JPH01199521A - Production of butterbur by shoot apex culture - Google Patents

Production of butterbur by shoot apex culture

Info

Publication number
JPH01199521A
JPH01199521A JP2242188A JP2242188A JPH01199521A JP H01199521 A JPH01199521 A JP H01199521A JP 2242188 A JP2242188 A JP 2242188A JP 2242188 A JP2242188 A JP 2242188A JP H01199521 A JPH01199521 A JP H01199521A
Authority
JP
Japan
Prior art keywords
shoot
butterbur
plant growth
transplanted
growth hormone
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.)
Granted
Application number
JP2242188A
Other languages
Japanese (ja)
Other versions
JPH0351377B2 (en
Inventor
Ryuso Tanaka
田中 隆荘
Kenji Taniguchi
谷口 研至
Hideki Miyagawa
宮川 秀樹
Akira Murakami
章 村上
Kunimutsu Murakami
邦睦 村上
Current Assignee (The listed assignees may be inaccurate. Google has not performed a legal analysis and makes no representation or warranty as to the accuracy of the list.)
Sanyo Kokusaku Pulp Co Ltd
Original Assignee
Sanyo Kokusaku Pulp Co Ltd
Priority date (The priority date 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 date listed.)
Filing date
Publication date
Application filed by Sanyo Kokusaku Pulp Co Ltd filed Critical Sanyo Kokusaku Pulp Co Ltd
Priority to JP2242188A priority Critical patent/JPH01199521A/en
Publication of JPH01199521A publication Critical patent/JPH01199521A/en
Publication of JPH0351377B2 publication Critical patent/JPH0351377B2/ja
Granted legal-status Critical Current

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  • Breeding Of Plants And Reproduction By Means Of Culturing (AREA)

Abstract

PURPOSE:To provide the title production process so designed that the shoot apexes of a butterbur are picked out and transplanted to an artificial culture medium containing inorganic salt and plant growth hormone to be put to rotary culture under lighting followed by conversion into a plant on a second artificial culture medium thus obtaining uniform nursery in a short period. CONSTITUTION:The shoot apexes of a butterbur are picked out and transplanted to an artificial culture medium containing inorganic salt and plant growth hormone to be put to rotary culture under lighting to effect induction, retention and proliferation of the nursery base. Thence, the resultant nursery base is transplanted to a second artificial culture medium with the kind and/or concentration of plant growth hormone differing from those (that) in the first culture medium to be put to subculture, thus converting said into a plant.

Description

【発明の詳細な説明】 (産業上の利用分野〕 本発明はフキ属の茎頂培養によるクローン増殖および無
病の苗を短期間に得ることを目的とした大量増殖法に関
するものであり、農業、組織培養等に利用出来る。
Detailed Description of the Invention (Industrial Application Field) The present invention relates to clonal propagation of the genus Butterbur by shoot apical culture and a mass propagation method aimed at obtaining disease-free seedlings in a short period of time, and relates to agriculture, It can be used for tissue culture, etc.

〔従来の技術及び発明が解決しようとする課題〕フキ(
Petasites japonicus)は日本に自
生し、広く栽培されている。しかし、栽培種は3倍体で
あるため不稔性で種子が得られない。したがって株分け
によってのみ繁殖が行われている。
[Problems to be solved by conventional technology and invention] Butterbur (
Petasites japonicus) is native to Japan and widely cultivated. However, since the cultivated species is triploid, it is sterile and cannot produce seeds. Therefore, propagation is carried out only by division.

そのためウィルス罹病株が増え、生長が不良となりまた
不揃いになるなど収量、品質の低下が大きな問題となっ
ている。その対策として現在茎頂を用い無病苗を作出す
る試みが行われているが、未だクローン増殖および無病
の苗を短期間に供給する方法は見いだされていない。
As a result, the number of virus-infected plants has increased, resulting in poor and uneven growth, resulting in a decline in yield and quality, which has become a major problem. As a countermeasure, attempts are currently being made to produce disease-free seedlings using shoot tips, but a method for clonal propagation and supplying disease-free seedlings in a short period of time has not yet been found.

〔課題を解決するための手段〕[Means to solve the problem]

そこで本発明者らは上記問題点を解決すべく鋭意努力し
た結果、フキ属の茎頂部を用い無機塩類および植物生長
ホルモンを含む人工培地に移植して鍼明下に回転培養す
ることにより、苗条原基を誘導、維持、増殖させ、ざら
に固定培地で植物体への転換をはかることによってクロ
ーン増殖および無病の苗を短時間に得る方法を見いだし
た。
As a result of our earnest efforts to solve the above-mentioned problems, the present inventors transplanted the shoot apices of the genus Butterbur to an artificial medium containing inorganic salts and plant growth hormones, and then cultured them in rotation under acupuncture. We have discovered a method for clonal propagation and disease-free seedlings in a short period of time by inducing, maintaining, and propagating primordia and converting them into plants using a fixed medium.

即ち本発明の要旨とする所はフキ属の茎頂部を摘出しこ
れを無機塩類および植物生長ホルモンを含む人工培地に
移植して照明下に回転培養することにより苗条原基の初
期誘導を行い、然る後該苗条原基を前記の人工培地とは
植物生長ホルモンの種類および/または濃度の異った人
工培地に移植して継代培養することにより継持増殖させ
ることを特徴とするフキ属の茎頂培養による生産方法に
存する。
That is, the gist of the present invention is to remove the shoot apex of the genus Butterbur, transplant it to an artificial medium containing inorganic salts and plant growth hormones, and perform rotational culture under illumination to initially induce shoot primordia. Thereafter, the shoot primordium is transplanted to an artificial medium containing a different type and/or concentration of a plant growth hormone from the artificial medium and subcultured, thereby allowing the genus Butterbur to be continuously propagated. It consists in a production method using shoot apical culture.

本発明法で増殖される苗条原基は遺伝的に極めて安定で
かつ増殖率が高くまた植物体への転換も容易に起きる分
裂細胞集塊である。即ち、本発明は一種の栄養体生殖に
よる大量増殖法であり、また、フキ属の茎頂部を用いる
ことにより無病苗の作出を可能としている。
The shoot primordia propagated by the method of the present invention are clumps of dividing cells that are genetically extremely stable, have a high multiplication rate, and easily convert into plants. That is, the present invention is a type of mass propagation method using vegetative reproduction, and also makes it possible to produce disease-free seedlings by using the shoot apex of the genus Butterbur.

〔作 用〕[For production]

本発明をざらに、詳しく説明する。 The present invention will be briefly explained in detail.

フキ属の茎頂部を含む茎を滅菌後、茎頂部を摘出しこれ
を無機塩類および植物生長ホルモン、炭素源を含む人工
培地に移植する。次いで、照明下回転培養を行い苗条原
基を誘導する。
After sterilizing the stem containing the shoot apex of the genus Butterbur, the stem apex is removed and transplanted to an artificial medium containing inorganic salts, plant growth hormones, and a carbon source. Next, rotational culture under illumination is performed to induce shoot primordia.

上記の照明下の回転培養としては照明度下限2.000
ルクス、上限io、oooルクス、温度18〜28°C
および回転数0.5〜5/分の条件があげられる。
For rotating culture under the above illumination, the lower limit of illumination intensity is 2.000.
Lux, upper limit io, ooo lux, temperature 18-28°C
and conditions of rotation speed 0.5 to 5/min.

静置培養では、苗条原基は誘導されても継持増殖するこ
とは出来ず、回転培養によって苗条原基の初期誘導を行
うことが必要である。
In static culture, shoot primordia cannot be continuously propagated even if they are induced, and it is necessary to perform initial induction of shoot primordia by rotational culture.

上記の無機塩類としては、ムラシゲ−スクーグ(Mur
ashige−skoog) 、ガンポーグ(Gamb
org)、ホワイト(White)等の組成を有する培
地を用いることが出来る。
The above-mentioned inorganic salts include Murashige-Skoog (Murashige-Skoog)
ashige-skoog), Gamboug (Gamb)
A medium having a composition such as White, White, etc. can be used.

上記の植物生長ホルモンとしては、オーキシンとしてイ
ンドール酢酸、α−ナフタレン酢酸等、サイトカイニン
として、6−ベンジルアミノプリン、カイネチン等を用
いることが出来るが、オーキシンの少くとも1種とサイ
トカイニンの少くとも1種を併用することが望ましい。
As the above plant growth hormone, indoleacetic acid, α-naphthaleneacetic acid, etc. can be used as auxin, and 6-benzylaminopurine, kinetin, etc. can be used as cytokinin, but at least one type of auxin and at least one type of cytokinin can be used. It is desirable to use them together.

この初期誘導時の人工培地としては例えばサイトカイニ
ンとして6−ベンジルアミノプリン1〜3ppm、オー
キシンとしてα−ナフタレン酢酸0.2〜2.0ppm
および蔗糖1〜5%を含むものが用いられる。
The artificial medium for this initial induction may contain, for example, 1 to 3 ppm of 6-benzylaminopurine as cytokinin and 0.2 to 2.0 ppm of α-naphthaleneacetic acid as auxin.
and those containing 1 to 5% sucrose are used.

誘導した苗条原基は、最初淡赤色の小突起の多い形態を
している。
The induced shoot primordia initially has a pale red shape with many small protrusions.

初期誘導後約1カ月経過した時点で初期誘導時の人工培
地とは植物生長ホルモンの種類および/または濃度の異
った無機塩類および植物生長ホルモンを含む新たな別個
の人工培地に初期誘導した苗条原基を移植して継代培養
を行って継持増殖させる。この継持増殖時の人工培地は
初期誘導時の人工培地に比し植物生長ホルモンの種類お
よび/または濃度を変更することが必要であり、これを
変更しないと形態がかわりカルス化する。この植物生長
ホルモンの種類および/または濃度を変更する手段のう
ち、サイトカイニンの濃度は不変としオーキシンのみそ
の濃度を低減して両者を併用するものが最も有効である
。この継持増殖によって苗条原基は次第に大きくなり、
淡緑色の集塊になる。適当な大きざ(約5M〜10.)
に増殖した時点で植物生長ホルモンのみを変更した同一
培地に移植することにより、以前と同様に苗条原基が増
殖する。以後、定期的に新しい培地に分割移植すること
により、半永久的に維持増殖することが出来る。
Approximately 1 month after the initial induction, the shoots were initially induced on a new, separate artificial medium containing inorganic salts and plant growth hormones with different types and/or concentrations of plant growth hormones. The primordium is transplanted and subcultured for continuous proliferation. It is necessary to change the type and/or concentration of the plant growth hormone in the artificial medium used for this continuous propagation compared to the artificial medium used for initial induction; if this is not changed, the morphology will change and callus will form. Among the means for changing the type and/or concentration of plant growth hormones, the most effective is to use both in combination, keeping the concentration of cytokinin unchanged and reducing the concentration of only auxin. Through this continuous multiplication, the shoot primordium gradually becomes larger,
It becomes a pale green agglomerate. Appropriate size (approximately 5M~10.)
When the shoot primordium grows, it is transplanted to the same medium with only the plant growth hormone changed, and the shoot primordium grows as before. Thereafter, by periodically dividing and transplanting into new medium, it is possible to maintain and proliferate semi-permanently.

継代培養時の人工培地としては例えばサイトカイニンと
して6−ベンジルアミノプリン1〜3ppm 、オーキ
シンとしてα−ナフタレン酢酸0.02〜0.2ppm
および1〜5%蔗糖を含むものが用いられる。
As an artificial medium during subculture, for example, 1 to 3 ppm of 6-benzylaminopurine as cytokinin and 0.02 to 0.2 ppm of α-naphthaleneacetic acid as auxin.
and those containing 1 to 5% sucrose are used.

この様にして得られた苗条原基は大量増殖させたのち固
定培地(静置培養)に移植すると、先端に葉原基を形成
し茎葉をもつ植物体へと分化し生長する。その後、徐々
に順化させると野外栽培出来る。得られた苗条原基およ
び再生させた植物体の染色体数は2 n=90 (X=
30>と安定している。
The shoot primordium thus obtained is grown in large quantities and then transplanted to a fixed medium (stationary culture), whereupon a leaf primordium forms at the tip and differentiates into a plant body with stems and leaves, which then grows. After that, after gradual acclimatization, it can be grown outdoors. The number of chromosomes in the obtained shoot primordium and regenerated plant was 2 n=90 (X=
30> and stable.

〔実施例) フキの茎頂部を含む茎を約1 cmに切り塩化ベンザル
コニウム溶液0.1%に5分間更に次亜塩素酸ナトリウ
ム溶液1%に5分間浸して殺菌処理を行った後、実体顕
微鏡下で茎頂を摘出し植え付は材料とした。人工液体培
地はムラシゲ−スクーグ培地を用い蔗糖3%と植物生長
ホルモンとして、6−ベンジルアミノプリン、α−ナフ
タレン酢酸をそれぞれ0,0.02.0.2又は2pp
m I7)8度になるように添加して調製した。
[Example] After cutting the stem including the top of the butterbur into approximately 1 cm pieces and sterilizing them by soaking them in a 0.1% benzalkonium chloride solution for 5 minutes and then in a 1% sodium hypochlorite solution for 5 minutes, The shoot tips were removed under a stereomicroscope and used as material for planting. The artificial liquid medium was Murashige-Skoog medium, containing 3% sucrose and plant growth hormones such as 6-benzylaminopurine and α-naphthalene acetic acid at 0, 0.02, 0.2, or 2 pp, respectively.
m I7) It was prepared by adding it to a temperature of 8 degrees.

培地のpHは5.7〜5.8に調整した。The pH of the medium was adjusted to 5.7-5.8.

それぞれの培地を試験管(27x 200 mm >に
25成分注し、次いで高圧滅菌器で121℃、15分間
滅菌した。調製した培地に無菌的に茎頂を植え付け、照
明24時間(照度下限2,000〜上限10、000ル
クス)、温度22(±2)°Cの条件下に回転培養(1
分間2回転)を行った。約1カ月経過すると6−ベンジ
ルアミノプリン2t)l)m、α−ナフタレン酢112
ppmの組合せにおいて、小突起の多い淡赤色の苗条原
基が得られた。他の組合せにおいてはカルスおよび早生
分枝状態となった。この時点で6−ベンジルアミノプリ
ン2ppm 、α−ナフタレン酢酸0. O2ppmに
植物生長ホルモン濃度を変更した培地に移植することに
より、苗条原基を維持増殖させることか出来た。
25 components of each medium were poured into test tubes (27 x 200 mm), and then sterilized in an autoclave at 121°C for 15 minutes.The shoot tips were aseptically planted in the prepared medium, and exposed to light for 24 hours (minimum illuminance 2, 000 to upper limit 10,000 lux) and a temperature of 22 (±2) °C.
2 revolutions per minute). After about a month, 6-benzylaminopurine 2t)l)m, α-naphthalene vinegar 112
In the ppm combination, pale red shoot primordia with many small protrusions were obtained. Other combinations resulted in callus and early branching. At this point, 6-benzylaminopurine 2 ppm, α-naphthalene acetic acid 0. The shoot primordia could be maintained and propagated by transplanting to a medium containing a plant growth hormone concentration of O2 ppm.

次第に生長し、小突起の多い淡緑色の苗条原基塊となっ
た。以後、定期的に分割移植することにより増殖をつづ
けた。
It gradually grew and became a pale green shoot primordium mass with many small protrusions. From then on, it continued to grow by dividing and transplanting periodically.

苗条原基から植物体への転換は固定培地で静置培養によ
り行った。
Conversion from shoot primordia to plants was performed by static culture on a fixed medium.

固定培地としては、無機塩類組成1/2希釈ムラシゲ−
スクーグに、蔗糖1%、寒天0.9%を加え、pl−1
5,7〜5.8に調製したものを用いた。
As a fixed medium, Murashige diluted with an inorganic salt composition of 1/2
Add 1% sucrose and 0.9% agar to Skoog, pl-1
5.7 to 5.8 was used.

この時の培養条件は光16時間1〜2千ルクス照明、温
度22(±2)°Cでおった。
The culture conditions at this time were 1 to 2,000 lux illumination for 16 hours and a temperature of 22 (±2)°C.

調製した培地に苗条原基を植え付けると約1週間増殖を
続けた後、先端に緑色の葉原基を形成し次第に茎葉を持
つ小植物体へ生長した。約30〜45日月には発根が見
られロックウール上で順化した後、野外栽培へ移した。
When the shoot primordium was planted in the prepared medium, it continued to proliferate for about one week, and then a green leaf primordium was formed at the tip and gradually grew into a plantlet with stems and leaves. Rooting was observed in about 30 to 45 days, and after acclimatization on rock wool, the plants were transferred to outdoor cultivation.

この植物体の染色体は親植物つまり茎頂を摘出した株と
同数の2 n=90 (X=30)であり形態的にも変
異は見られなかった。
This plant had the same number of chromosomes as the parent plant, ie, the strain from which the shoot apex was removed, 2 n = 90 (X = 30), and no variation was observed in morphology.

苗条原基の増殖率は約1カ月間に重量比で約7〜8倍に
なる。また、苗条原基からの植物体への転換は苗条原基
17から約500本の植物体が得られた。つまり、1つ
の茎頂から1年間で2手刀株以上の苗が1けられる。ま
た、既知の方法で検査した結果ウィルスは検出されず無
病苗であった。
The growth rate of shoot primordia increases by about 7 to 8 times by weight in about one month. Regarding the conversion of shoot primordia to plants, approximately 500 plants were obtained from shoot primordium 17. In other words, one seedling of two or more plants can be produced from one stem apex in one year. In addition, as a result of testing using a known method, no virus was detected and the seedlings were disease-free.

(発明の効果) 本発明の方法を用いれば不稔で株分は以外の増殖法がな
いフキ属においても苗条原基を誘導維持、増殖させその
後、植物体へ転換することにより、均一な苗を常に安定
して供給出来る。
(Effect of the invention) By using the method of the present invention, even in the genus Butterbur, which is sterile and for which there is no other method of propagating plants, the shoot primordia can be maintained and propagated, and then converted into plants, resulting in uniform seedlings. can always be stably supplied.

また、短期間に大量増殖できることによりコストか軽減
出来る。
In addition, costs can be reduced by being able to multiply in large numbers in a short period of time.

更に、無病苗による収量の増加、品質の均−化等が可能
となり、効果は絶大でおる。
Furthermore, the disease-free seedlings make it possible to increase yields, equalize quality, etc., and the effects are tremendous.

手続補正書泊発) 昭和63年4月26日 特許庁長官 、/JX月1$i’3大 殿1、事件の表
示 昭和63年 特許願 第22421号 2、発明の名称 フキ属の茎頂培養による生産方法 3、補正をする者 事件との関係 特許出願人 4、代理人 住 所   東京都千代田区神田北乗物町16番地5、
補正の対象 明細書の発明の詳細な説明の欄 6、補正の内容 明細書第8頁11〜12行目に 「苗条原基塊」とあるを「苗条原基集塊」と訂正。
April 26, 1985 Commissioner of the Japan Patent Office / JX Month 1 $i'3 Daido 1, Indication of the case 1988 Patent Application No. 22421 2, Name of the invention Stem tip of the genus Butterbur Production method by culture 3, relationship with the case of the person making the amendment Patent applicant 4, agent address: 16-5, Kanda Kita Jorimono-cho, Chiyoda-ku, Tokyo;
In Column 6 of the detailed description of the invention in the specification subject to amendment, on page 8, lines 11-12 of the description of the contents of the amendment, the term "shoot primordium mass" has been corrected to "shoot primordium mass".

Claims (3)

【特許請求の範囲】[Claims] (1)フキ属の茎頂部を摘出しこれを無機塩類および植
物生長ホルモンを含む人工培地に移植して照明下に回転
培養することにより苗条原基の初期誘導を行い、然る後
該苗条原基を前記の人工培地とは植物生長ホルモンの種
類および/または濃度の異つた人工培地に移植して継代
培養することにより継持増殖させることを特徴とするフ
キ属の茎頂培養による生産方法。
(1) Initial induction of shoot primordia is performed by removing the shoot apex of the genus Butterbur and transplanting it to an artificial medium containing inorganic salts and plant growth hormones and rotating culture under illumination. A production method by shoot apical culture of the genus Butterbur, characterized in that the base is transplanted to an artificial medium containing a different type and/or concentration of plant growth hormone from the artificial medium and subcultured for continuous propagation. .
(2)植物生長ホルモンの種類および/または濃度のみ
を変更した同一組成の人工培地に移植して継持増殖を反
覆することを特徴とする請求項1記載の生産方法。
(2) The production method according to claim 1, characterized in that the continuous multiplication is repeated by transplanting to an artificial medium of the same composition in which only the type and/or concentration of the plant growth hormone is changed.
(3)苗条原基を初期誘導した後継持増殖させ、これを
更に固定培地に移植して植物体へ分化成長せしめること
を特徴とする請求項1記載の生産方法。
(3) The production method according to claim 1, characterized in that the shoot primordium is initially induced, sustained propagation is carried out, and the shoot primordium is further transplanted to a fixed medium to differentiate and grow into a plant body.
JP2242188A 1988-02-02 1988-02-02 Production of butterbur by shoot apex culture Granted JPH01199521A (en)

Priority Applications (1)

Application Number Priority Date Filing Date Title
JP2242188A JPH01199521A (en) 1988-02-02 1988-02-02 Production of butterbur by shoot apex culture

Applications Claiming Priority (1)

Application Number Priority Date Filing Date Title
JP2242188A JPH01199521A (en) 1988-02-02 1988-02-02 Production of butterbur by shoot apex culture

Publications (2)

Publication Number Publication Date
JPH01199521A true JPH01199521A (en) 1989-08-10
JPH0351377B2 JPH0351377B2 (en) 1991-08-06

Family

ID=12082213

Family Applications (1)

Application Number Title Priority Date Filing Date
JP2242188A Granted JPH01199521A (en) 1988-02-02 1988-02-02 Production of butterbur by shoot apex culture

Country Status (1)

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CN115918540A (en) * 2022-12-26 2023-04-07 广西壮族自治区药用植物园 Method for eliminating bacterial contamination in artemisia annua tissue culture

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Publication number Priority date Publication date Assignee Title
CN112772413A (en) * 2020-12-31 2021-05-11 向丽 Tissue culture method and culture medium composition of artemisia annua
CN115918540A (en) * 2022-12-26 2023-04-07 广西壮族自治区药用植物园 Method for eliminating bacterial contamination in artemisia annua tissue culture
CN115918540B (en) * 2022-12-26 2024-01-12 广西壮族自治区药用植物园 Elimination of Artemisia annua L tissue culture contamination with medium bacteria is a method of (2)

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