an eye on transcription in the course of improvement and in illness states. As such, it really is serious for researchers to achieve a great knowing of the connection among the constitution of assorted households of transcription components and their functionality, in addition to roles in human affliction. in view that booklet of the Read more...

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These methylated G residues can then be cleaved by exposure to piperidine, whereas no cleavage occurs at unmethylated G residues (Maxam and Gilbert, 1980). A protein bound to the DNA will protect the guanine residues which it contacts from methylation and hence they will not be cleaved upon subsequent piperidine treatment. As in the other footprinting techniques therefore, specific bands produced by such treatment of naked DNA are absent in the protein–DNA sample. Unlike the other methods, however, because cleavage occurs at specific guanine residues, this method identifies specific bases within the DNA which are contacted by the transcription factor protein.

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Following binding, the DNA is treated with a small amount of the enzyme deoxyribonuclease I (DNAseI) which will digest DNA. The digestion conditions are chosen, however, so that each molecule of DNA will be cut once or a very few times by the enzyme. Following digestion the bound protein is removed and the DNA fragments separated by electrophoresis on a polyacrylamide gel capable of resolving DNA fragments differing in size by only one base. This produces a ladder of bands representing the products of DNAseI cutting either one or two or three or four etc.

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