Recombinant Tol2 transposase with activity in Xenopus embryos.
The Tol2 transposon system is a useful gene transduction technique, but the injection of mRNA is not sufficiently effective in Xenopus embryos to express Tol2 transposase (Tol2TP). To overcome this, we bacterially synthesized recombinant Tol2TP (rTol2TP) protein and showed that rTol2TP efficiently excised the Tol2 element from an injected donor plasmid in Xenopus embryos. Furthermore, injected embryos exhibited uniform and ubiquitous expression of an EGFP reporter gene placed within the Tol2 element. Importantly, size-exclusion chromatography suggests that rTol2TP forms a tetramer, which differs from the reported hexamer formed by Hermes transposase, although both belong to the same hAT family. The use of rTol2TP may facilitate efficient gene transduction in Xenopus, and the biochemical characterization of Tol2TP.
Shibano
Takashi
T
Department of Biological Sciences, Graduate School of Science, University of Tokyo, 7-3-1, Hongo, Bunkyo-ku, Tokyo 113-0033, Japan.
Takeda
Masatoshi
M
Suetake
Isao
I
Kawakami
Koichi
K
Asashima
Makoto
M
Tajima
Shoji
S
Taira
Masanori
M
D000818:Animals; D001483:Base Sequence; D002850:Chromatography, Gel; D004251:DNA Transposable Elements / Q000235:genetics; D004926:Escherichia coli / Q000235:genetics; D049452:Green Fluorescent Proteins / Q000235:genetics / Q000378:metabolism; D008845:Microinjections; D009990:Oryzias / Q000235:genetics; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*; D014170:Transformation, Genetic; D019895:Transposases / Q000096:biosynthesis / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*; D014981:Xenopus / Q000145:classification / Q000196:embryology* / Q000235:genetics / Q000378:metabolism*
D004251:DNA Transposable Elements; D011994:Recombinant Proteins; C485184:enhanced green fluorescent protein; D049452:Green Fluorescent Proteins; D019895:Transposases