ARS | Publication request: CHIMERIC GENE CONSTRUCTION WITHOUT REFERENCE TO RESTRICTION SITES

Submitted to: Biotechniques
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: January 22, 2002
Publication Date: April 4, 2002

Interpretive Summary: In this report, we describe how we developed a method that will help genetic researchers perform very challenging laboratory work in order to obtain scientific answers to important medical questions. This work involves the manipulation of DNA to assemble what are called chimeras, which artistically may be said to represent genetic mosaics, or creative new combinations of different entities. The word "chimera" is well chosen for this scientific application, as the mythological chimera had the head of a lion, the body of a goat, and the tail of a serpent. Genetic researchers may make these chimeras by putting together parts of two different genes in different ways to make comparisons, determine commonalities, and ultimately identify the specific area in a gene that enables it to function in a certain way, for example, to provide a particular benefit. Thus, they may reconfigure pieces of a genetic puzzle in a manner not unlike solving a Rubik's cube. This innovative method bypasses thorny obstacles and tricky problems often encountered by genetic researchers in attempting to execute chimeric constructs, and therefore should prove very helpful.

Technical Abstract: This manuscript details a novel method to interchange similar regions between related open-reading frames. This method utilizes class II restriction enzymes and does not require any common restriction sites between the interchanged sequences. This method requires only that PCR be performed and the final product be sequenced. This rapid methodology avoids many of the current complications associated with chimeric gene construction and is universally applicable.