The homing endonucleases are a collection of endonucleases encoded either as freestanding genes within introns, as fusions with host proteins, or as self-splicing inteins. They catalyze the hydrolysis of genomic DNA within the cells that synthesize them, but do so at very few, or even singular, locations. Repair of the hydrolyzed DNA by the host cell frequently results in the gene encoding the homing endonuclease having been copied into the cleavage site, hence the term 'homing' to describe the movement of these genes. Homing endonucleases can thereby transmit their genes horizontally within a host population, increasing their allele frequency at greater than Mendelian rates.

Although the origin and function of homing endonucleases is still being researched, the most established hypothesis considers them as selfish genetic elements, similar to transposons, because they facilitate the perpetuation of the genetic elements that encode them independent of providing a functional attribute to the host organism.

Homing endonuclease recognition sequences are long enough to occur randomly only with a very low probability (approximately once every 7009700000000000000♠7×10⁹ bp), and are normally found in one or very few instances per genome. Generally, owing to the homing mechanism, the gene encoding the endonuclease (the HEG, "homing endonuclease gene") is located within the recognition sequence which the enzyme cuts, thus interrupting the homing endonuclease recognition sequence and limiting DNA cutting only to sites that do not (yet) carry the HEG.

Prior to transmission, one allele carries the gene (HEG⁺) while the other does not (HEG⁻), and is therefore susceptible to being cut by the enzyme. Once the enzyme is synthesized, it breaks the chromosome in the HEG⁻ allele, initiating a response from the cellular DNA repair system. The damage is repaired using recombination, taking the pattern of the opposite, undamaged DNA allele, HEG⁺, that contains the gene for the endonuclease. Thus, the gene is copied to the allele that initially did not have it and it is propagated through successive generations. This process is called "homing".

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