Identification of a novel strong and ubiquitous promoter/enhancer in the silkworm Bombyx mori.
Identification of a novel strong and ubiquitous promoter/enhancer in the silkworm Bombyx mori.
复制标题
DOI：
10.1534/g3.114.011643
复制
发表时间：
2014-05-28
期刊：
影响因子：
--
通讯作者：
Sezutsu H
中科院分区：
文献类型：
--
作者：
Tsubota T;Uchino K;Suzuki TK;Tanaka H;Kayukawa T;Shinoda T;Sezutsu H
转基因技术为确定基因功能提供了一个有价值的工具。虽然各种启动子可用于基因过表达，基因敲低，和转基因个体的鉴定，但仍然缺乏通用的启动子用于此类研究，这种缺乏成为瓶颈，特别是对于非模式生物。在这里，我们成功地确定了一个新的强大的和普遍存在的启动子/增强子在家蚕。我们确定了一个独特的家蚕菌株，其报告基因在增强子陷阱菌株的建立过程中表现出强烈和普遍的表达。在该菌株中，转座子被插入到hsp 90的5′UTR中，hsp 90是一种在一系列组织中大量表达的管家基因。为了确定hsp 90的启动子/增强子是否可用于诱导强基因表达，分离了hsp 90的2.9-kb上游基因组片段（hsp90P2.9k），并检测了其转录激活活性。引人注目的是，hsp90P2.9k在家蚕细胞培养物中诱导强烈的基因表达，并且在家蚕的各种组织和发育阶段中也强烈诱导基因表达。hsp90P2.9k在粘虫细胞培养物Sf 9中也表现出显著的启动子/增强子活性，表明该片段可能用作其它鳞翅目昆虫的基因表达工具。我们进一步发现，2.0kb的hsp90P2.9k足以诱导强基因表达。我们相信，该元件将是一系列的研究，如目标基因的过表达，基因敲低和标记基因的表达，不仅在家蚕，而且在其他昆虫物种的价值。
Transgenic techniques offer a valuable tool for determining gene functions. Although various promoters are available for use in gene overexpression, gene knockdown, and identification of transgenic individuals, there is nevertheless a lack of versatile promoters for such studies, and this dearth acts as a bottleneck, especially with regard to nonmodel organisms. Here, we succeeded in identifying a novel strong and ubiquitous promoter/enhancer in the silkworm. We identified a unique silkworm strain whose reporter gene showed strong and ubiquitous expression during the establishment of enhancer trap strains. In this strain, the transposon was inserted into the 5′UTR of hsp90, a housekeeping gene that is abundantly expressed in a range of tissues. To determine whether the promoter/enhancer of hsp90 could be used to induce strong gene expression, a 2.9-kb upstream genomic fragment of hsp90 was isolated (hsp90P2.9k), and its transcriptional activation activity was examined. Strikingly, hsp90P2.9k induced strong gene expression in silkworm cell cultures and also strongly induced gene expression in various tissues and developmental stages of the silkworm. hsp90P2.9k also exhibited significant promoter/enhancer activity in Sf9, a cell culture from the armyworm, suggesting that this fragment might possibly be used as a gene expression tool in other Lepidoptera. We further found that 2.0 kb of hsp90P2.9k is sufficient for the induction of strong gene expression. We believe that this element will be of value for a range of studies such as targeted gene overexpression, gene knockdown and marker gene expression, not only in the silkworm but also in other insect species.
登录
查看更多内容
影响因子：
4.6
作者：
Kayukawa, Takumi;Tateishi, Ken;Shinoda, Tetsuro
通讯作者：
Shinoda, Tetsuro
影响因子：
56.9
作者：
COOLEY, L;KELLEY, R;SPRADLING, A
通讯作者：
SPRADLING, A
影响因子：
2.6
作者：
Gkouvitsas, T.;Kontogiannatos, D.;Kourti, A.
通讯作者：
Kourti, A.
影响因子：
64.8
作者：
Catteruccia, F;Nolan, T;Crisanti, A
通讯作者：
Crisanti, A
影响因子：
2.6
作者：
Lorenzen, M. D.;Kimzey, T.;Beeman, R. W.
通讯作者：
Beeman, R. W.