Sunday, May 3, 2009

Cloning Stategy

Cloning Procedures:
1. Method for generating DNA fragments
> Mechanical shearing
>Mixing, glass beads
>Restriction enzyme digestion
>Complete or incomplete digestion
>PCR (polymerase chain reaction)
>cDNA synthesis
>Direct synthesis

2. Reaction which join together the foreign DNA to vector
>Homopolymer tailing
>Ligation of cohesive termini
>Blunt end ligation (no linker)
>Linker molecule

3. A mean of introducing DNA to cells
>Transfection with phage DNA
>Transformation with plasmid DNA
>In vitro packaging in to phage coat; transduction with phage or cosmid

4. Screening and selection

A diagram of the overall process of cloning