This document describes the transformation of competent E. coli cells using the CaCl2 method. It defines competent cells as cells that can uptake exogenous DNA and discusses plasmids, which are self-replicating DNA that can carry useful genes. The document explains that transformation can be natural or artificial, and the artificial process involves making cells competent through ice-cold CaCl2 treatment, then applying heat shock to induce the cells to uptake exogenous DNA like plasmids.

1. TRANSFORMATION
of competent E. coli using CaCl2
GULPREET KAUR
Department of Biological Sciences
Florida Institute of Technology, FL 32901

2. OBJECTIVES
• Define a Competent Cell and its
Transformation
• Describe the theory of transformation of
bacterial cells with plasmid using CaCl2
method.
• Describe the method of preparation of
Competent cells using CaCl2 and their
transformation with plasmids

3. What is Transformation?

4. PLASMIDS
• Self-replicating DNA
• Double stranded, Circular
• 1 - 1000kbps
• May carry useful genes
– Antibiotic Resistance
– Ability to undergo sexual conjugation

5. TRANSFORMATION MAY BE NATURAL OR
ARTIFICIAL (induced in the laboratory)

6. COMPETENT CELLS
• Treated cells
• Can uptake exogenous DNA
• Uptake is through membrane

7. THEORY OF TRANSFORMATION
1. Ice-cold CaCl2 Treatment – Cell becomes competent
2. Heat shock Treatment - Cell uptakes exogeneous DNA
Fig 5.3. Brown, T.A. 1990. Gene Cloning and DNA Analysis

8. Preparation of Competent Cells
CALCIUM CHLORIDE TREATMENT –
KEEP CELLS COLD!

9. Transformation
of Competent
Cells
TRANSFORMANTS!

10. Transformation efficiency of this procedure:
VERY LOW

11. QUESTIONS?
• What will happen to the cells if they are not
kept cold while preparing competent cells?

12. QUESTIONS?
• Describe the theory of transformation of a
bacterial cell with a plasmid.

13. QUESTIONS?
• A competent cell can take up linear DNA from
its surroundings. True or False?