transformation

Transformation is the process of altering a cell so that it acquires and expresses genetic material acquired from the surrounding environment. Transformation is commonly used to introduce recombinant plasmid DNA into bacterial strains which can transform naturally or can be made by competent for transformation by artificial means. These latter are known as competent bacteria or simply competent cells.

Types of Competent Cells for Transformation

There are two main types of competent cells that can be used for transformation; chemically competent and
electrocompetent
cells.

Chemically Competent Cells

Chemically competent cells are calcium chloride-treated to facilitate attachment of the plasmid DNA to the competent cell membrane. During chemical transformation, the cells are heat-shocked cells are heat-shocked at a precise temperature and length of time (usually 42°C for 30 seconds). This helps to open the pores in the cell membrane so plasmid DNA from the surrounding environment can enter the cell.

Electrocompetent Cells

Electrocompetent cells are prepared for transformation using Electroporation for a few milliseconds. This method uses an electrical pulse to creates pores so that the material can enter the cell from the surrounding environment.

 

 

Chemical transformation


Overview of chemical transformation

   

The basics of chemical transformation.

Transformation is the process by which bacterial cells take up foreign DNA from their environment. Typically, this is done in the lab for two main reasons: to propagate a recombinant plasmid or to obtain the results of a sub-cloning reaction. The procedure shown in this video can be used with most chemically competent cells. We are focusing on the protocol provided with Invitrogen's Top10 strain of competent cell.

Transformation efficiency

An important consideration in selecting either chemically competent or electrocompetent cells is their transformation efficiency. This is expressed as the number of transformants (or colony forming units) per microgram of plasmid DNA (cfu/µg).

The transformation efficiency you need will be largely determined by your application. To meet different cloning requirements, competent cells are available in a wide range of transformation efficiencies - from >1 × 106 for routine cloning to >3 × 1010 cfu/μg pUC DNA for library construction or working with limited amounts of DNA.

 

transformation efficiency

Figure 1.
A range of transformation efficiencies are available to meet your application needs. One Shot® cells are provided at  >5 x 109, 1 x 109, or >1 x 108 cfu/µg.

Determining transformation efficiency

Usually the transformation efficiency of competent cells is measured by transforming them with subsaturating amounts of supercoiled pUC DNA (~10–500 pg). The results are expressed in number of transformants (or colony forming units) per microgram of plasmid DNA (cfu/μg).

Usually the transformation efficiency of competent cells is measured by transforming them with subsaturating amounts of supercoiled pUC DNA (~10–500 pg). The results are expressed in number of transformants (or colony forming units) per microgram of plasmid DNA (cfu/μg).

Transformation Efficiency Calculations

# of coloniesx106 pgx300 µl total
transformation volume
=# of transformants




100 pg transformed DNAµg? µl platedµg plasmid DNA