发明名称 |
Identification of eukaryotic internal ribosome entry site (IRES) elements |
摘要 |
A method of search for and identification of a eukaryotic IRES element active in cap-independent translation of RNA in eukaryotic cells is provided, comprising the following steps: i) screening eukaryotic mRNA sequences or corresponding DNA sequences for a potential IRES element having a block of nucleotides having: a) a length of at least 30 nucleotides; b) an adenine nucleotide content of at least 40 mol-%; and c) a pyrimidine nucleotide content of less than 40 mol-%; ii) inserting said potential IRES element into a linear dicistronic construct between an upstream gene and a downstream GUS reporter gene, whereby said potential IRES element is positioned for IRES-dependent translation of said downstream GUS gene and whereby said upstream gene is preceded by a stable hairpin structure to prevent IRES-independent translation of said genes; and iii) testing said potential IRES element for IRES-dependent translation of said GUS gene in a rabbit reticulocyte lysate or in a wheat germ extract in vitro translation assay, whereby GUS gene expression is quantitated preferably relative to a construct having a reference IRES element or a non-IRES element between said upstream gene and said GUS gene. |
申请公布号 |
US8257945(B2) |
申请公布日期 |
2012.09.04 |
申请号 |
US20040489136 |
申请日期 |
2004.03.16 |
申请人 |
ATABEKOV JOSEPH;DOROKHOV YURII;SKULACHEV MAXIM;IVANOV PETER;GLEBA YURI;ICON GENETICS, INC. |
发明人 |
ATABEKOV JOSEPH;DOROKHOV YURII;SKULACHEV MAXIM;IVANOV PETER;GLEBA YURI |
分类号 |
C12N15/00;C12N15/09;A01H5/00;A01K67/027;C12N1/15;C12N1/19;C12N5/10;C12N15/10;C12N15/11;C12N15/63;C12N15/67;C12N15/82;C12N15/85;C12Q1/68;G06F19/00 |
主分类号 |
C12N15/00 |
代理机构 |
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代理人 |
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主权项 |
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地址 |
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