Excision of the Tol2 transposable element of the medaka fish Oryzias latipes in Xenopus laevis and Xenopus tropicalis
Excision of the Tol2 transposable element of the medaka fish Oryzias latipes in Xenopus laevis and Xenopus tropicalis
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DOI：
10.1016/j.gene.2004.05.013
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发表时间：
2004-08-18
期刊：
影响因子：
3.5
通讯作者：
Taira, M
中科院分区：
文献类型：
--
作者：
Kawakami, K;Imanaka, K;Taira, M
medaka鱼的Tol2转座因子属于hAT转座子家族。在之前的研究中，我们已经确定了该元件的一个自主成员，它编码一个功能齐全的转座酶，并表明它可以催化斑马鱼生殖谱系中的转座。迄今为止，Tol2元件是脊椎动物中唯一被鉴定出独立成员的天然转座子。我们在此报道了非洲爪蟾和热带爪蟾胚胎中Tol2元素的转座酶依赖性切除。我们将含有非自主Tol2元件的质粒DNA和体外合成的转座酶mRNA共注射到两细胞期胚胎中，并通过聚合酶链反应（PCR）分析注射胚胎中提取的DNA。我们证明了Tol2元件只有在与转座酶mRNA共注射时才能从X laevis和X tropicalis的质粒DNA中切除。在大多数情况下，Tol2序列的完全缺失伴随着在靶序列上添加短DNA序列，表明发生了转座酶依赖性切除。虽然这些足迹是hAT家族转座子切除后产生的特征，但在非洲爪蟾中发现的额外碱基比在斑马鱼中发现的更长，结构更复杂。这可能反映了鱼和青蛙之间参与转位、修复或两者兼而有之的宿主因子活动的差异。我们目前的研究表明，Tol2转座子系统应该作为一种新的遗传工具来开发非洲爪蟾的转基因和诱变方法。(C) 2004 Elsevier B.V.版权所有
The Tol2 transposable element from the medaka fish belongs to the hAT family of transposons. In the previous studies, we have identified an autonomous member of this element, which encodes a fully functional transposase, and have shown that it can catalyze transposition in the zebrafish germ lineage. To date, the Tol2 element is the only natural transposon in vertebrates from which an autonomous member has been identified. We report here transposase-dependent excision of the Tol2 element in Xenopus laevis and Xenopus (Silurana) tropicalis embryos. We coinjected a plasmid DNA containing a nonautonomous Tol2 element and the transposase mRNA synthesized in vitro into two-cell-stage embryos, and analyzed DNA extracted from the injected embryos by polymerase chain reaction (PCR). We demonstrated that the Tol2 element could be excised from the plasmid DNA in both X laevis and X tropicalis only when it was coinjected with the transposase mRNA. In most cases, a complete loss of the Tol2 sequence was accompanied by addition of a short DNA sequence to the target sequence, indicating that transposase-dependent excision occurred. While these footprints were characteristic to those created upon excision of transposons of the hAT family, the additional bases found in Xenopus were longer and their structures were more complicated than those detected upon excision in zebrafish. This may reflect differences in the activities of host factors involved in either transposition, repair, or both between fish and frog. Our present study suggests that the Tol2 transposon system should be used as a novel genetic tool to develop transgenesis and mutagenesis methods in Xenopus. (C) 2004 Elsevier B.V. All rights reserved.