Transformation

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Introduction of exogenous DNA into a cell is "transformation". Transformation is stimulated by by physically disrupting the cellmembranes so that the genetic material may enter, for example by electroporation:


ii) When the electric field is applied, the ions move according to their charge.

iii) Pathways are formed across the cell membrane allowing DNA to enter.

iv) When the electric field is deactivated, the membrane heals.


Transfection

Transfection and selection
Transfection and selection

One special case of transformation is transfection. Here a vector, like either a viral genome or a plasmid is used, to carry specific DNA of interest into a cell. The cells that are used for transfection are special - they lack genes for production of some nutrients required for growth. Such host cells are called auxotrophic mutants. Some auxotrophic mutants will not receive any plasmids, some will receive plasmids of only one type and some will receive both types of plasmids.

Auxotrophic mutants containing both types of plasmids are selected for by growing them on minimal media lacking two essential nutrients. Only cells that contain the "bait" and "hunter" plasmids will survive. This is because the "bait" and the "hunter" encode the missing nutrients and hence provide the auxotrophic hosts with the required nutrients. Untransfected cells will not grow.

Transfected cells contain both types of plasmids. If the plasmids combine in a proper way transcriptional activity will be restored. Such restoration is detected by a reporter gene. A common reporter gene is LacZ. If LacZ is inserted immediately after the Gal4 promoter so that binding occurs, LacZ is transcribed when the Gal4 promoter is activated and turn those transfected cells blue.

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