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    • 5. 发明授权
    • Glyphosate-resistant plants
    • 草甘膦抗性植物
    • US4940835A
    • 1990-07-10
    • US879814
    • 1986-07-07
    • Dilip M. ShahStephen G. RogersRobert B. HorschRobert T. Fraley
    • Dilip M. ShahStephen G. RogersRobert B. HorschRobert T. Fraley
    • C12N15/82
    • C12N15/8275
    • This invention involves a cloning or expression vector comprising a gene which encodes 5-enolpyruvylshikimate-3-phosphate synthase (EPSPS) polypeptide which, when expressed in a plant cell contains a chloroplast transit peptide which allows the polypeptide, or an enzymatically active portion thereof, to be transported from the cytoplasm of the plant cell into a chloroplast in the plant cell, and confers a substantial degree of glyphosate resistance upon the plant cell and plants regenerated therefrom.The EPSPS coding sequence may be ligated to a strong promoter, such as the 35S promoter from cauliflower mosaic virus, to create a chimeric gene. Such genes can be inserted into plant transformation vectors, and subsequently introduced into plant cells. Plant cells transformed using such genes and plants regenerated therefrom have been shown to exhibit a substantial degree of glyphosate resistance.
    • 本发明涉及包含编码5-烯醇丙酮酸莽草酸-3-磷酸合成酶(EPSPS)多肽的基因的克隆或表达载体,其在植物细胞中表达时含有允许多肽的叶绿体转运肽或其酶活性部分, 从植物细胞的细胞质转移到植物细胞中的叶绿体中,并且赋予植物细胞和植物再生的植物大量的草甘膦抗性。 EPSPS编码序列可以连接到强启动子,例如来自花椰菜花叶病毒的35S启动子,以产生嵌合基因。 可将这些基因插入植物转化载体中,随后导入植物细胞。 已经显示使用从其再生的这些基因和植物转化的植物细胞表现出相当程度的草甘膦抗性。
    • 8. 发明授权
    • Plasmids for transforming plant cells
    • 用于转化植物细胞的质粒
    • US08334139B1
    • 2012-12-18
    • US06783336
    • 1985-10-04
    • Robert T. FraleyStephen G. Rogers
    • Robert T. FraleyStephen G. Rogers
    • C12N15/84C12N15/54C12N1/20C12N1/21
    • C12N15/8205
    • This invention relates to several plasmids which are useful for genetically transforming plant cells. A first plasmid, such as pMON120, contains a T-DNA border, one or more marker genes, a unique cleavage site, and a region of Ti plasmid homology. A gene which is expressed in plant cells may be inserted into this plasmid to obtain a derivative plasmid, such as pMON128 which expresses neomycin phosphotransferase in plant cells. The derivative plasmid is inserted into a suitable microorganism, such as A. tumefaciens which contains a Ti plasmid. The inserted plasmids recombine with Ti plasmids to form co-integrate plasmids. Only a single crossover event is required to create the desired co-integrate plasmid. A. tumefaciens cells with co-integrate plasmids are selected and co-cultured with plant cells. The co-integrate Ti plasmids enter the plant cells and insert a segment of T-DNA which does not contain tumorigenic genes into the plant genome. The transformed plant cell(s) may be regenerated into a morphologically normal plant which will pass the inserted gene(s) to its descendants.
    • 本发明涉及可用于遗传转化植物细胞的几种质粒。 第一种质粒如pMON120含有T-DNA边界,一个或多个标记基因,独特的切割位点和Ti质粒同源性的区域。 可以将在植物细胞中表达的基因插入该质粒中,得到在植物细胞中表达新霉素磷酸转移酶的pMON128等衍生质粒。 将衍生质粒插入合适的微生物,如含有Ti质粒的根瘤土壤杆菌。 插入的质粒与Ti质粒重组以形成共同整合质粒。 只需要一个交叉事件来创建所需的共同整合质粒。 选择具有共同整合质粒的根癌农杆菌细胞并与植物细胞共培养。 共同整合的Ti质粒进入植物细胞,并将不含致瘤基因的一段T-DNA插入植物基因组。 转化的植物细胞可以再生成形态上正常的植物,其将插入的基因传递给其后代。
    • 9. 发明授权
    • Modified transcriptionally active SP6 plasmid vector
    • 修饰的转录活性SP6质粒载体
    • US4948731A
    • 1990-08-14
    • US893482
    • 1986-08-05
    • Lee GehrkeRobert T. FraleyStephen G. Rogers
    • Lee GehrkeRobert T. FraleyStephen G. Rogers
    • C12N1/21C12N15/70C12P19/34
    • C12P19/34C12N15/70
    • A recombinant plasmid which may be used for propagation of cloned cDNAs and also for the in vitro synthesis of RNA which is an exact copy of the natural sequence, wherein the transcript is devoid of vector-derived sequence. The novel vector was generated de novo by genetic engineering procedures using a synthetic double strand oligodeoxyribonucleotide fragment and a larger DNA fragment derived from plasmid pSP64. The vector, plasmid pHST-O, carried by Escherichia coli HB101, deposited with the ATCC on Dec. 30, 1985 and designated ATCC 53381, is distinguished from other vectors containing the SP6 promoter by the following characteristics: it contains a unique site for the restriction endonuclease BglII; the engineered GBlII site overlaps the downstream border of the SP6 promoter sequence; and the presence and positioning of the BglII restriction site permit insertion of cDNA molecules in such a way that transcription by the SP6 RNA polymerase begins exactly at the 5' terminus of the RNA, providing that the 5' terminal nucleotide of the mRNA transcript is a guanosine (G) residue, thus excluding the transcription of nucleotides derived from the vector. The novel vector permits synthesis of RNA molecules which have a defined 5' terminus and which are devoid of vector-derived sequence. The vector has potential use in research in molecular biology and in the in vitro production of RNA and proteins.
    • 可用于繁殖克隆cDNA的重组质粒以及用于体外合成RNA的重组质粒,其是天然序列的精确拷贝,其中转录本缺乏载体衍生的序列。 通过使用合成双链寡脱氧核糖核苷酸片段和源自质粒pSP64的较大DNA片段的遗传工程程序从头生成新载体。 1985年12月30日保藏于ATCC并命名为ATCC 53381的大肠杆菌HB101携带的载体质粒pHST-O与含有SP6启动子的其他载体具有以下特征的区别:其含有 限制性内切核酸酶BglII; 设计的GBlII位点与SP6启动子序列的下游边界重叠; 并且BglII限制性位点的存在和定位允许插入cDNA分子,使得由SP6 RNA聚合酶转录正好位于RNA的5'末端,条件是mRNA转录物的5'末端核苷酸是 鸟苷(G)残基,因此不包括衍生自载体的核苷酸的转录。 该新载体允许合成具有定义的5'末端并且没有载体衍生序列的RNA分子。 该载体在分子生物学和RNA和蛋白质的体外生产研究中具有潜在的用途。