Before cloning a gene of interest you have to obtain sufficient amounts of DNA which is usually done by a polmerase chain reaction (PCR). There are variuos methods for cloning PCR amplified DNA fragment, e.g. using restriction enzymes, TA-cloning, TOPOTM-cloning, ligation independent cloning (LIC) or cloning via recombination (GatewayTM, CreatorTM). Furthermore, you can modify your DNA-fragment by site directed mutagenesis, truncation, fusion to other fragments, etc.

Here, we want to give you a selection of guidelines that might be helpful for planning your experiments. We are aware of the fact that we cannot cover all issues in detail, therefore you are invited to send us your questions or comments and make suggestions for improving the content of our website, especially if you find any mistakes.

 

References and further reading

Critical Factors for Successful PCR (1999), Qiagen

Sambrook, J., Fritsch, E.F. & Maniatis, T. (1989) Molecular Cloning: A Laboratory Manual, 2nd Ed., Cold Spring Harbor Laboratory Press, Cold Spring Harbor, NY

PCR Cloning Protocols. From molecular cloning to genetic engineering (1997), (White, B.A., ed.), Humana Press, Totowa, N.J.