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Efficient Chromosomal Transposition of a Tc1/mariner-like Transposon Sleeping Beauty in Mice

 



 

Introduction

In post-genome-sequence era, one of the major goals is to identify the functions of a large number of novel genes that have been determined by sequencing. The relationship between a gene and its functions can be analyzed by generating a gene knock-out animal. Analysis of the phenotype of such an animal makes it possible to search for gene functions in vivo.

Availability of mouse embryonic stem (ES) cells allows for the knocking out of a gene of interest. This has resulted in a genetic tool for examining gene functions of a gene of interest in mice. This tool, known as “reverse genetics”, requires gene information before a gene of interest is knocked out. Therefore, it is not suitable for a genome-wide analysis of gene functions due to the lack of comprehensive genome information. In contrast, “forward genetics”, the identification of mutated genes responsible for specific phenotypes, does not require such information, so that genome-wide analysis of genes and identification of functions of new genes become possible.

To date, no efficient method for “forward genetics” in mice has been established. In the study presented here, we demonstrated that a novel transposon, Sleeping Beauty , transposes efficiently in mice and facilitates “forward genetics”.

Transposon is a mobile genetic element

Dr. Barbara McClintock, who won the 1983 Nobel Prize in Physiology or Medicine, discovered a mobile genetic element, transposon, in corn more than fifty years ago. Before this discovery it was believed that genes have fixed positions on chromosomes and never move, a belief which is at the core of Mendelian genetics.

The discovery that the transposon is able to move around chromosomes (Fig. 1) dramatically changed the concept of the genome from static to dynamic.

The transposon system has two components, an active transposase and the DNA sequences that are recognized and mobilized by the transposase. The transposon system was later found in many organisms such as C. elegans, fruitfly, plant and even in yeast and was applied to “forward genetics” by using the characteristics of random insertion into the genome.

Transposon system did not work in mammals until recently

The fact that the transposon system cannot be used in mammals has hampered an efficient “forward genetics” in mice. However, Ivics et. al reported in 1997 that Sleeping Beauty (SB) transposon derived from fish was active in mammalian cells1. Since the SB transposon system was originally inactive in fish due to many mutations in transposase, the authors reconstructed an active transposase from an inactive one in vitro. Although the result suggested the possibility that the transposon system could be utilized for “forward genetics”, a subsequent study demonstrated that the efficiency of transposon-mediated transposition was extremely low in mammalian culture cells2.

 

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