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    • 6. 发明申请
    • Insect Desiccation Resistance Genes and Uses Thereof
    • 昆虫抗病基因及其用途
    • US20110281349A1
    • 2011-11-17
    • US13103806
    • 2011-05-09
    • Takahiro KikawadaTakashi OkudaMasahiko WatanabeKazuei MitaKeiko Kadono
    • Takahiro KikawadaTakashi OkudaMasahiko WatanabeKazuei MitaKeiko Kadono
    • C12N5/10C12N15/63C07H21/04
    • C07K14/43563
    • An objective of the present invention is to provide polynucleotides encoding insect desiccation resistance proteins, and uses thereof cDNA libraries were produced from Polypedilum vanderplanki larvae in a desiccated state, a P. vanderplanki EST database was constructed, and genes encoding LEA proteins were isolated. This resulted in the successful isolation of three types of novel gene encoding LEA-like proteins (PvLEA1, PvLEA2, and PvLEA3.) When secondary structure predictions and motif searches were performed on the proteins deduced from each of the genes, all three proteins had α-helix-rich structures and LEA_4 motifs, which are characteristic of LEA proteins. Moreover, the recombinant proteins synthesized from PvLEA1, 2 and 3 genes were heat soluble even when boiling, so that PvLEA1, 2 and 3 proteins have highly hydrophilic property as well as plant LEA proteins. Therefore, the three isolated genes were found to be novel P. vanderplanki-derived LEA genes. Furthermore, introduction of these PvLEA 1, 2 and 3 genes into animal cells successfully conferred desiccation resistance to the cells. The present invention provides the first example of LEA genes isolated from insects.
    • 本发明的目的在于提供编码昆虫抗干扰性蛋白质的多核苷酸及其使用方法,从干燥状态的Polypedilum vanderplanki幼虫生产cDNA文库,构建P.vanderplanki EST数据库,分离编码LEA蛋白质的基因。 这导致了三种类型的编码LEA样蛋白的新基因(PvLEA1,PvLEA2和PvLEA3)的成功分离。当对从每个基因推导的蛋白进行二级结构预测和基序搜索时,所有三种蛋白都具有α 富含螺旋的结构和LEA_4基序,是LEA蛋白的特征。 此外,由PvLEA1,2和3基因合成的重组蛋白即使在沸腾时也是热溶性的,因此PvLEA1,2和3蛋白具有高亲水性以及植物LEA蛋白。 因此,发现三个分离的基因是新的P. vanderplanki衍生的LEA基因。 此外,将这些PvLEA 1,2和3基因引入动物细胞成功地赋予细胞干燥抗性。 本发明提供了从昆虫分离的LEA基因的第一个实例。
    • 8. 发明申请
    • Trehalose transporter gene and method of introducing trehalose into cells
    • 海藻糖转运蛋白基因和将海藻糖引入细胞的方法
    • US20090111176A1
    • 2009-04-30
    • US12073457
    • 2008-03-05
    • Takahiro KikawadaTakashi OkudaMasahiko WatanabeAyako SaitoYasushi KanamoriYuichi Nakahara
    • Takahiro KikawadaTakashi OkudaMasahiko WatanabeAyako SaitoYasushi KanamoriYuichi Nakahara
    • C12N5/06C12N15/11C07K14/00C12N15/00
    • C07K14/43563C07K14/43577
    • There are provided trehalose transporter gene and a method of introducing trehalose into cells by using the gene. Candidates for the trehalose transporter genes were searched in P. vanderplanki EST, resulting in being obtained cDNA designated as Tret1. Tret1 encodes a 504 amino acid protein with 12 trans-membrane structures. Tret1 expression was induced by desiccation stress and predominant in the fat body. Functional expression of TRET1 in Xenopus oocytes showed that transport activity was specific for trehalose and independent of extracellular pH and electrochemical membrane potential. The direction of transport of TRET1 was reversible depending on the concentration gradient of trehalose. Apparent Km and Vmax of TRET1 for trehalose were extraordinarily high values. These results indicate that TRET1 is a facilitated, high-capacity trehalose-specific transporter. Tret1 is widespread in insects. Furthermore, TRET1 conferred trehalose permeability upon cells including those of vertebrates as well as insects.
    • 提供海藻糖转运蛋白基因和通过使用该基因将海藻糖引入细胞的方法。 在P. vanderplanki EST中搜索海藻糖转运蛋白基因的候选物,得到获得的命名为Tret1的cDNA。 Tret1编码具有12个跨膜结构的504个氨基酸的蛋白质。 Tret1表达由干燥应激诱导,在脂肪体中占优势。 非洲爪蟾卵母细胞中TRET1的功能表达显示,转运活性对海藻糖是特异性的,独立于细胞外pH和电化学膜电位。 根据海藻糖的浓度梯度,TRET1的转运方向是可逆的。 TRET1对于海藻糖的表观Km和Vmax是非常高的值。 这些结果表明TRET1是一种促进的高容量海藻糖特异性转运蛋白。 Tret1在昆虫中广泛存在。 此外,TRET1在包括脊椎动物以及昆虫的细胞中赋予海藻糖通透性。