Top > Search of International Patents > DNA MOLECULE CODING FOR 5’ UTR ENABLING HIGH RECOMBINANT PROTEIN EXPRESSION IN MONOCOTYLEDONS

DNA MOLECULE CODING FOR 5’ UTR ENABLING HIGH RECOMBINANT PROTEIN EXPRESSION IN MONOCOTYLEDONS

Foreign code F200010004
File No. S2018-0460-C0
Posted date Jan 29, 2020
Country WIPO
International application number 2019JP015506
International publication number WO 2019198724
Date of international filing Apr 9, 2019
Date of international publication Oct 17, 2019
Priority data
  • P2018-077249 (Apr 13, 2018) JP
Title DNA MOLECULE CODING FOR 5’ UTR ENABLING HIGH RECOMBINANT PROTEIN EXPRESSION IN MONOCOTYLEDONS
Abstract The purpose of the present invention is to provide technology for identifying a 5’ UTR having excellent translation ability while taking into account 5’ UTR variants in monocotyledons, providing a DNA molecule that codes for the 5’ UTR, and efficiently producing a recombinant protein in the monocotyledons. It is possible to efficiently produce a recombinant protein in a monocotyledon by using a nucleic acid structure in a polynucleotide that codes for a foreign protein, said structure comprising a base sequence among the base sequences shown in sequences 1-4 and linking to a 5’ UTR or a variant thereof.
Outline of related art and contending technology BACKGROUND ART
Conventionally, the introduction of foreign genes to the plant-established techniques, in addition, the high expression system are constructed, using the production of recombinant proteins in a plant has been actively performed. Is gene expression in plants, largely divided into a translation transfer processes in the course, translation initiation reaction becomes a rate-determining step of the production of a protein are known (Non-Patent Document 1). Translation processes, of the mRNA 5 'located at the end of the structure of the translation initiation factor is bonded to the Cap, ribosomal subunit 40S of the 5' untranslated region (5' UTR) is to start to recruit. Ribosome to the recruitment of mRNA translation efficiency is significantly influenced by the efficiency, the scaffold and the 5' UTR of mRNA translation efficiency is defined to be a very important factor.
Plants, monocots and dicotyledonous plant is divided into a display. Among these, a monocotyledonous or dicotyledonous plant Arabidopsis tobacco and alchol dehydrogenase (ADH) gene isolated from the 5 'UTR sequence (NtADH 5' UTR, AtADH 5' UTR) that incorporates an expression vector, a very high translation capability, and the Arabidopsis tobacco protoplasts using transient expression experiments in, commercially available 87-150 times as compared with the vector in the translational stage pBI221 the expression have been reported to have been improved (Non-Patent Document 2). In addition, a plurality of genome-wide analysis it has been found that, even under various conditions (AtCOR 47) At1g20440 having a high translation capabilities of the 5' UTR sequence, and the Arabidopsis tobacco plants, is shown with its ability, of a plant growth, development or due to thermal stress and to avoid inhibition of translation, whereas the actively contribute to the translation have been reported (Non-Patent Document 3). In this way, in monocotyledonous or dicotyledonous plant such as Arabidopsis thaliana and tobacco, high translation capability 5' UTR sequence is acquired.
On the other hand, in monocotyledonous plants, unlike a monocotyledonous or dicotyledonous plant, as described above has a high translation capabilities is obtained 5' UTR sequence and the ability to provide a translation may not be reported. Specifically, a monocotyledonous or dicotyledonous plant is isolated from the AtADH 5 'UTR dicotyledonous also exhibits the effect to improve the amount of expression, the expression in cultured cells of rice is only 2 times and the improvement effect, in addition, isolated from rice ADH of the 5' UTR sequence (OsADH 5' UTR) even in the case of using, the effect of improving the amount of expression in cultured cells of the rice is at most 9 times which has been reported (Non-Patent Document 2).
Scope of claims (In Japanese)[請求項1]
 以下の(i)~(iii)のいずれかに示すポリヌクレオチドからなることを特徴とする、5'UTRをコードするDNA分子:
(i)配列番号1~4のいずれかに示す塩基配列からなるポリヌクレオチド、
(ii)配列番号1~4のいずれかに示す塩基配列において、1又は数個の塩基が置換、欠失若しくは付加された塩基配列からなり、且つ配列番号1~4のいずれかに示す塩基配列からなるポリヌクレオチドと同等の5'UTR活性を示すポリヌクレオチド、
(iii)配列番号1~4のいずれかに示す塩基配列と相補的な塩基配列からなるDNA断片とストリンジェントな条件下でハイブリダイズし、且つ配列番号1~4のいずれかに示す塩基配列からなるポリヌクレオチドと同等の5'UTR活性を示すポリヌクレオチド。

[請求項2]
 請求項1に記載のDNA分子が、外来タンパク質をコードするポリヌクレオチドの5'末端側に連結されている、核酸構築物。

[請求項3]
 請求項2の記載の核酸構築物を含む、ベクター。

[請求項4]
 請求項3に記載のベクターを単子葉植物又は単子葉植物細胞に導入する、形質転換体の製造方法。

[請求項5]
 請求項3に記載のベクターで、単子葉植物又は単子葉植物細胞が形質転換されてなる、形質転換体。

[請求項6]
 請求項5に記載の形質転換体を培養又は栽培する、組み換えタンパク質の製造方法。
  • Applicant
  • ※All designated countries except for US in the data before July 2012
  • NARA INSTITUTE OF SCIENCE AND TECHNOLOGY
  • Inventor
  • KATO Ko
IPC(International Patent Classification)
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