Ligation-During-Amplification (LDA) for DNA modification

Polymerase chain reaction has traditionally been used for in vitro modification and amplification of linear DNA. On the other hand, circular closed loop DNA is often the form replicated in host cells. In order to modify host cell DNA constructs, the traditional method has been to work with linearized DNA that must then be cloned back into a circular form before its reintroduction into the host cell. Ligation-during-amplification (LDA) represents a novel approach for the in vitro amplification of closed circular DNA, thereby eliminating the usual required cloning steps following gene manipulation. The essence of LDA is the inclusion of a thermostable DNA ligase in a PCR reaction that uses closed circular DNA as template. Thus, LDA is a powerful tool to improve not only the processes of site-directed mutagenesis and DNA modification, but also for mutation detection, DNA library screening and circular DNA production.

This invention describes a molecular biology process for improved DNA manipulation. Methods to improve genetic engineering have very broad applications in the development of kits for use not only in basic research, but also agricultural and general healthcare needs.

Stage of Development
Patent 6,620,597 has been issued by the United States Patent and Trademark Office for this invention. The invention�s technology is part of an active research program and has been shown to work in proof of concept experiments which include a working prototype. It is available for licensing through either an exclusive or non-exclusive license.

Additional Info
*Zhidong Chen and Duane Ruffner, �Amplification of closed circular DNA in vitro�, Nuc. Acids Res. Vol. 26(4), 1126-1127 (Feb. 1998).
*Also see corrections in Nuc. Acids Res. Vol. 26(23), 1126-1127 (Dec. 1998).

Inventor(s): Zhidong Chen, Duane Ruffner

Type of Offer: Licensing

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