TRANSFECTION

INDEX Introduction Types of transfection Methods of transfection

INTRODUCTION Introduction of foreign DNA into nucleus of eukaryotic cells is called transfection. TRANSFECTANTS: Cells that have incorporated the foreign DNA either by biochemical or physical processes. Physical methods includes- Electroporation, gene gun method, microinjection. Biochemical method includes- calcium phosphate method, DEAE- dextron, PEG method.

TYPES OF TRANSFECTION TRANSIENT TRANSFECTION In transient transfected cells plasmid  is inserted into the nucleus of the cell, but don’t get integrated Into the chromosome. STABLE TRANSFECTION In stable transfection plasmid DNA is integrated into chromosome and passed on to next generation.

METHODS OF TRANSFECTION Gene gun method/Biolistics Used for injecting cells with genetic information. DNA bearing tungsten/gold particles are called microprojectile which are carried by macrocarrier. When the gun is turned on and the helium flows through. The helium moves the macrocarrier toward the stopping screen. The helium pushes the particles through the stopping screen the target cells for transfection..

MICROINJECTION Direct mechanical insertion of DNA into cell by use of insertion pipette and holding pipette. Diameter of injection pipette is about 0.5- 10.0µm which injects DNA into cytoplasm or nucleus. suction holding pipettes immobilizes the recipient cell.

Calcium phosphate method Discovered in 1973 DNA is mixed in calcium chloride and phosphate buffer forming DNA- CaPO₄ precipitates. Reaction between DNA- CaPO₄ precipitates and actively dividing cells occurs for hours. Cells treated with dimethyl sulfoxide (DMSO) to allow DNA to pass through cell membrane.