The Tol1 element of medaka fish is transposed with only terminal regions and can deliver large DNA fragments into the chromosomes

Abstract

Tol1 is an active DNA-based transposable element residing in the genome of the medaka fish Oryzias latipes. This element belongs to the hAT transposable element family, of which complete copies have relatively long sequences. In addition, we found that Tol1 elements as long as 18 and 20 kb occur in the medaka fish genome. These facts suggest that Tol1 is suitable for carrying large DNA fragments as a gene transfer vector. Focusing on this, we conducted two kinds of manipulations of the element. The first was to eliminate internal regions dispensable for transposition. It was revealed that a Tol1 element consisting of 157-bp left- and 106-bp right-terminal regions could be transposed without a loss of transposition efficiency. Next, we prepared long Tol1 elements by incorporating unrelated DNA fragments into this short Tol1 clone and examined their transposition efficiencies. The transposition frequency decreased as the element size increased. The longest Tol1 element we examined measured 22.1 kb, and its transposition frequency was approximately one fifth that of a 2.1-kb element. However, this frequency was still significantly higher than that of a random integration of DNA into the chromosomes. The element size of 22.1 kb is the longest ever reported for DNA-based elements currently used for mammals. Thus, Tol1 is a superior gene-transfer vector with a large cargo capacity.