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1. 发明公开

EP1016729A1 METHOD FOR SYNTHESIZING cDNA FROM mRNA SAMPLE 审中-公开
标题翻译： VERFAHREN ZUR HERSTELLUNG VON cDNA，AUSGEHEND VON EINER mRNA PROBE
公开(公告)号：EP1016729A1
公开(公告)日：2000-07-05
申请号：EP98943012.9
申请日：1998-09-17
申请人： Nippon Gene Co., Ltd , Agene Research Institute, Co., Ltd.
发明人： SHIMAMOTO, Akira , FURUICHI, Yasuhiro , SHIBATA, Yuko , FUNAKI, Hiroko , OHARA, Eiji , WATAHIKI, Masanori
IPC分类号： C12Q1/68 , C12N15/11 , C12N9/16
CPC分类号： C12N15/1096
摘要： cDNA including the 5'-terminal sequence of full-length mRNA with a cap structure is synthesized from a mRNA sample containing the full-length mRNA with the cap structure and non-full-length mRNA without any cap structure in mixture. At the first step, the phosphate group at 5'-terminus of the non-full-length mRNA in the mRNA sample is removed. At the second step, the cap structure at the 5'-terminus of the full-length mRNA in the mRNA sample is removed. At the third step, an oligoribonucleotide is ligated to the phosphate group at 5'-terminus of mRNA generated through the first and second steps. At the fourth step, mRNA with the oligoribonucleotide ligated at the 5'-terminus thereof at the third step is subjected to a reverse transcriptase process using a short-chain oligonucleotide capable of being annealed to an intermediate sequence within the mRNA as primer, to synthesize a first-strand cDNA.
摘要翻译：包含具有帽结构的全长mRNA的5'末端序列的cDNA由含有具有帽结构的全长mRNA的mRNA样品和不具有任何盖结构的非全长mRNA合成。在第一步，除去mRNA样品中非全长mRNA的5'端的磷酸基。在第二步，除去mRNA样品中全长mRNA的5'端的帽结构。在第三步骤中，通过第一和第二步产生的mRNA的5'末端的寡核苷酸与磷酸基连接。在第四步骤中，使用能够与mRNA内部的中间序列退火的短链寡核苷酸作为引物，在第三步骤的将其在其5'端连接的寡核糖核苷酸的mRNA进行逆转录酶处理，合成第一链cDNA。

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