JPH0430790A - Genetic manipulation method - Google Patents
Genetic manipulation methodInfo
- Publication number
- JPH0430790A JPH0430790A JP13407790A JP13407790A JPH0430790A JP H0430790 A JPH0430790 A JP H0430790A JP 13407790 A JP13407790 A JP 13407790A JP 13407790 A JP13407790 A JP 13407790A JP H0430790 A JPH0430790 A JP H0430790A
- Authority
- JP
- Japan
- Prior art keywords
- gene
- ion beam
- fusion
- damage
- manipulation
- Prior art date
- Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
- Pending
Links
- 238000000034 method Methods 0.000 title claims description 11
- 238000010353 genetic engineering Methods 0.000 title claims description 6
- 108090000623 proteins and genes Proteins 0.000 claims abstract description 26
- 238000010884 ion-beam technique Methods 0.000 claims abstract description 22
- 230000006378 damage Effects 0.000 claims abstract description 17
- 230000004927 fusion Effects 0.000 claims abstract description 14
- 238000003776 cleavage reaction Methods 0.000 claims description 3
- 230000007017 scission Effects 0.000 claims description 3
- 230000001678 irradiating effect Effects 0.000 abstract description 5
- 108090000790 Enzymes Proteins 0.000 abstract description 4
- 102000004190 Enzymes Human genes 0.000 abstract description 4
- 239000000126 substance Substances 0.000 abstract description 3
- GYHNNYVSQQEPJS-UHFFFAOYSA-N Gallium Chemical compound [Ga] GYHNNYVSQQEPJS-UHFFFAOYSA-N 0.000 description 7
- 229910052733 gallium Inorganic materials 0.000 description 7
- 229910052751 metal Inorganic materials 0.000 description 6
- 239000002184 metal Substances 0.000 description 6
- 150000002500 ions Chemical class 0.000 description 5
- WFKWXMTUELFFGS-UHFFFAOYSA-N tungsten Chemical compound [W] WFKWXMTUELFFGS-UHFFFAOYSA-N 0.000 description 5
- CKHJYUSOUQDYEN-UHFFFAOYSA-N gallium(3+) Chemical compound [Ga+3] CKHJYUSOUQDYEN-UHFFFAOYSA-N 0.000 description 4
- 238000000605 extraction Methods 0.000 description 3
- 230000002068 genetic effect Effects 0.000 description 3
- 230000000694 effects Effects 0.000 description 2
- 230000015572 biosynthetic process Effects 0.000 description 1
- 230000007910 cell fusion Effects 0.000 description 1
- 238000006243 chemical reaction Methods 0.000 description 1
- 210000000078 claw Anatomy 0.000 description 1
- 238000007796 conventional method Methods 0.000 description 1
- 238000009792 diffusion process Methods 0.000 description 1
- 230000005686 electrostatic field Effects 0.000 description 1
- 238000010438 heat treatment Methods 0.000 description 1
- 238000003384 imaging method Methods 0.000 description 1
- 238000009940 knitting Methods 0.000 description 1
Abstract
Description
【発明の詳細な説明】
［産業上の利用分野］
本発明は遺伝子−操作の方法に係り、特定の化学溶液あ
るいは酵素等を用いることなく、しかも遺伝子の任意の
箇所での切断、融合、ダメージ発生を行うことができる
遺伝子操作方法に関する。[Detailed Description of the Invention] [Industrial Application Field] The present invention relates to a method of gene manipulation, which allows cutting, fusion, and damage at any location of a gene without using specific chemical solutions or enzymes. The present invention relates to a genetic manipulation method capable of carrying out development.
［従来の技術］
この種の遺伝子操作には、これまで、特定の化学溶液あ
るいは酵素等を用いて遺伝子と遺伝子との間の化学的な
反応を利用して遺伝子の切断、融合あるいはダメージ発
生を生じさせることによって操作する方法が一般に用い
られてきていた。[Prior art] This type of genetic manipulation has so far involved cutting, fusion, or causing damage to genes by utilizing chemical reactions between genes using specific chemical solutions or enzymes. Techniques of manipulation have been commonly used.
［発明が解決しようとする課題］
しかしながら、このような方法による場合には切断、融
合、ダメージ等の発生する箇所が使用溶液あるいは酵素
等の内容によって決定されてしまい、遺伝Ｙ−の任意の
箇所での切断、融合、ダメージを発生させることが不可
能で、適用範囲が自ずから限定されていた。[Problems to be Solved by the Invention] However, when using such a method, the location where cleavage, fusion, damage, etc. occur is determined by the content of the solution or enzyme used, and any location of the genetic Y- It was impossible to cut, fuse, or cause damage, and the range of application was naturally limited.
本発明の目的は、上記従来技術の有していた課題を解決
して、遺伝子の任意の箇所での切断、融合、ダメージ発
生をμｆ能にし、遺伝子操作の適用範囲を拡大すること
にある。The purpose of the present invention is to solve the above-mentioned problems of the prior art, to make it possible to cleave, fuse, and cause damage at any location of a gene, and to expand the scope of application of genetic manipulation.
［課題を解決するための手段］
上記目的は、集束イオンビームを用いて遺伝子を切断、
融合、ダメージ発生を生じさせることによって達成する
ことができる。すなわち、高真空の集束イオンビーム装
置を用い、ビーム径を数十ｎｍから数百ｎｍに絞った高
速の集束イオンビームな遺伝子の所望の箇所に照射する
ことによって達成することができる。[Means for solving the problem] The above purpose is to cut genes using a focused ion beam,
This can be achieved by fusion and damage generation. That is, this can be achieved by using a high-vacuum focused ion beam device and irradiating a desired location on the gene with a high-speed focused ion beam with a beam diameter of several tens of nanometers to several hundred nanometers.
この場合、照射所、望箇所の選定は、上記集束イオンビ
ーム装置においてまず低速低電流のイオンビームを遺伝
子に照射、走査し、放出される電子を利用した結像の観
察によって１１標箇所を選定することによって行い、そ
の後イオンビームを高速高電流のビームに切替え、照射
することによって所望箇所の切断、融合、ダメージを発
生させる。In this case, the irradiation site and desired location were selected by first irradiating and scanning the gene with a low-speed, low-current ion beam using the focused ion beam device described above, and then selecting 11 target locations by observing the imaging using the emitted electrons. After that, the ion beam is switched to a high-speed, high-current beam, and irradiation is performed to cause cutting, fusion, and damage at the desired location.
［作用］
１−記第１段階の低速低電流の集束イオンビームの走査
によって遺伝子に損傷を与えることなしに切断、融合、
ダメージ発生を生ゼしめるべき遺伝子の箇所を任意に選
定することができ、その後の高速高電流集束イオンビー
ムへの切替えによって、該選定箇所での切断、融合、ダ
メージ発生を生じさせることができる。これによって、
従来技術では不可能であった遺伝子の任意箇所での切断
、融合、ダメージの発生を可能にすることができる。[Effect] 1- The scanning of the low-speed, low-current focused ion beam in the first step allows cutting, fusion, and fusion without damaging genes.
The location of the gene where damage should occur can be arbitrarily selected, and by subsequent switching to a high-speed, high-current focused ion beam, cutting, fusion, and damage can be caused at the selected location. by this,
It is possible to cut, fuse, or cause damage at any location in a gene, which was not possible with conventional techniques.
［実施例］
以下、本発明の遺伝子操作方法について実施例によって
具体的に説明する。[Example] Hereinafter, the gene manipulation method of the present invention will be specifically explained with reference to Examples.
第１図は本発明方法の実施に使用する集束イオンビーム
装置の概略構成を示す模式断面図で、先端を尖らせたタ
ングステン線ｌ、金属ガリウム２を収納するガリウム容
器３、ヒータ４、引出し電極５、電源６および７、金属
製静電レンズ極板９、ｌおよび９ｂ、１を源１０、偏向
電極１２ａおよび１２１）、α品向信シＪ−電源１３ａ
および１３１）、試料台口からなり、電源（Ｓ、７．１
０、偏向信′Ｉ！ｆｆｆｌ源１３ａおよび＋３ｂを除い
て容器（図示せず）によって高真空に保持される。FIG. 1 is a schematic cross-sectional view showing the schematic configuration of a focused ion beam device used to carry out the method of the present invention, including a tungsten wire 1 with a sharp tip, a gallium container 3 containing metal gallium 2, a heater 4, and an extraction electrode. 5, power sources 6 and 7, metal electrostatic lens plates 9, l and 9b, 1 as a source 10, deflection electrodes 12a and 121), α quality signal J-power source 13a
and 131), a sample stage opening, and a power supply (S, 7.1
0, deflection 'I! All but the fffl sources 13a and +3b are maintained at high vacuum by vessels (not shown).
ここで、まずヒータ４を加熱することによってガリウム
容器３中のガリウム２を溶融し、表面拡散によりタング
ステン線ｌの表面を軽く儒らす。次に、電源７によって
引出し電極５に（−）、タングステン線ｌに（＋）の高
電圧を印加するとタングステン線１の先端からガリウム
イオンビーム８が放射される。該ガリウムイオンビーム
８は静電レンズ極板９ａおよび９ｂの中心開口部を通過
する際、電源１０による静電界によって集束され、試料
台！１上に置いた試料Ｈの表面に微細スポットとなって
照射される。この微細スポットは２枚の対向する金属板
からなる偏向電極１２ａおよびこれと直角に配置された
同じく２枚の対向する金属板からなる偏向爪１斬１２ｂ
にそれぞれ接続された編向信号・電隙１３．１および１
３！〕の７１圧によって任意に偏向することができ、試
料１１」二の任意の点を選択的に照射させることかでき
る。First, the gallium 2 in the gallium container 3 is melted by heating the heater 4, and the surface of the tungsten wire 1 is slightly softened by surface diffusion. Next, when a high voltage (-) is applied to the extraction electrode 5 and a (+) voltage is applied to the tungsten wire 1 by the power source 7, a gallium ion beam 8 is emitted from the tip of the tungsten wire 1. When the gallium ion beam 8 passes through the center openings of the electrostatic lens plates 9a and 9b, it is focused by the electrostatic field from the power source 10, and is focused on the sample stage! The surface of the sample H placed on the sample H is irradiated in the form of a fine spot. This fine spot is formed by a deflection electrode 12a made of two metal plates facing each other and a deflection claw 12b made of two metal plates placed at right angles thereto.
Knitting direction signal/electric gap 13.1 and 1 connected respectively to
3! ] can be arbitrarily deflected by the 71 pressure, and any point on the sample 11' can be selectively irradiated.
本５Ｑ明の方法を実施するに当っては、まず試料Ｍ＋Ｌ
Ｉ−に載置した試料１１を低速低電流の集束ガリウムイ
オンビームによって照射、該ビームを走査して試ｔ’）
遺伝子１ＪＮＡ上の切断、融合、ダメージを生ぜしめる
べき１１標箇所を電子粘１象による像の観察によって決
定し、その後イオンビームを高速高電流の集束イオンビ
ームに切替え、該箇所の照射を行うことによって切断、
融合、ダメージの発生を生ゼしぬる。In carrying out the method of this 5Q, first, sample M+L
The sample 11 placed on I- is irradiated with a focused gallium ion beam of low speed and low current, and the beam is scanned to perform a test t')
The 11 target locations on gene 1 JNA that should cause cleavage, fusion, and damage are determined by observing the image using electron viscosity, and then the ion beam is switched to a high-speed, high-current focused ion beam to irradiate the locations. cut by,
Fusion, generate damage.
なお、試料を照射するイオン種は上記金属ガリウムに限
定されるものではなく、各種元素の使用が可能であり、
また、イオン源の構成も図示のような構成のイオン源に
限られるものではなく各種イオン源の使用が可能である
。Note that the ion species for irradiating the sample is not limited to the metal gallium mentioned above, and various elements can be used.
Further, the configuration of the ion source is not limited to the ion source having the configuration shown in the drawings, and various ion sources can be used.
細胞の融合の場合には遺伝子形成に必要なＣ１Ｎ、１１
などのイオンビームを用いる。これらのイオン種を用い
、遺伝子の構成を損なわない適切なエネルギーで所望箇
所を照射することにより、生物固有の取り込み能力によ
って自然に融合か行われる。In the case of cell fusion, C1N, 11, which is necessary for gene formation,
An ion beam such as By using these ion species and irradiating the desired location with appropriate energy that does not damage the genetic structure, fusion occurs naturally due to the organism's inherent uptake ability.
［発明の効果］
以１．述べてきたように、遺伝子操作に本発明の操イ１
ノＪθ二を適用することによって、従来技術のイｊして
いた課題を解決して、遺伝子の任意の箇所での切断、融
合、ダメージ発生をｉ＋Ｊ能にし、遺伝子操イ１の適用
範囲をさらに拡大させることができた。[Effects of the invention] Below 1. As mentioned above, the present invention's procedure 1 is useful for genetic manipulation.
By applying NoJθ2, we can solve the problems of conventional technology and make it possible to cut, fuse, and cause damage at any location of a gene, further expanding the scope of application of Genetic Manipulation1. I was able to expand it.
第１図は本発明の遺伝子操作方法の実施に使用する集束
イオンビーム装置の概略構成を示す模式断面図である。
ｌ・・・タングステン線、２・・・金属ガリウム、＝５
・・・ガリウム容器、 ４・・・ヒータ、５・・・引
出し電極、 ６．７．１０・・・電源、８・・・ガ
リウムイオンビーム、９・・・静電レンズ、１１・・・
試料、 １２・・・偏向電極、１３・・・偏向
信号電源
１４・・・試料台。
特許出願人 口本雪信電話味式会社FIG. 1 is a schematic cross-sectional view showing the schematic configuration of a focused ion beam device used to carry out the gene manipulation method of the present invention. l...Tungsten wire, 2...Metal gallium, =5
...Gallium container, 4...Heater, 5...Extraction electrode, 6.7.10...Power source, 8...Gallium ion beam, 9...Electrostatic lens, 11...
Sample, 12... Deflection electrode, 13... Deflection signal power source 14... Sample stage. Patent applicant: Kuchimoto Yukinobu Telephone Ajishiki Company
Claims (1)
メージ発生を生じさせることにより遺伝子操作を行うこ
とを特徴とする遺伝子操作方法。1. A gene manipulation method characterized by performing genetic manipulation by causing gene cleavage, fusion, and damage generation using a focused ion beam.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| JP13407790A JPH0430790A (en) | 1990-05-25 | 1990-05-25 | Genetic manipulation method |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| JP13407790A JPH0430790A (en) | 1990-05-25 | 1990-05-25 | Genetic manipulation method |
Publications (1)
| Publication Number | Publication Date |
|---|---|
| JPH0430790A true JPH0430790A (en) | 1992-02-03 |
Family
ID=15119861
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| JP13407790A Pending JPH0430790A (en) | 1990-05-25 | 1990-05-25 | Genetic manipulation method |
Country Status (1)
| Country | Link |
|---|---|
| JP (1) | JPH0430790A (en) |
-
1990
- 1990-05-25 JP JP13407790A patent/JPH0430790A/en active Pending
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