JPH10174586A - Tyrosinase gene of shiitake (lentinus edodes) - Google Patents
Tyrosinase gene of shiitake (lentinus edodes)Info
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- JPH10174586A JPH10174586A JP8337127A JP33712796A JPH10174586A JP H10174586 A JPH10174586 A JP H10174586A JP 8337127 A JP8337127 A JP 8337127A JP 33712796 A JP33712796 A JP 33712796A JP H10174586 A JPH10174586 A JP H10174586A
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Abstract
Description
【０００１】[0001]
【発明の属する技術分野】本発明は、シイタケチロシナ
ーゼをコードするＤＮＡに関する。TECHNICAL FIELD The present invention relates to a DNA encoding shiitake tyrosinase.
【０００２】[0002]
【従来の技術】チロシナーゼとは、モノフェノール化合
物、ポリフェノール化合物などを酸化する酵素で、広く
生物が含有する酵素である。チロシナーゼは生体内でメ
ラニン生成に関与し、種々の生理作用を示す重要な酵素
である。また、チロシナーゼは種々の産業に利用されて
いる。食品工業ではカカオやコーヒー豆の酵素的褐変化
にカビ産生のチロシナーゼが用いられており（特開昭54
-50586号公報）、また、医薬品工業においてはメラニン
の製造にチロシナーゼが用いられている（特開昭53-382
90号公報）。2. Description of the Related Art Tyrosinase is an enzyme that oxidizes monophenolic compounds, polyphenolic compounds, and the like, and is widely contained in living organisms. Tyrosinase is an important enzyme that participates in melanin production in vivo and exhibits various physiological actions. Tyrosinase is used in various industries. In the food industry, mold-producing tyrosinase is used for enzymatic browning of cacao and coffee beans.
No. -50586), and in the pharmaceutical industry, tyrosinase is used for the production of melanin (JP-A-53-382).
No. 90).
【０００３】従来より、チロシナーゼは各種生体から精
製されており、例えばマッシュルーム（T. Nakamura et
al., J. Biochemistry, 59, 481-486（1966））、アカ
パンカビ（M. Fling et al., J. Biol. Chem., 238, 20
45-2053（1963））などのほか、リンゴ（M. Murata et
al., Biosci. Biotech. Biochem., 56, 1705-1710（199
2））、モモ（T. C. Wong et al., Plant Physiology,
48, 19-23（1971））等の果樹、ジャガイモ（S.U. Path
ak et al., Phytochemistry, 36, 1165-1167（1994））
等の野菜等およびチャ（R. P. F. Gregory et al., Bio
chem. J., 101, 569-581（1966））等から得られてい
る。[0003] Conventionally, tyrosinase has been purified from various organisms, for example, mushrooms (T. Nakamura et al.).
al., J. Biochemistry, 59, 481-486 (1966)), Neurospora crassa (M. Fling et al., J. Biol. Chem., 238, 20).
45-2053 (1963)) and apples (M. Murata et
al., Biosci. Biotech. Biochem., 56, 1705-1710 (199
2)), peach (TC Wong et al., Plant Physiology,
48, 19-23 (1971), etc., potatoes (SU Path
ak et al., Phytochemistry, 36, 1165-1167 (1994))
Such as vegetables and tea (RPF Gregory et al., Bio
chem. J., 101, 569-581 (1966)).
【０００４】しかし、チロシナーゼを大量かつ効率よく
得るには遺伝子工学的手法によるチロシナーゼの製造が
望まれており、そのためには、チロシナーゼをコードす
るＤＮＡ（チロシナーゼ遺伝子ともいう）をクローニン
グすることが必要とされる。これまでに、チロシナーゼ
遺伝子の塩基配列は若干報告されている。例えば、麹菌
（Aspergillus oryzae；Y. Fujita et al., Biochemica
et Biophysica Acta, 1261, 151-154（1995））、アカ
パンカビ（Neurospora crassa；U. Kupper et al., J.
Biol. Chem., 264, 17250-17258（1989））、トマト
（T. Shahar etal., Plant Cell, 4, 135-147（199
2））などのチロシナーゼ遺伝子について塩基配列が示
されているが、シイタケのチロシナーゼ（以下「シイタ
ケチロシナーゼ」という）については塩基配列が明らか
ではない。However, in order to obtain tyrosinase in large quantities and efficiently, production of tyrosinase by a genetic engineering technique is desired. For that purpose, it is necessary to clone a DNA encoding tyrosinase (also referred to as tyrosinase gene). Is done. The nucleotide sequence of the tyrosinase gene has been reported to some extent. For example, Aspergillus oryzae; Y. Fujita et al., Biochemica
et Biophysica Acta, 1261, 151-154 (1995)), Neurospora crassa; U. Kupper et al., J.
Biol. Chem., 264, 17250-17258 (1989)), tomato (T. Shahar et al., Plant Cell, 4, 135-147 (199)
Although the nucleotide sequence of the tyrosinase gene such as 2)) is shown, the nucleotide sequence of shiitake tyrosinase (hereinafter referred to as “shiitake tyrosinase”) is not clear.
【０００５】[0005]
【発明が解決しようとする課題】本発明は、シイタケチ
ロシナーゼをコードするＤＮＡを提供することを目的と
する。An object of the present invention is to provide a DNA encoding shiitake tyrosinase.
【０００６】[0006]
【課題を解決するための手段】本発明者等は、上記課題
を解決すべく鋭意研究した結果、シイタケ（Lentinused
odes）由来のｃＤＮＡからチロシナーゼ遺伝子をクロー
ニングすることに成功し、本発明を完成するに至った。
すなわち、本発明は、配列番号１で表されるアミノ酸配
列、又は該アミノ酸配列において１若しくは複数のアミ
ノ酸が欠失、置換若しくは付加されたものを含み、且つ
チロシナーゼ活性をもたらすタンパク質をコードするＤ
ＮＡである。該ＤＮＡとしては、シイタケの子実体又は
菌糸由来のものが挙げられ、配列番号２又は３で表され
る塩基配列を含むものが挙げられる。Means for Solving the Problems The inventors of the present invention have conducted intensive studies to solve the above-mentioned problems, and as a result, have found that shiitake mushrooms (Lentinused)
odes) was successfully cloned from the tyrosinase gene, and the present invention was completed.
That is, the present invention relates to a D-encoding protein comprising the amino acid sequence represented by SEQ ID NO: 1 or a protein having one or more amino acids deleted, substituted or added in the amino acid sequence, and encoding tyrosinase activity.
NA. Examples of the DNA include those derived from shiitake mushroom fruit bodies or hyphae, and include those containing the nucleotide sequence represented by SEQ ID NO: 2 or 3.
【０００７】ここで、「１若しくは複数のアミノ酸が欠
失、置換若しくは付加された」とは、配列番号１で表さ
れるアミノ酸配列を含むタンパク質がチロシナーゼ活性
を有する限り、そのアミノ酸配列に１個又は複数の欠
失、置換、付加、挿入等の変異が生じてもよいことを意
味する。[0007] Here, "one or more amino acids are deleted, substituted or added" means that as long as the protein containing the amino acid sequence represented by SEQ ID NO: 1 has tyrosinase activity, one amino acid sequence is added to the protein. Alternatively, it means that a plurality of mutations such as deletion, substitution, addition, and insertion may occur.
【０００８】したがって、例えばシイタケチロシナーゼ
のアミノ酸配列に含まれる第１番目のメチオニン（配列
番号１で表されるアミノ酸配列中の第１番目の Met）が
欠失しているものなども、このアミノ酸配列の変化によ
るチロシナーゼに含まれる。また、シイタケチロシナー
ゼをコードする塩基配列のほか、縮重コドンにおいての
み異なる同一のタンパク質をコードする縮重異性体も本
発明のＤＮＡに含まれる。以下、本発明を詳細に説明す
る。Accordingly, for example, the amino acid sequence of Shiitake tyrosinase in which the first methionine (the first Met in the amino acid sequence represented by SEQ ID NO: 1) contained in the amino acid sequence is deleted, Is included in tyrosinase due to the change of In addition to the nucleotide sequence encoding shiitake tyrosinase, degenerate isomers encoding the same protein that differs only in degenerate codons are included in the DNA of the present invention. Hereinafter, the present invention will be described in detail.
【０００９】[0009]
【発明の実施の形態】本発明のＤＮＡは、下記の工程に
よりクローニングされる。 (1) ｍＲＮＡの調製およびｃＤＮＡの作製 まず、レンチナス属に属するシイタケ、例えばレンチナ
ス・エドデス（Lentinus edodes）からｍＲＮＡを抽出
する。ｍＲＮＡの抽出は、公知の方法、例えばオリゴ
（dT）セルロースカラム法、マグネタイトオリゴ（dT）
パーティクル法、あるいは Novagen社の Straight A's
TM mRNA Isolation Kit を用いて行うことができる。ｍ
ＲＮＡの抽出に使用する組織としては、例えば子実体、
菌糸等が挙げられる。BEST MODE FOR CARRYING OUT THE INVENTION The DNA of the present invention is cloned by the following steps. (1) Preparation of mRNA and Preparation of cDNA First, mRNA is extracted from Lentinus edodes belonging to the genus Lentinas, for example, Lentinus edodes. mRNA can be extracted by known methods, for example, oligo (dT) cellulose column method, magnetite oligo (dT)
Particle method or Novagen's Straight A's
This can be performed using the TM mRNA Isolation Kit. m
Tissues used for RNA extraction include, for example, fruiting bodies,
Hyphae and the like.
【００１０】次に、これらの組織を凍結し、液体窒素存
在下で磨砕し、ＲＮＡ抽出液を加えて全ＲＮＡの粗抽出
物を得る。さらに、タンパク質、多糖類、その他の不純
物を除去し、マグネタイトオリゴ（dT）パーティクルを
用いて更に精製する。ポリ A（ポリ A＋）鎖画分を溶出
し、溶出液を集めた後、同様の精製を２〜３回繰り返す
ことによってｍＲＮＡを高度に濃縮し、前記組織の全Ｒ
ＮＡを単離する。一方、シイタケチロシナーゼの部分ア
ミノ酸配列から合成した２種類の縮重オリゴヌクレオチ
ドプライマーを設計する。[0010] Next, these tissues are frozen, ground in the presence of liquid nitrogen, and an RNA extract is added to obtain a crude extract of total RNA. Furthermore, proteins, polysaccharides, and other impurities are removed, and further purified using magnetite oligo (dT) particles. The poly A (poly A +) chain fraction was eluted, the eluate was collected, and the same purification was repeated two to three times to highly concentrate the mRNA, thereby obtaining the total R of the tissue.
Isolate NA. On the other hand, two types of degenerate oligonucleotide primers synthesized from the partial amino acid sequence of Shiitake tyrosinase are designed.
【００１１】前記のように精製したｍＲＮＡをテンプレ
ートにし、縮重オリゴヌクレオチドプライマーを用いて
RT-PCRを行い、得られたシイタケチロシナーゼ遺伝子の
部分塩基配列をクローニングした後、その塩基配列を解
析する。塩基配列の解析は、例えばABI PRISMTM Dye Te
rminator Cycle Sequencing Ready Reaction kit, FS
(Perkin Elmer)を用いて行うことができる。RT-PCR産物
の塩基配列を基に、改めてシイタケチロシナーゼ遺伝子
に特異的なプライマーを合成する。[0011] Using the mRNA purified as described above as a template, and using a degenerate oligonucleotide primer
After performing RT-PCR and cloning the partial nucleotide sequence of the obtained Shiitake tyrosinase gene, the nucleotide sequence is analyzed. For analysis of the nucleotide sequence, for example, ABI PRISM ™ Dye Te
rminator Cycle Sequencing Ready Reaction kit, FS
(Perkin Elmer). Based on the base sequence of the RT-PCR product, a primer specific to the shiitake tyrosinase gene is synthesized again.
【００１２】上記プライマーを用いてRACE法によりシイ
タケチロシナーゼ遺伝子の 5' 末端側および 3' 末端側
の部分塩基配列を増幅する。RACE法は、例えば 5' RACE
System for Rapid Amplification of cDNA Endsおよび
3' RACE System for RapidAmplification of cDNA End
s（GibcoBRL）を用いて行うことができる。Using the primers described above, the partial base sequences at the 5 'end and the 3' end of the shiitake tyrosinase gene are amplified by the RACE method. The RACE method is, for example, 5 'RACE
System for Rapid Amplification of cDNA Ends and
3 'RACE System for RapidAmplification of cDNA End
s (GibcoBRL).
【００１３】RACE法によって増幅されたシイタケチロシ
ナーゼ遺伝子の部分塩基配列をプラスミドベクターに組
み込んだ後、トランスフォーメーションすることによっ
てクローニングする。RACE産物のクローニングは、例え
ば Original TA Cloning Kit（Invitrogen）を用いて行
うことができる。[0013] The partial base sequence of the shiitake tyrosinase gene amplified by the RACE method is inserted into a plasmid vector, and then cloned by transformation. Cloning of the RACE product can be performed, for example, using the Original TA Cloning Kit (Invitrogen).
【００１４】(2) プローブＤＮＡの作製 シイタケから単離精製されたチロシナーゼの部分アミノ
酸配列から２種類の縮重オリゴヌクレオチドプライマー
（N プライマーおよび Cプライマー）を合成する。(2) Preparation of Probe DNA Two kinds of degenerate oligonucleotide primers (N primer and C primer) are synthesized from the partial amino acid sequence of tyrosinase isolated and purified from Shiitake mushroom.
【００１５】N プライマー：5'-T(C/T)CA(A/G)AT(A/C/
T)GG(A/C/G/T)GG(A/C/G/T)AT(A/C/T)CA(C/T)GG -3'
（配列番号４） C プライマー：5'-(A/G)(A/C/G/T)C(G/T)(A/G)TC(A/C/G
/T)AC(C/T)TG(A/C/G/T)GC(A/G)TG(A/G)TG-3'（配列番号
５）N primer: 5'-T (C / T) CA (A / G) AT (A / C /
T) GG (A / C / G / T) GG (A / C / G / T) AT (A / C / T) CA (C / T) GG -3 '
(SEQ ID NO: 4) C primer: 5 '-(A / G) (A / C / G / T) C (G / T) (A / G) TC (A / C / G
/ T) AC (C / T) TG (A / C / G / T) GC (A / G) TG (A / G) TG-3 '(SEQ ID NO: 5)
【００１６】この２種類のオリゴヌクレオチドプライマ
ーを用いてRT-PCRを行い、得られるＤＮＡ断片を精製し
た後、クローニング及びシークエンシングを行う。得ら
れた塩基配列を基に改めて２種類のプライマー（N>GL.S
Q01 プライマーおよび GL<C.SQ01プライマー）を合成す
る。[0016] RT-PCR is performed using these two kinds of oligonucleotide primers, and the obtained DNA fragment is purified, followed by cloning and sequencing. Two types of primers (N> GL.S.
Synthesize Q01 primer and GL <C.SQ01 primer).
【００１７】N>GL.SQ01 プライマー：5'-GCGCAGGAAATAA
GCCAGTAGACAC-3'(配列番号６） GL<C.SQ01 プライマー：5'-GCGTGGTGCATAAAGAAAAT-3'
（配列番号７） 上記２種類のプライマーを用いて得られる650 bpのRT-P
CR産物を精製した後、クローニングする。クローニング
は、例えばOriginal TA Cloning kit (Invitrogen)を用
いて行うことができる。得られたクローンからプラスミ
ド調製した後、Eco RI消化によってインサート（RT-PCR
産物）を回収し、プローブとして用いる。N> GL.SQ01 Primer: 5'-GCGCAGGAAATAA
GCCAGTAGACAC-3 '(SEQ ID NO: 6) GL <C.SQ01 Primer: 5'-GCGTGGTGCATAAAGAAAAT-3'
(SEQ ID NO: 7) 650 bp RT-P obtained using the above two primers
After purifying the CR product, it is cloned. Cloning can be performed using, for example, Original TA Cloning kit (Invitrogen). After plasmid preparation from the obtained clone, insert (RT-PCR
Product) and use it as a probe.
【００１８】(3) RACE産物からのスクリーニング クローニングしたRACE産物から、チロシナーゼ遺伝子の
部分塩基配列を有するクローンをスクリーニングする。
スクリーニングは、例えば上記プローブを用いたコロニ
ーハイブリダイゼーション法、あるいは上記２種類のプ
ライマーを用いたＰＣＲ法によって行われる。(3) Screening from RACE product A clone having a partial nucleotide sequence of the tyrosinase gene is screened from the cloned RACE product.
The screening is performed by, for example, a colony hybridization method using the above-mentioned probe or a PCR method using the above-mentioned two kinds of primers.
【００１９】(4) RACE産物のシークエンシング RACE産物の塩基配列については、(3) で得られたクロー
ンからプラスミドを調製し、市販のキット（ABI PRISM
TM Dye Terminator Cycle Sequencing Ready Reaction
Kit, FS (PERKIN ELMER)）を用いたいわゆる「Primer W
alking 法」によって決定することができる。(4) Sequencing of the RACE product For the nucleotide sequence of the RACE product, a plasmid was prepared from the clone obtained in (3) and a commercially available kit (ABI PRISM
TM Dye Terminator Cycle Sequencing Ready Reaction
Kit, FS (PERKIN ELMER))
alking method ".
【００２０】配列番号３に本発明のＤＮＡを例示する
が、本質的にチロシナーゼ活性を発現する配列であれば
この配列に限定されるものではなく、欠失、置換、挿
入、付加などによってその塩基配列を変異させることが
可能である。SEQ ID NO: 3 exemplifies the DNA of the present invention, but is not limited to this sequence as long as it essentially expresses tyrosinase activity, and its base may be deleted, substituted, inserted, added or the like. It is possible to mutate the sequence.
【００２１】シイタケチロシナーゼ遺伝子の塩基配列が
一旦決定されると、その後は、その配列をもとに互いに
３’末端に10〜12bpの相補鎖又はオーバーラップを有す
る80-100mer のオリゴヌクレオチドプライマーを化学合
成し、この合成オリゴヌクレオチドプライマーのみを使
用したＰＣＲ（藤本英也，細胞工学別冊植物細胞工学シ
リーズ２，植物のＰＣＲ実験プロトコール，p84-89, 19
95) によって、あるいは該塩基配列を有するＤＮＡ断片
をプローブとしてハイブリダイズさせることにより、本
発明の遺伝子を得ることができる。なお、塩基配列の変
異は、公知の突然変異導入法（例えば In vitro Mutage
nesis primers 法（TAKARA shuzou Co., LTD.））を用
いて決定することができる。Once the nucleotide sequence of the shiitake tyrosinase gene has been determined, an 80-100 mer oligonucleotide primer having a 10-12 bp complementary strand or overlap at the 3 'end with each other is then chemically based on the sequence. PCR using only synthetic oligonucleotide primers (Hideya Fujimoto, Cell Engineering Separate Volume, Plant Cell Engineering Series 2, Plant PCR Experiment Protocol, p84-89, 19
95) or by hybridizing using a DNA fragment having the nucleotide sequence as a probe, the gene of the present invention can be obtained. The mutation of the nucleotide sequence can be performed by a known mutagenesis method (eg, in vitro mutagenesis).
nesis primers method (TAKARA shuzou Co., LTD.).
【００２２】[0022]
【実施例】以下、実施例により本発明をさらに具体的に
説明する。ただし、本発明はこれら実施例によりその技
術的範囲を限定するものではない。The present invention will be described more specifically with reference to the following examples. However, the present invention does not limit the technical scope by these examples.
【００２３】〔実施例１〕 (1) シイタケ子実体採取 シイタケ菌株として北研57号（（株）北研）を用いた。
子実体形成は松本ら（T. Matsumoto et al., Rept. Tot
tori Mycol. Inst., 26, 46-54（1988））の方法に準
じて行い、菌傘底部の被膜が切れ始めた状態の子実体を
収穫し、25℃で３日間保存した褐変開始前の菌褶部から
総 ＲＮＡの抽出を行った。[Example 1] (1) Collection of shiitake mushroom fruit body Kitaken No. 57 (Kitaken Co., Ltd.) was used as a shiitake mushroom strain.
Fruiting body formation is by Matsumoto et al. (T. Matsumoto et al., Rept. Tot
Tori Mycol. Inst., 26, 46-54 (1988)), harvest the fruiting body in which the coating on the bottom of the fungus began to be cut off and stored at 25 ° C for 3 days before browning. Total RNA was extracted from fungal folds.
【００２４】(2) 総ＲＮＡの抽出 試料から総ＲＮＡの抽出はISOGEN（Nippon Gene 社）の
プロトコールに準じて行った。その結果、試料１g から
精製された総ＲＮＡ 0.5mgを得た。 (3) ｍＲＮＡの調製 ＲＮＡ 0.5mgに対して、Straight A'sTM mRNA Isolatio
n Kit （Novagen 社）を利用し、Anneal MagnetightTM
Oligo（dT）Particleを用いたアフィニティー精製法を
行うことによりｍＲＮＡを精製した。最終的に27μg の
ｍＲＮＡが精製された。(2) Extraction of Total RNA Extraction of total RNA from the sample was performed according to the protocol of ISOGEN (Nippon Gene). As a result, 0.5 mg of purified total RNA was obtained from 1 g of the sample. (3) Preparation of mRNA For 0.5 mg of RNA, Straight A's TM mRNA Isolation
using the n Kit (Novagen, Inc.), Anneal Magnetight TM
MRNA was purified by performing an affinity purification method using Oligo (dT) Particles. Finally, 27 μg of mRNA was purified.
【００２５】(4) ｃＤＮＡの調製 ｍＲＮＡからのｃＤＮＡの調製には、逆転写酵素とし
て、SuperScript IITM RNaseH- Reverse Transcriptase
(GibcoBRL) を用いた。反応は常法にしたがって42℃で6
0分反応し、95℃で10分加熱処理して反応を停止した。
反応液からｃＤＮＡを常法にて精製単離した。(4) Preparation of cDNA For preparation of cDNA from mRNA, SuperScript II ™ RNaseH - Reverse Transcriptase was used as a reverse transcriptase.
(GibcoBRL) was used. The reaction is carried out at 42 ° C
The reaction was performed for 0 minutes, and the reaction was stopped by heating at 95 ° C. for 10 minutes.
CDNA was purified and isolated from the reaction solution by a conventional method.
【００２６】(5) シイタケチロシナーゼ遺伝子部分塩基
配列の調製 シイタケチロシナーゼの部分アミノ酸配列（K. Kanda e
t al., Biosci. Biotech. Biochem., 60, 1273-1278
（1996））から２種類の縮重オリゴヌクレオチドプライ
マー（N プライマーおよび Cプライマー）を合成した。(5) Preparation of partial nucleotide sequence of Shiitake tyrosinase gene Partial amino acid sequence of Shiitake tyrosinase (K. Kanda e
t al., Biosci. Biotech. Biochem., 60, 1273-1278
(1996)), two types of degenerate oligonucleotide primers (N primer and C primer) were synthesized.
【００２７】N プライマー：5'- T(C/T)CA(A/G)AT(A/C/
T)GG(A/C/G/T)GG(A/C/G/T)AT(A/C/T)CA(C/T)GG-3'(配列
番号４) C プライマー：5'-(A/G)(A/C/G/T)C(G/T)(A/G)TC(A/C/G
/T)AC(C/T)TG(A/C/G/T)GC(A/G)TG(A/G)TG-3 '(配列番号
５)N primer: 5'-T (C / T) CA (A / G) AT (A / C /
T) GG (A / C / G / T) GG (A / C / G / T) AT (A / C / T) CA (C / T) GG-3 '(SEQ ID NO: 4) C primer: 5' -(A / G) (A / C / G / T) C (G / T) (A / G) TC (A / C / G
/ T) AC (C / T) TG (A / C / G / T) GC (A / G) TG (A / G) TG-3 '(SEQ ID NO: 5)
【００２８】これら縮重オリゴヌクレオチドプライマー
を用いて、上記ｃＤＮＡをテンプレートにして PCRを行
った。反応は、94℃で１分、50℃で１分、72℃で２分の
反応を１サイクルとしてこれを30サイクル行った。得ら
れた700 bpのRT-PCR産物は常法にしたがって精製した
後、クローニングを行った。Using these degenerate oligonucleotide primers, PCR was performed using the above cDNA as a template. The reaction was carried out 30 cycles, with the reaction at 94 ° C. for 1 minute, 50 ° C. for 1 minute and 72 ° C. for 2 minutes as one cycle. The obtained 700 bp RT-PCR product was purified according to a conventional method, and then cloned.
【００２９】(6) RT-PCR産物のシークエンシング RT-PCR産物の塩基配列については、上記(5) で得られた
クローンから RPM Kit（Bio101）でプラスミドを調製
し、このプラスミドをテンプレートとして ABI PRISMTM
Dye Terminator Cycle Sequencing Ready Reaction Ki
t, FS （Perkin Elmer）を用いた Primer Walking 法に
よってシークエンシングを行い、373A DNASequencer（A
pplied Biosystems）を利用して塩基配列を決定した。
この塩基配列から改めてシイタケチロシナーゼ遺伝子に
特異的な２種類のプライマー（N>GL.SQ01 プライマーお
よび GL<C.SQ01プライマー）を合成した。(6) Sequencing of RT-PCR product For the nucleotide sequence of the RT-PCR product, a plasmid was prepared from the clone obtained in the above (5) using RPM Kit (Bio101), and this plasmid was used as a template for ABI. PRISM TM
Dye Terminator Cycle Sequencing Ready Reaction Ki
Sequencing was performed by the Primer Walking method using t, FS (Perkin Elmer), and the 373A DNASequencer (A
Nucleotide sequence was determined using pplied Biosystems).
Two types of primers (N> GL.SQ01 primer and GL <C.SQ01 primer) specific to the shiitake tyrosinase gene were synthesized again from this nucleotide sequence.
【００３０】N>GL.SQ01 プライマー：5'-GCGCAGGAAATAA
GCCAGTAGACAC-3'(配列番号６) GL<C.SQ01 プライマー：5'-GCGTGGTGCATAAAGAAAAT-3'
（配列番号７)N> GL.SQ01 Primer: 5'-GCGCAGGAAATAA
GCCAGTAGACAC-3 '(SEQ ID NO: 6) GL <C.SQ01 Primer: 5'-GCGTGGTGCATAAAGAAAAT-3'
(SEQ ID NO: 7)
【００３１】(7) RACE法によるシイタケチロシナーゼ遺
伝子の 5' 末端側の塩基配列の調製GL<C.SQ01 プライマ
ーを用いて、5' RACE System for Rapid Amplification
of cDNA Ends（GibcoBRL）を利用してシイタケチロシ
ナーゼ遺伝子の 5' 末端側の塩基配列をキットのプロト
コールにしたがって増幅した。得られた 5' RACE産物は
常法にしたがって精製した後、クローニングを行った。(7) Preparation of base sequence at 5 'end of shiitake tyrosinase gene by RACE method 5' RACE System for Rapid Amplification using GL <C.SQ01 primer
Using the cDNA of Ends (GibcoBRL), the 5′-terminal nucleotide sequence of the shiitake tyrosinase gene was amplified according to the protocol of the kit. The obtained 5 'RACE product was purified according to a conventional method and then cloned.
【００３２】(8) RACE法によるシイタケチロシナーゼ遺
伝子の 3' 末端側の塩基配列の調製N>GL.SQ01 プライマ
ーを用いて、3' RACE System for Rapid Amplification
of cDNA Ends（GibcoBRL）を利用してシイタケチロシ
ナーゼ遺伝子の 3' 末端側の塩基配列をキットのプロト
コールにしたがって増幅した。得られた 3' RACE産物は
常法にしたがって精製した後、クローニングを行った。(8) Preparation of base sequence at 3 'end of shiitake tyrosinase gene by RACE method Using N> GL.SQ01 primer, 3' RACE System for Rapid Amplification
Using the cDNA Ends (GibcoBRL), the nucleotide sequence at the 3 'end of the shiitake tyrosinase gene was amplified according to the protocol of the kit. The obtained 3 'RACE product was purified according to a conventional method and then cloned.
【００３３】(9) プローブＤＮＡの調製 前記(4) で調製したｃＤＮＡをテンプレートにし、前記
(6) で合成した２種類のプライマー（N>GL.SQ01 プライ
マーおよび GL<C.SQ01プライマー) を用いてPCR を行っ
た。反応は、94℃で30秒、60℃で30秒、72℃で１分の反
応を１サイクルとしてこれを30サイクル行った。得られ
た650 bpの PCR産物を常法にしたがって精製した後、ク
ローニングを行った。得られたクローンからプラスミド
を調製した後、Eco RI消化によってインサートを回収
し、プローブＤＮＡとした。(9) Preparation of Probe DNA Using the cDNA prepared in (4) above as a template,
PCR was performed using the two types of primers synthesized in (6) (N> GL.SQ01 primer and GL <C.SQ01 primer). The reaction was performed for 30 cycles at 94 ° C. for 30 seconds, 60 ° C. for 30 seconds, and 72 ° C. for 1 minute. The obtained 650 bp PCR product was purified according to a conventional method, and then cloned. After preparing a plasmid from the resulting clone, the insert was recovered by digestion with Eco RI and used as probe DNA.
【００３４】(10) シイタケチロシナーゼ遺伝子の部分
塩基配列を有するRACE産物のスクリーニング 前記(7) および(8) で作製したRACE産物を有するクロー
ンから、前記(9) で調製したプローブを用い、コロニー
ハイブリダイゼーション法でシイタケチロシナーゼ遺伝
子の部分塩基配列を有するクローンをスクリーニングし
た。(10) Screening of RACE Product Having Partial Nucleotide Sequence of Shiitake Tyrosinase Gene From the clone having the RACE product prepared in (7) and (8) above, colony hybridization was performed using the probe prepared in (9) above. A clone having a partial base sequence of the shiitake tyrosinase gene was screened by a hybridization method.
【００３５】(11) シイタケチロシナーゼ遺伝子部分塩
基配列を有するRACE産物のシークエンシング RACE産物の塩基配列については、前記(6) の方法にした
がって解析した。5'RACE産物および3'RACE産物は、とも
に前記(9) で得られたシイタケチロシナーゼ遺伝子に特
異的な650 bpの塩基配列を有するので、この配列を基準
にしてシイタケチロシナーゼ遺伝子の全塩基配列を決定
した。決定した塩基配列を配列番号３に示す。(11) Sequencing of RACE Product Having Shiitake Tyrosinase Gene Partial Nucleotide Sequence The nucleotide sequence of the RACE product was analyzed according to the method described in (6) above. Since the 5′RACE product and the 3′RACE product both have a nucleotide sequence of 650 bp specific to the shiitake tyrosinase gene obtained in the above (9), the entire nucleotide sequence of the shiitake tyrosinase gene is determined based on this sequence. Decided. The determined nucleotide sequence is shown in SEQ ID NO: 3.
【００３６】本発明のシイタケチロシナーゼをコードす
るＤＮＡの塩基配列は、配列番号３で表される塩基配列
を含み、また、第21-23 番目の塩基配列によってコード
されるメチオニンから第1902-1904 番目のTGA で終了す
る単一のオープンリーディングフレーム（アミノ酸 627
残基）が存在していた。このオープンリーディングフレ
ームのアミノ酸配列を配列番号１に、その塩基配列を配
列番号２に記載した。なお、このオープンリーディング
フレームの上流には、20塩基の非翻訳領域が存在してい
た。The nucleotide sequence of the DNA encoding the shiitake tyrosinase of the present invention comprises the nucleotide sequence represented by SEQ ID NO: 3, and the nucleotide sequence from the methionine encoded by the nucleotide sequence at positions 21-23 to 1902-1904. Single open reading frame (amino acid 627) ending in TGA
Residue) was present. The amino acid sequence of this open reading frame is shown in SEQ ID NO: 1, and its nucleotide sequence is shown in SEQ ID NO: 2. Note that an untranslated region of 20 bases was present upstream of this open reading frame.
【００３７】さらに、オープンリーディングフレームの
下流にはポリ(A) 領域が存在しているので、このｃＤＮ
Ａは完全長のものであると言える。これらの塩基配列か
ら、シイタケから得られたチロシナーゼのアミノ酸残基
数は 627個で、その分子量は69.7kDa と推定される。Furthermore, since a poly (A) region exists downstream of the open reading frame, this cDN
A can be said to be full length. From these nucleotide sequences, it is estimated that the number of amino acid residues of tyrosinase obtained from Shiitake mushroom is 627 and its molecular weight is 69.7 kDa.
【００３８】ここに得られたシイタケチロシナーゼのア
ミノ酸配列領域を、麹菌（Aspergillus oryzae）、アカ
パンカビ（Neurospora crassa）、マッシュルーム（Aga
ricus bisporus ）やトマト(Lycopersicon esculentum)
等のチロシナーゼのアミノ酸配列領域と比較すると、
それぞれ 36.3 ％、30.8％、54.0％、19.8％までの相同
性が認められるが、かなりの部分で異なっていることか
ら、配列番号１〜３で表される塩基配列およびアミノ酸
配列は、シイタケ特有のものであることが示された。The amino acid sequence region of the shiitake tyrosinase thus obtained was subjected to Aspergillus oryzae, Neurospora crassa, and mushroom (Aga).
ricus bisporus) and tomato (Lycopersicon esculentum)
Compared to the amino acid sequence region of tyrosinase, etc.
Although homology of up to 36.3%, 30.8%, 54.0%, and 19.8% is observed, the nucleotide sequence and the amino acid sequence represented by SEQ ID NOs: 1 to 3 are unique to Shiitake mushrooms. Was shown.
【００３９】[0039]
【発明の効果】本発明により、シイタケチロシナーゼ遺
伝子が提供される。本発明の遺伝子は、同種または異種
の菌茸、微生物（細菌、酵母、かび等）、植物、動物細
胞等に導入することにより、チロシナーゼを大量生産さ
せ、代謝調節に作用させ、生理作用に影響を与えること
ができる等、幅広く活用できる点で有用である。According to the present invention, a shiitake tyrosinase gene is provided. The gene of the present invention is introduced into homologous or heterologous fungi, microorganisms (bacteria, yeasts, molds, etc.), plants, animal cells, etc., thereby producing tyrosinase in large quantities, affecting metabolic regulation, and affecting physiological functions. It is useful in that it can be used widely, for example,
【００４０】[0040]
配列番号：１ 配列の長さ：６２７ 配列の型：アミノ酸 トポロジー：直鎖状 配列の種類：タンパク質 起源 生物名：レンチナス・エドデス（Lentinus edodes） 組織の種類：子実体 配列 Met Ser His Tyr Leu Val Thr Gly Ala Thr Gly Gly Ser Thr Ser Gly 1 5 10 15 Ala Ala Ala Pro Asn Arg Leu Glu Ile Asn Asp Phe Val Lys Gln Glu 20 25 30 Asp Gln Phe Ser Leu Tyr Ile Gln Ala Leu Gln Tyr Ile Tyr Ser Ser 35 40 45 Lys Ser Gln Asp Asp Ile Asp Ser Phe Phe Gln Ile Gly Gly Ile His 50 55 60 Gly Leu Pro Tyr Val Pro Trp Asp Gly Ala Gly Asn Lys Pro Val Asp 65 70 75 80 Thr Asp Ala Trp Glu Gly Tyr Cys Thr His Gly Ser Val Leu Phe Pro 85 90 95 Thr Phe His Arg Pro Tyr Val Leu Leu Ile Glu Gln Ala Ile Gln Ala 100 105 110 Ala Ala Val Asp Ile Ala Ala Thr Tyr Ile Val Asp Arg Ala Arg Tyr 115 120 125 Gln Asp Ala Ala Leu Asn Leu Arg Gln Pro Tyr Trp Asp Trp Ala Arg 130 135 140 Asn Pro Val Pro Pro Pro Glu Val Ile Ser Leu Asp Glu Val Thr Ile 145 150 155 160 Val Asn Pro Ser Gly Glu Lys Ile Ser Val Pro Asn Pro Leu Arg Arg 165 170 175 Tyr Thr Phe His Pro Ile Asp Pro Ser Phe Pro Glu Pro Tyr Gln Ser 180 185 190 Trp Ser Thr Thr Leu Arg His Pro Leu Ser Asp Asp Ala Asn Ala Ser 195 200 205 Asp Asn Val Pro Glu Leu Lys Ala Thr Leu Arg Ser Ala Gly Pro Gln 210 215 220 Leu Lys Thr Lys Thr Tyr Asn Leu Leu Thr Arg Val His Thr Trp Pro 225 230 235 240 Ala Phe Ser Asn His Thr Pro Asp Asp Gly Gly Ser Thr Ser Asn Ser 245 250 255 Leu Glu Gly Ile His Asp Ser Val His Val Asp Val Gly Gly Asn Gly 260 265 270 Gln Met Ser Asp Pro Ser Val Ala Gly Phe Asp Pro Ile Phe Phe Met 275 280 285 His His Ala Gln Val Asp Arg Leu Leu Ser Leu Trp Ser Ala Leu Asn 290 295 300 Pro Arg Val Trp Ile Thr Asp Gly Pro Ser Gly Asp Gly Thr Trp Thr 305 310 315 320 Ile Pro Pro Asp Thr Val Val Gly Lys Asp Thr Asp Leu Thr Pro Phe 325 330 335 Trp Asn Thr Gln Ser Ser Tyr Trp Ile Ser Ala Asn Val Thr Asp Thr 340 345 350 Ser Lys Met Gly Tyr Thr Tyr Pro Glu Phe Asn Asn Leu Asp Met Gly 355 360 365 Asn Glu Val Ala Val Arg Ser Ala Ile Ala Ala Gln Val Asn Lys Leu 370 375 380 Tyr Gly Gly Pro Phe Thr Lys Phe Ala Ala Ala Ile Gln Gln Pro Ser 385 390 395 400 Ser Gln Thr Thr Ala Asp Ala Ser Thr Ile Gly Asn Val Thr Ser Asp 405 410 415 Ala Ser Ser His Leu Val Asp Ser Lys Ile Asn Pro Thr Pro Asn Arg 420 425 430 Ser Ile Asp Asp Ala Pro Gln Val Lys Ile Ala Ser Thr Leu Arg Asn 435 440 445 Asn Glu Gln Lys Glu Phe Trp Glu Trp Thr Ala Arg Val Gln Val Lys 450 455 460 Lys Tyr Glu Ile Gly Gly Ser Phe Lys Val Leu Phe Phe Leu Gly Ser 465 470 475 480 Val Pro Ser Asp Pro Lys Glu Trp Ala Thr Asp Pro His Phe Val Gly 485 490 495 Ala Phe His Gly Phe Val Asn Ser Ser Ala Glu Arg Cys Ala Asn Cys 500 505 510 Arg Arg Gln Gln Asp Val Val Leu Glu Gly Phe Val His Leu Asn Glu 515 520 525 Gly Ile Ala Asn Ile Ser Asn Leu Asn Ser Phe Asp Pro Ile Val Val 530 535 540 Glu Pro Tyr Leu Lys Glu Asn Leu His Trp Arg Val Gln Lys Val Ser 545 550 555 560 Gly Glu Val Val Asn Leu Asp Ala Ala Thr Ser Pro Gly Ser Arg Ser 565 570 575 Cys Arg Tyr Ala Phe Gly Val Ala Ser Trp Arg Asp Phe Pro Ser Thr 580 585 590 Cys Arg Asp Thr Pro Pro Ser Pro Tyr His Thr Trp Ser Ser Trp Trp 595 600 605 Phe Ser Pro Gln Arg Arg Ile Phe Lys Pro Leu Ile Arg Gln Arg Val 610 615 620 Glu Phe Glu 625 SEQ ID NO: 1 Sequence length: 627 Sequence type: Amino acid Topology: Linear Sequence type: Protein Origin Organism name: Lentinus edodes Tissue type: Fruiting body Sequence Met Ser His Tyr Leu Val Thr Gly Ala Thr Gly Gly Ser Thr Ser Gly 1 5 10 15 Ala Ala Ala Pro Asn Arg Leu Glu Ile Asn Asp Phe Val Lys Gln Glu 20 25 30 Asp Gln Phe Ser Leu Tyr Ile Gln Ala Leu Gln Tyr Ile Tyr Ser Ser 35 40 45 Lys Ser Gln Asp Asp Ile Asp Ser Phe Phe Gln Ile Gly Gly Ile His 50 55 60 Gly Leu Pro Tyr Val Pro Trp Asp Gly Ala Gly Asn Lys Pro Val Asp 65 70 75 80 Thr Asp Ala Trp Glu Gly Tyr Cys Thr His Gly Ser Val Leu Phe Pro 85 90 95 Thr Phe His Arg Pro Tyr Val Leu Leu Ile Glu Gln Ala Ile Gln Ala 100 105 110 Ala Ala Val Asp Ile Ala Ala Thr Tyr Ile Val Asp Arg Ala Arg Tyr 115 120 125 Gln Asp Ala Ala Leu Asn Leu Arg Gln Pro Tyr Trp Asp Trp Ala Arg 130 135 140 Asn Pro Val Pro Pro Pro Glu Val Ile Ser Leu Asp Glu Val Thr Ile 145 150 155 160 Val Asn Pro Ser Gly Glu Lys Ile Ser Val Pro Asn Pro Leu Arg Arg 165 170 175 Tyr Thr Phe His Pro Ile Asp Pro Ser Phe Pro Glu Pro Tyr Gln Ser 180 185 190 Trp Ser Thr Thr Leu Arg His Pro Leu Ser Asp Asp Ala Asn Ala Ser 195 200 205 Asp Asn Val Pro Glu Leu Lys Ala Thr Leu Arg Ser Ala Gly Pro Gln 210 215 220 Leu Lys Thr Lys Thr Tyr Asn Leu Leu Thr Arg Val His Thr Trp Pro 225 230 235 240 Ala Phe Ser Asn His Thr Pro Asp Asp Gly Gly Ser Thr Ser Asn Ser 245 250 255 Leu Glu Gly Ile His Asp Ser Val His Val Asp Val Gly Gly Asn Gly 260 265 270 Gln Met Ser Asp Pro Ser Val Ala Gly Phe Asp Pro Ile Phe Phe Mhe 275 280 285 His His Ala Gln Val Asp Arg Leu Leu Ser Leu Trp Ser Ala Leu Asn 290 295 300 Pro Arg Val Trp Ile Thr Asp Gly Pro Ser Gly Asp Gly Thr Trp Thr 305 310 315 320 Ile Pro Pro Asp Thr Val Val Gly Lys Asp Thr Asp Leu Thr Pro Phe 325 330 335 Trp Asn Thr Gln Ser Ser Tyr Trp Ile Ser Ala Asn Val Thr Asp Thr 340 345 350 Ser Lys Met Gly Tyr Thr Tyr Pro Glu Phe Asn Asn Leu Asp Met Gly 355 360 365 Asn Glu Va l Ala Val Arg Ser Ala Ile Ala Ala Gln Val Asn Lys Leu 370 375 380 Tyr Gly Gly Pro Phe Thr Lys Phe Ala Ala Ala Ile Gln Gln Pro Ser 385 390 395 400 400 Ser Gln Thr Thr Ala Asp Ala Ser Thr Ile Gly Asn Val Thr Ser Asp 405 410 415 Ala Ser Ser His Leu Val Asp Ser Lys Ile Asn Pro Thr Pro Asn Arg 420 425 430 Ser Ile Asp Asp Ala Pro Gln Val Lys Ile Ala Ser Thr Leu Arg Asn 435 440 445 Asn Glu Gln Lys Glu Phe Trp Glu Trp Thr Ala Arg Val Gln Val Lys 450 455 460 Lys Tyr Glu Ile Gly Gly Ser Phe Lys Val Leu Phe Phe Leu Gly Ser 465 470 475 475 480 Val Pro Ser Asp Pro Lys Glu Trp Ala Thr Asp Pro His Phe Val Gly 485 490 495 Ala Phe His Gly Phe Val Asn Ser Ser Ala Glu Arg Cys Ala Asn Cys 500 505 510 Arg Arg Gln Gln Asp Val Val Leu Glu Gly Phe Val His Leu Asn Glu 515 520 525 Gly Ile Ala Asn Ile Ser Asn Leu Asn Ser Phe Asp Pro Ile Val Val 530 535 540 Glu Pro Tyr Leu Lys Glu Asn Leu His Trp Arg Val Gln Lys Val Ser 545 550 555 560 Gly Glu Val Val Asn Leu Asp Ala Ala Thr Ser Pro Gly Ser Arg Ser 565 570 575 Cys Arg Ty r Ala Phe Gly Val Ala Ser Trp Arg Asp Phe Pro Ser Thr 580 585 590 Cys Arg Asp Thr Pro Pro Ser Pro Tyr His Thr Trp Ser Ser Trp Trp 595 600 600 605 Phe Ser Pro Gln Arg Arg Ile Phe Lys Pro Leu Ile Arg Gln Arg Val 610 615 620 Glu Phe Glu 625
【００４１】配列番号：２ 配列の長さ：１８８１ 配列の型：核酸 鎖の数：二本鎖 トポロジー：直鎖状 配列の種類：cDNA to mRNA 起源 生物名：レンチナス・エドデス（Lentinus edodes） 組織の種類：子実体 配列 ATGTCTCATT ATCTTGTCAC TGGCGCAACT GGAGGATCAA CCTCTGGGGC AGCAGCACCC 60 AATCGTCTCG AAATTAATGA TTTCGTCAAA CAAGAAGACC AGTTTTCTCT CTATATTCAG 120 GCTTTGCAAT ACATTTATTC AAGTAAAAGC CAAGACGATA TTGACTCCTT CTTCCAAATC 180 GGAGGGATCC ATGGTCTTCC GTATGTCCCT TGGGACGGCG CAGGAAATAA GCCAGTAGAC 240 ACTGACGCCT GGGAGGGATA TTGCACTCAT GGCAGCGTGT TATTTCCAAC CTTCCACCGT 300 CCGTATGTTC TACTCATCGA GCAAGCAATC CAGGCTGCGG CCGTCGATAT CGCCGCAACA 360 TACATCGTAG ATAGAGCTCG TTACCAGGAC GCCGCGTTGA ATCTACGTCA GCCATACTGG 420 GATTGGGCCC GAAACCCAGT TCCTCCGCCG GAAGTAATAT CTCTGGACGA GGTTACCATC 480 GTTAACCCAA GCGGAGAGAA AATCTCTGTT CCCAACCCTC TCCGACGTTA TACATTCCAC 540 CCCATAGATC CGTCCTTCCC TGAACCATAT CAGTCTTGGT CGACTACTCT TCGACATCCT 600 TTGTCCGATG ATGCCAATGC ATCGGACAAT GTTCCAGAAT TGAAAGCGAC GTTGAGAAGT 660 GCTGGTCCCC AACTCAAGAC CAAGACGTAC AACCTTCTGA CGCGAGTTCA TACATGGCCG 720 GCGTTCAGTA ACCATACGCC CGACGATGGA GGGAGTACCA GCAATAGTCT TGAAGGTATC 780 CACGACAGTG TCCACGTCGA TGTTGGTGGA AACGGGCAAA TGTCAGATCC TTCAGTAGCA 840 GGATTCGATC CCATTTTCTT TATGCACCAC GCCCAGGTCG ACCGCTTGCT TTCATTGTGG 900 TCTGCATTGA ATCCGAGGGT GTGGATTACC GACGGACCTT CTGGCGATGG GACATGGACT 960 ATCCCTCCCG ACACTGTAGT TGGAAAGGAT ACTGATCTTA CTCCGTTCTG GAACACCCAG 1020 TCATCGTATT GGATTTCTGC CAATGTGACC GATACGTCCA AGATGGGATA TACATATCCA 1080 GAATTTAACA ATCTCGATAT GGGAAATGAA GTTGCAGTTC GATCTGCTAT AGCTGCACAA 1140 GTTAACAAGC TCTATGGTGG ACCATTCACG AAATTCGCGG CAGCAATTCA ACAACCTTCT 1200 TCTCAAACTA CTGCAGACGC TTCCACGATT GGCAATGTCA CAAGCGATGC CTCTTCGCAC 1260 CTGGTAGACA GCAAAATCAA TCCGACGCCA AATAGAAGCA TTGATGATGC CCCTCAAGTA 1320 AAAATAGCTT CCACTCTAAG GAACAACGAA CAAAAGGAGT TTTGGGAATG GACTGCCCGT 1380 GTGCAGGTCA AGAAGTACGA AATAGGTGGA AGCTTCAAGG TCTTATTCTT CTTAGGCAGT 1440 GTGCCCAGTG ATCCCAAGGA ATGGGCTACT GATCCCCATT TTGTCGGAGC ATTCCACGGG 1500 TTCGTGAATA GCTCTGCCGA ACGATGCGCA AACTGTCGGC GTCAACAGGA TGTCGTTCTC 1560 GAAGGATTCG TGCATCTCAA CGAAGGCATT GCGAACATTT CCAACTTGAA CTCATTCGAC 1620 CCAATCGTTG TGGAACCGTA TCTTAAAGAG AACCTCCACT GGCGTGTGCA AAAGGTATCG 1680 GGCGAGGTAG TCAATTTGGA TGCAGCGACA TCCCCTGGAA GTCGTAGTTG TCGCTACGCG 1740 TTTGGAGTTG CCTCCTGGAG AGATTTTCCC AGTACCTGCA GAGACACACC ACCATCACCA 1800 TATCACACAT GGTCGTCCTG GTGGTTCTCG CCACAGCGTC GCATTTTCAA GCCCCTAATC 1860 AGACAAAGAG TGGAATTCGA A 1881SEQ ID NO: 2 Sequence length: 1881 Sequence type: nucleic acid Number of strands: double-stranded Topology: linear Sequence type: cDNA to mRNA Origin Organism name: Lentinus edodes type: fruiting sequence ATGTCTCATT ATCTTGTCAC TGGCGCAACT GGAGGATCAA CCTCTGGGGC AGCAGCACCC 60 AATCGTCTCG AAATTAATGA TTTCGTCAAA CAAGAAGACC AGTTTTCTCT CTATATTCAG 120 GCTTTGCAAT ACATTTATTC AAGTAAAAGC CAAGACGATA TTGACTCCTT CTTCCAAATC 180 GGAGGGATCC ATGGTCTTCC GTATGTCCCT TGGGACGGCG CAGGAAATAA GCCAGTAGAC 240 ACTGACGCCT GGGAGGGATA TTGCACTCAT GGCAGCGTGT TATTTCCAAC CTTCCACCGT 300 CCGTATGTTC TACTCATCGA GCAAGCAATC CAGGCTGCGG CCGTCGATAT CGCCGCAACA 360 TACATCGTAG ATAGAGCTCG TTACCAGGAC GCCGCGTTGA ATCTACGTCA GCCATACTGG 420 GATTGGGCCC GAAACCCAGT TCCTCCGCCG GAAGTAATAT CTCTGGACGA GGTTACCATC 480 GTTAACCCAA GCGGAGAGAGAAAATCTCTGTT CCCAACCCTC TCCGACGTTA TACATTCCAC 540 CCCATAGATC CGTCCTTCCTTCCTCTGACTCATCTAGCTACTG CGGACAAT GTTCCAGAAT TGAAAGCGAC GTTGAGAAGT 660 GCTGGTCCCC AACTCAAGAC CAAGACGTAC AACCTTCTGA CGCGAGTTCA TACATGGCCG 720 GCGTTCAGTA ACCATACGCC CGACGATGGA GGGAGTACCA GCAATAGTCT TGAAGGTATC 780 CACGACAGTG TCCACGTCGA TGTTGGTGGA AACGGGCAAA TGTCAGATCC TTCAGTAGCA 840 GGATTCGATC CCATTTTCTT TATGCACCAC GCCCAGGTCG ACCGCTTGCT TTCATTGTGG 900 TCTGCATTGA ATCCGAGGGT GTGGATTACC GACGGACCTT CTGGCGATGG GACATGGACT 960 ATCCCTCCCG ACACTGTAGT TGGAAAGGAT ACTGATCTTA CTCCGTTCTG GAACACCCAG 1020 TCATCGTATT GGATTTCTGC CAATGTGACC GATACGTCCA AGATGGGATA TACATATCCA 1080 GAATTTAACA ATCTCGATAT GGGAAATGAA GTTGCAGTTC GATCTGCTAT AGCTGCACAA 1140 GTTAACAAGC TCTATGGTGG ACCATTCACG AAATTCGCGG CAGCAATTCA ACAACCTTCT 1200 TCTCAAACTA CTGCAGACGC TTCCACGATT GGCAATGTCA CAAGCGATGC CTCTTCGCAC 1260 CTGGTAGACA GCAAAATCAA TCCGACGCCA AATAGAAGCA TTGATGATGC CCCTCAAGTA 1320 AAAATAGCTT CCACTCTAAG GAACAACGAA CAAAAGGAGT TTTGGGAATG GACTGCCCGT 1380 GTGCAGGTCA AGAAGTACGA AATAGGTGGA AGCTTCAAGG TCTTATTCTT CTTAGGCAGT 1440 GTGCCCAGTG ATCCCAAGGA ATGGGCTACT GAT CCCCATT TTGTCGGAGC ATTCCACGGG 1500 TTCGTGAATA GCTCTGCCGA ACGATGCGCA AACTGTCGGC GTCAACAGGA TGTCGTTCTC 1560 GAAGGATTCG TGCATCTCAA CGAAGGCATT GCGAACATTT CCAACTTGAA CTCATTCGAC 1620 CCAATCGTTG TGGAACCGTA TCTTAAAGAG AACCTCCACT GGCGTGTGCA AAAGGTATCG 1680 GGCGAGGTAG TCAATTTGGA TGCAGCGACA TCCCCTGGAA GTCGTAGTTG TCGCTACGCG 1740 TTTGGAGTTG CCTCCTGGAG AGATTTTCCC AGTACCTGCA GAGACACACC ACCATCACCA 1800 TATCACACAT GGTCGTCCTG GTGGTTCTCG CCACAGCGTC GCATTTTCAA GCCCCTAATC 1860 AGACAAAGAG TGGAATTCGA A 1881
【００４２】配列番号：３ 配列の長さ：２０５１ 配列の型：核酸 鎖の数：二本鎖 トポロジー：直鎖状 配列の種類：cDNA to mRNA 起源 生物名：レンチナス・エドデス（Lentinus edodes） 組織の種類：子実体 配列の特徴 特徴を表す記号：CDS 存在位置：21..1901 特徴を決定した方法：E 配列 TTTTCACAGA GTTCATTTAG 20 ATG TCT CAT TAT CTT GTC ACT GGC GCA ACT GGA GGA TCA ACC TCT GGG 68 Met Ser His Tyr Leu Val Thr Gly Ala Thr Gly Gly Ser Thr Ser Gly 1 5 10 15 GCA GCA GCA CCC AAT CGT CTC GAA ATT AAT GAT TTC GTC AAA CAA GAA 116 Ala Ala Ala Pro Asn Arg Leu Glu Ile Asn Asp Phe Val Lys Gln Glu 20 25 30 GAC CAG TTT TCT CTC TAT ATT CAG GCT TTG CAA TAC ATT TAT TCA AGT 164 Asp Gln Phe Ser Leu Tyr Ile Gln Ala Leu Gln Tyr Ile Tyr Ser Ser 35 40 45 AAA AGC CAA GAC GAT ATT GAC TCC TTC TTC CAA ATC GGA GGG ATC CAT 212 Lys Ser Gln Asp Asp Ile Asp Ser Phe Phe Gln Ile Gly Gly Ile His 50 55 60 GGT CTT CCG TAT GTC CCT TGG GAC GGC GCA GGA AAT AAG CCA GTA GAC 260 Gly Leu Pro Tyr Val Pro Trp Asp Gly Ala Gly Asn Lys Pro Val Asp 65 70 75 80 ACT GAC GCC TGG GAG GGA TAT TGC ACT CAT GGC AGC GTG TTA TTT CCA 308 Thr Asp Ala Trp Glu Gly Tyr Cys Thr His Gly Ser Val Leu Phe Pro 85 90 95 ACC TTC CAC CGT CCG TAT GTT CTA CTC ATC GAG CAA GCA ATC CAG GCT 356 Thr Phe His Arg Pro Tyr Val Leu Leu Ile Glu Gln Ala Ile Gln Ala 100 105 110 GCG GCC GTC GAT ATC GCC GCA ACA TAC ATC GTA GAT AGA GCT CGT TAC 404 Ala Ala Val Asp Ile Ala Ala Thr Tyr Ile Val Asp Arg Ala Arg Tyr 115 120 125 CAG GAC GCC GCG TTG AAT CTA CGT CAG CCA TAC TGG GAT TGG GCC CGA 452 Gln Asp Ala Ala Leu Asn Leu Arg Gln Pro Tyr Trp Asp Trp Ala Arg 130 135 140 AAC CCA GTT CCT CCG CCG GAA GTA ATA TCT CTG GAC GAG GTT ACC ATC 500 Asn Pro Val Pro Pro Pro Glu Val Ile Ser Leu Asp Glu Val Thr Ile 145 150 155 160 GTT AAC CCA AGC GGA GAG AAA ATC TCT GTT CCC AAC CCT CTC CGA CGT 548 Val Asn Pro Ser Gly Glu Lys Ile Ser Val Pro Asn Pro Leu Arg Arg 165 170 175 TAT ACA TTC CAC CCC ATA GAT CCG TCC TTC CCT GAA CCA TAT CAG TCT 596 Tyr Thr Phe His Pro Ile Asp Pro Ser Phe Pro Glu Pro Tyr Gln Ser 180 185 190 TGG TCG ACT ACT CTT CGA CAT CCT TTG TCC GAT GAT GCC AAT GCA TCG 644 Trp Ser Thr Thr Leu Arg His Pro Leu Ser Asp Asp Ala Asn Ala Ser 195 200 205 GAC AAT GTT CCA GAA TTG AAA GCG ACG TTG AGA AGT GCT GGT CCC CAA 692 Asp Asn Val Pro Glu Leu Lys Ala Thr Leu Arg Ser Ala Gly Pro Gln 210 215 220 CTC AAG ACC AAG ACG TAC AAC CTT CTG ACG CGA GTT CAT ACA TGG CCG 740 Leu Lys Thr Lys Thr Tyr Asn Leu Leu Thr Arg Val His Thr Trp Pro 225 230 235 240 GCG TTC AGT AAC CAT ACG CCC GAC GAT GGA GGG AGT ACC AGC AAT AGT 788 Ala Phe Ser Asn His Thr Pro Asp Asp Gly Gly Ser Thr Ser Asn Ser 245 250 255 CTT GAA GGT ATC CAC GAC AGT GTC CAC GTC GAT GTT GGT GGA AAC GGG 836 Leu Glu Gly Ile His Asp Ser Val His Val Asp Val Gly Gly Asn Gly 260 265 270 CAA ATG TCA GAT CCT TCA GTA GCA GGA TTC GAT CCC ATT TTC TTT ATG 884 Gln Met Ser Asp Pro Ser Val Ala Gly Phe Asp Pro Ile Phe Phe Met 275 280 285 CAC CAC GCC CAG GTC GAC CGC TTG CTT TCA TTG TGG TCT GCA TTG AAT 932 His His Ala Gln Val Asp Arg Leu Leu Ser Leu Trp Ser Ala Leu Asn 290 295 300 CCG AGG GTG TGG ATT ACC GAC GGA CCT TCT GGC GAT GGG ACA TGG ACT 980 Pro Arg Val Trp Ile Thr Asp Gly Pro Ser Gly Asp Gly Thr Trp Thr 305 310 315 320 ATC CCT CCC GAC ACT GTA GTT GGA AAG GAT ACT GAT CTT ACT CCG TTC 1028 Ile Pro Pro Asp Thr Val Val Gly Lys Asp Thr Asp Leu Thr Pro Phe 325 330 335 TGG AAC ACC CAG TCA TCG TAT TGG ATT TCT GCC AAT GTG ACC GAT ACG 1076 Trp Asn Thr Gln Ser Ser Tyr Trp Ile Ser Ala Asn Val Thr Asp Thr 340 345 350 TCC AAG ATG GGA TAT ACA TAT CCA GAA TTT AAC AAT CTC GAT ATG GGA 1124 Ser Lys Met Gly Tyr Thr Tyr Pro Glu Phe Asn Asn Leu Asp Met Gly 355 360 365 AAT GAA GTT GCA GTT CGA TCT GCT ATA GCT GCA CAA GTT AAC AAG CTC 1172 Asn Glu Val Ala Val Arg Ser Ala Ile Ala Ala Gln Val Asn Lys Leu 370 375 380 TAT GGT GGA CCA TTC ACG AAA TTC GCG GCA GCA ATT CAA CAA CCT TCT 1220 Tyr Gly Gly Pro Phe Thr Lys Phe Ala Ala Ala Ile Gln Gln Pro Ser 385 390 395 400 TCT CAA ACT ACT GCA GAC GCT TCC ACG ATT GGC AAT GTC ACA AGC GAT 1268 Ser Gln Thr Thr Ala Asp Ala Ser Thr Ile Gly Asn Val Thr Ser Asp 405 410 415 GCC TCT TCG CAC CTG GTA GAC AGC AAA ATC AAT CCG ACG CCA AAT AGA 1316 Ala Ser Ser His Leu Val Asp Ser Lys Ile Asn Pro Thr Pro Asn Arg 420 425 430 AGC ATT GAT GAT GCC CCT CAA GTA AAA ATA GCT TCC ACT CTA AGG AAC 1364 Ser Ile Asp Asp Ala Pro Gln Val Lys Ile Ala Ser Thr Leu Arg Asn 435 440 445 AAC GAA CAA AAG GAG TTT TGG GAA TGG ACT GCC CGT GTG CAG GTC AAG 1412 Asn Glu Gln Lys Glu Phe Trp Glu Trp Thr Ala Arg Val Gln Val Lys 450 455 460 AAG TAC GAA ATA GGT GGA AGC TTC AAG GTC TTA TTC TTC TTA GGC AGT 1460 Lys Tyr Glu Ile Gly Gly Ser Phe Lys Val Leu Phe Phe Leu Gly Ser 465 470 475 480 GTG CCC AGT GAT CCC AAG GAA TGG GCT ACT GAT CCC CAT TTT GTC GGA 1508 Val Pro Ser Asp Pro Lys Glu Trp Ala Thr Asp Pro His Phe Val Gly 485 490 495 GCA TTC CAC GGG TTC GTG AAT AGC TCT GCC GAA CGA TGC GCA AAC TGT 1556 Ala Phe His Gly Phe Val Asn Ser Ser Ala Glu Arg Cys Ala Asn Cys 500 505 510 CGG CGT CAA CAG GAT GTC GTT CTC GAA GGA TTC GTG CAT CTC AAC GAA 1604 Arg Arg Gln Gln Asp Val Val Leu Glu Gly Phe Val His Leu Asn Glu 515 520 525 GGC ATT GCG AAC ATT TCC AAC TTG AAC TCA TTC GAC CCA ATC GTT GTG 1652 Gly Ile Ala Asn Ile Ser Asn Leu Asn Ser Phe Asp Pro Ile Val Val 530 535 540 GAA CCG TAT CTT AAA GAG AAC CTC CAC TGG CGT GTG CAA AAG GTA TCG 1700 Glu Pro Tyr Leu Lys Glu Asn Leu His Trp Arg Val Gln Lys Val Ser 545 550 555 560 GGC GAG GTA GTC AAT TTG GAT GCA GCG ACA TCC CCT GGA AGT CGT AGT 1748 Gly Glu Val Val Asn Leu Asp Ala Ala Thr Ser Pro Gly Ser Arg Ser 565 570 575 TGT CGC TAC GCG TTT GGA GTT GCC TCC TGG AGA GAT TTT CCC AGT ACC 1796 Cys Arg Tyr Ala Phe Gly Val Ala Ser Trp Arg Asp Phe Pro Ser Thr 580 585 590 TGC AGA GAC ACA CCA CCA TCA CCA TAT CAC ACA TGG TCG TCC TGG TGG 1844 Cys Arg Asp Thr Pro Pro Ser Pro Tyr His Thr Trp Ser Ser Trp Trp 595 600 605 TTC TCG CCA CAG CGT CGC ATT TTC AAG CCC CTA ATC AGA CAA AGA GTG 1892 Phe Ser Pro Gln Arg Arg Ile Phe Lys Pro Leu Ile Arg Gln Arg Val 610 615 620 GAA TTC GAA TGA ACTATCGCGA TAAATAAATA ATGTCCTCGT TGTGCGTATG 1944 Glu Phe Glu 625 TGTAATGTTG GTTTTTTTAG CGGTTGAAGA CAGGTAGCGC TGAGCCTGGC CATTAATGGA 2004 GAATGATTCG GTACTCAAAA AAAAAAAAAA AAAAAAAAAA AAAAAAA 2051SEQ ID NO: 3 Sequence length: 2051 Sequence type: number of nucleic acid strands: double-stranded Topology: linear Sequence type: cDNA to mRNA Origin Organism: Lentinus edodes Type: fruiting body Features of the sequence Symbol indicating the feature: CDS Location: 21.1901 Method for determining the feature: E sequence TTTTCACAGA GTTCATTTAG 20 ATG TCT CAT TAT CTT GTC ACT GGC GCA ACT GGA GGA TCA ACC TCT GGG 68 Met Ser His Tyr Leu Val Thr Gly Ala Thr Gly Gly Ser Thr Ser Gly 1 5 10 15 GCA GCA GCA CCC AAT CGT CTC GAA ATT AAT GAT TTC GTC AAA CAA GAA 116 Ala Ala Ala Pro Asn Arg Leu Glu Ile Asn Asp Phe Val Lys Gln Glu 20 25 30 GAC CAG TTT TCT CTC TAT ATT CAG GCT TTG CAA TAC ATT TAT TCA AGT 164 Asp Gln Phe Ser Leu Tyr Ile Gln Ala Leu Gln Tyr Ile Tyr Ser Ser 35 40 45 AAA AGC CAA GAC GAT ATT GAC TCC TTC TTC CAA ATC GGA GGG ATC CAT 212 Lys Ser Gln Asp Asp Ile Asp Ser Phe Phe Gln Ile Gly Gly Ile His 50 55 60 GGT CTT CCG TAT GTC CCT TGG GAC G GC GCA GGA AAT AAG CCA GTA GAC 260 Gly Leu Pro Tyr Val Pro Trp Asp Gly Ala Gly Asn Lys Pro Val Asp 65 70 75 80 ACT GAC GCC TGG GAG GGA TAT TGC ACT CAT GGC AGC GTG TTA TTT CCA 308 Thr Asp Ala Trp Glu Gly Tyr Cys Thr His Gly Ser Val Leu Phe Pro 85 90 95 ACC TTC CAC CGT CCG TAT GTT CTA CTC ATC GAG CAA GCA ATC CAG GCT 356 Thr Phe His Arg Pro Tyr Val Leu Leu Ile Glu Gln Ala Ile Gln Ala 100 105 110 GCG GCC GTC GAT ATC GCC GCA ACA TAC ATC GTA GAT AGA GCT CGT TAC 404 Ala Ala Val Asp Ile Ala Ala Thr Tyr Ile Val Asp Arg Ala Arg Tyr 115 120 125 CAG GAC GCC GCG TTG AAT CTA CGT CAG CCA TAC TGG GAT TGG GCC CGA 452 Gln Asp Ala Ala Leu Asn Leu Arg Gln Pro Tyr Trp Asp Trp Ala Arg 130 135 140 AAC CCA GTT CCT CCG CCG GAA GTA ATA TCT CTG GAC GAG GTT ACC ATC 500 Asn Pro Val Pro Pro Pro Glu Val Ile Ser Leu Asp Glu Val Thr Ile 145 150 155 160 GTT AAC CCA AGC GGA GAG AAA ATC TCT GTT CCC AAC CCT CTC CGA CGT 548 Val Asn Pro Ser Gly Glu Lys Ile Ser Val Pro Asn Pro Leu Arg Arg 165 170 175 TAT ACA TTC CAC CCC ATA GAT CCG TCC TTC CCT GAA CCA TAT CAG TCT 596 Tyr Thr Phe His Pro Ile Asp Pro Ser Phe Pro Glu Pro Tyr Gln Ser 180 185 190 TGG TCG ACT ACT CTT CGA CAT CCT TTG TCC GAT GAT GCC AAT GCA TCG 644 Trp Ser Thr Thr Leu Arg His Pro Leu Ser Asp Asp Ala Asn Ala Ser 195 200 205 GAC AAT GTT CCA GAA TTG AAA GCG ACG TTG AGA AGT GCT GGT CCC CAA 692 Asp Asn Val Pro Glu Leu Lys Ala Thr Leu Arg Ser Ala Gly Pro Gln 210 215 220 CTC AAG ACC AAG ACG TAC AAC CTT CTG ACG CGA GTT CAT ACA TGG CCG 740 Leu Lys Thr Lys Thr Tyr Asn Leu Leu Thr Arg Val His Thr Trp Pro 225 230 235 240 GCG TTC AGT AAC CAT ACG CCC GAC GAT GGA GGG AGT ACC AGC AAT AGT 788 Ala Phe Ser Asn His Thr Pro Asp Asp Gly Gly Ser Thr Ser Asn Ser 245 250 255 CTT GAA GGT ATC CAC GAC AGT GTC CAC GTC GAT GTT GGT GGA AAC GGG 836 Leu Glu Gly Ile His Asp Ser Val His Val Asp Val Gly Gly Asn Gly 260 265 270 CAA ATG TCA GAT CCT TCA GTA GCA GGA TTC GAT CCC ATT TTC TTT ATG 884 Gln Met Ser Asp Pro Ser Val Ala Gly Phe Asp Pro Ile Phe Phe Met 275 280 285 CAC CAC GCCCAG GTC GAC CGC TTG CTT TCA TTG TGG TCT GCA TTG AAT 932 His His Ala Gln Val Asp Arg Leu Leu Ser Leu Trp Ser Ala Leu Asn 290 295 300 CCG AGG GTG TGG ATT ACC GAC GGA CCT TCT GGC GAT GGG ACA TGG ACT 980 Pro Arg Val Trp Ile Thr Asp Gly Pro Ser Gly Asp Gly Thr Trp Thr 305 310 315 320 ATC CCT CCC GAC ACT GTA GTT GGA AAG GAT ACT GAT CTT ACT CCG TTC 1028 Ile Pro Pro Asp Thr Val Val Gly Lys Asp Thr Asp Leu Thr Pro Phe 325 330 335 TGG AAC ACC CAG TCA TCG TAT TGG ATT TCT GCC AAT GTG ACC GAT ACG 1076 Trp Asn Thr Gln Ser Ser Tyr Trp Ile Ser Ala Asn Val Thr Asp Thr 340 345 350 TCC AAG ATG GGA TAT ACA TAT CCA GAA TTT AAC AAT CTC GAT ATG GGA 1124 Ser Lys Met Gly Tyr Thr Tyr Pro Glu Phe Asn Asn Leu Asp Met Gly 355 360 365 AAT GAA GTT GCA GTT CGA TCT GCT ATA GCT GCA CAA GTT AAC AAG CTC 1172 Asn Glu Val Ala Val Arg Ser Ala Ile Ala Ala Gln Val Asn Lys Leu 370 375 380 TAT GGT GGA CCA TTC ACG AAA TTC GCG GCA GCA ATT CAA CAA CCT TCT 1220 Tyr Gly Gly Pro Phe Thr Lys Phe Ala Ala Ala Ile Gln Gln Pro Ser 385 390 395 400 TCT CAA ACT ACT GCA GAC GCT TCC ACG ATT GGC AAT GTC ACA AGC GAT 1268 Ser Gln Thr Thr Ala Asp Ala Ser Thr Ile Gly Asn Val Thr Ser Asp 405 410 415 GCC TCT TCG CAC CTG GTA GAC AGC AAA ATC AAT CCG ACG CCA AAT AGA 1316 Ala Ser Ser His Leu Val Asp Ser Lys Ile Asn Pro Thr Pro Asn Arg 420 425 430 AGC ATT GAT GAT GCC CCT CAA GTA AAA ATA GCT TCC ACT CTA AGG AAC 1364 Ser Ile Asp Asp Ala Pro Gln Val Lys Ile Ala Ser Thr Leu Arg Asn 435 440 445 AAC GAA CAA AAG GAG TTT TGG GAA TGG ACT GCC CGT GTG CAG GTC AAG 1412 Asn Glu Gln Lys Glu Phe Trp Glu Trp Thr Ala Arg Val Gln Val Lys 450 455 460 AAG TAC GAA ATA GGT GGA AGC TTC AAG GTC TTA TTC TTC TTA GGC AGT 1460 Lys Tyr Glu Ile Gly Gly Ser Phe Lys Val Leu Phe Phe Leu Gly Ser 465 470 475 475 480 GTG CCC AGT GAT CCC AAG GAA TGG GCT ACT GAT CCC CAT TTT GTC GGA 1508 Val Pro Ser Asp Pro Lys Glu Trp Ala Thr Asp Pro His Phe Val Gly 485 490 495 GCA TTC CAC GGG TTC GTG AAT AGC TCT GCC GAA CGA TGC GCA AAC TGT 1556 Ala Phe His Gly Phe Val Asn Ser Ser Ala Glu Arg Cys Ala Asn Cys 500 505 510 CGG CGT CAA CAG GAT GTC GTT CTC GAA GGA TTC GTG CAT CTC AAC GAA 1604 Arg Arg Gln Gln Asp Val Val Leu Glu Gly Phe Val His Leu Asn Glu 515 520 525 GGC ATT GCG AAC ATT TCC AAC TTG AAC TCA TTC GAC CCA ATC GTT GTG 1652 Gly Ile Ala Asn Ile Ser Asn Leu Asn Ser Phe Asp Pro Ile Val Val 530 535 540 GAA CCG TAT CTT AAA GAG AAC CTC CAC TGG CGT GTG CAA AAG GTA TCG 1700 Glu Pro Tyr Leu Lys Glu Asn Leu His Trp Arg Val Gln Lys Val Ser 545 550 555 560 GGC GAG GTA GTC AAT TTG GAT GCA GCG ACA TCC CCT GGA AGT CGT AGT 1748 Gly Glu Val Val Asn Leu Asp Ala Ala Thr Ser Pro Gly Ser Arg Ser 565 570 575 TGT CGC TAC GCG TTT GGA GTT GCC TCC TGG AGA GAT TTT CCC AGT ACC 1796 Cys Arg Tyr Ala Phe Gly Val Ala Ser Trp Arg Asp Phe Pro Ser Thr 580 585 590 590 TGC AGA GAC ACA CCA CCA TCA CCA TAT CAC ACA TGG TCG TCC TGG TGG 1844 Cys Arg Asp Thr Pro Pro Ser Pro Tyr His Thr Trp Ser Ser Trp Trp 595 600 605 TTC TCG CCA CAG CGT CGC ATT TTC AAG CCC CTA ATC AGA CAA AGA GTG 1892 Phe Ser Pro Gln Arg Arg Ile Phe Lys Pro Leu Ile Arg Gln Arg Val 610 615 620 GAA TTC GAA TGA ACTATCGCGA TAAATAAATA ATGTCCTCGT TGTGCGTATG 1944 Glu Phe Glu 625 TGTAATGTTG GTTTTTTTAG CGGTTGAAGA CAGGTAGCGC TGAGCCTGAAAAAATGAAGAAAAAATGAA
【００４３】配列番号：４ 配列の長さ：２２ 配列の型：核酸 鎖の数：一本鎖 トポロジー：直鎖状 配列の種類：他の核酸（合成ＤＮＡ） 配列： TYCARATHGG NGGNATHCAY GG 22SEQ ID NO: 4 Sequence length: 22 Sequence type: number of nucleic acid chains: single stranded Topology: linear Sequence type: other nucleic acid (synthetic DNA) Sequence: TYCARATHGG NGGNATHCAY GG 22
【００４４】配列番号：５ 配列の長さ：２２ 配列の型：核酸 鎖の数：一本鎖 トポロジー：直鎖状 配列の種類：他の核酸（合成ＤＮＡ） 配列： RNCKRTCNAC YTGNGCRTGR TG 22SEQ ID NO: 5 Sequence length: 22 Sequence type: number of nucleic acid chains: single-stranded Topology: linear Sequence type: other nucleic acid (synthetic DNA) Sequence: RNCKRTCNAC YTGNGCRTGR TG 22
【００４５】配列番号：６ 配列の長さ：２５ 配列の型：核酸 鎖の数：一本鎖 トポロジー：直鎖状 配列の種類：他の核酸（合成ＤＮＡ） 配列： GCGCAGGAAA TAAGCCAGTA GACAC 25Sequence number: 6 Sequence length: 25 Sequence type: Number of nucleic acid strands: Single strand Topology: Linear Sequence type: Other nucleic acid (synthetic DNA) Sequence: GCGCAGGAAA TAAGCCAGTA GACAC 25
【００４６】配列番号：７ 配列の長さ：２０ 配列の型：核酸 鎖の数：一本鎖 トポロジー：直鎖状 配列の種類：他の核酸（合成ＤＮＡ） 配列： GCGTGGTGCA TAAAGAAAAT ２０SEQ ID NO: 7 Sequence length: 20 Sequence type: number of nucleic acid chains: single-stranded Topology: linear Sequence type: other nucleic acid (synthetic DNA) Sequence: GCGTGGTGCA TAAAGAAAAT 20
───────────────────────────────────────────────────── フロントページの続き (51)Int.Cl.6 識別記号 ＦＩ Ｃ１２Ｒ 1:645） (Ｃ１２Ｎ 9/02 Ｃ１２Ｒ 1:645） (72)発明者 江井 仁 岩手県北上市新穀町１丁目６番地 メルベ イユ１−304 (72)発明者 江尻 愼一郎 岩手県盛岡市黒石野１丁目７番地──────────────────────────────────────────────────の Continuing on the front page (51) Int.Cl. 6 Identification code FI C12R 1: 645) (C12N 9/02 C12R 1: 645) (72) Inventor Hitoshi Ei 1-6-6 Shinshinkumachi, Kitakami-shi, Iwate Prefecture Merveille 1-304 (72) Inventor Shinichiro Ejiri 1-7-7 Kuroishino, Morioka City, Iwate Prefecture
Claims (3)
は該アミノ酸配列において１若しくは複数のアミノ酸が
欠失、置換若しくは付加されたものを含み、且つチロシ
ナーゼ活性をもたらすタンパク質をコードするＤＮＡ。Claims: 1. A DNA comprising an amino acid sequence represented by SEQ ID NO: 1 or a deletion or substitution or addition of one or more amino acids in the amino acid sequence, and a DNA encoding a protein that produces tyrosinase activity.
塩基配列を含むものである請求項１記載のＤＮＡ。2. The DNA according to claim 1, wherein the DNA comprises the nucleotide sequence represented by SEQ ID NO: 2 or 3.
来のものである請求項１又は２記載のＤＮＡ。3. The DNA according to claim 1, wherein the DNA is derived from a fruit body or a mycelium of Shiitake mushroom.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| JP8337127A JPH10174586A (en) | 1996-12-17 | 1996-12-17 | Tyrosinase gene of shiitake (lentinus edodes) |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| JP8337127A JPH10174586A (en) | 1996-12-17 | 1996-12-17 | Tyrosinase gene of shiitake (lentinus edodes) |
Publications (1)
| Publication Number | Publication Date |
|---|---|
| JPH10174586A true JPH10174586A (en) | 1998-06-30 |
Family
ID=18305705
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| JP8337127A Pending JPH10174586A (en) | 1996-12-17 | 1996-12-17 | Tyrosinase gene of shiitake (lentinus edodes) |
Country Status (1)
| Country | Link |
|---|---|
| JP (1) | JPH10174586A (en) |
Cited By (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| WO2001048210A1 (en) * | 1999-12-23 | 2001-07-05 | Fudan University | A novel polypeptide, tyrosinase 10 and the polynucleotide encoding the polypeptide |
| EP1848798A1 (en) * | 2005-02-10 | 2007-10-31 | Valtion Teknillinen Tutkimuskeskus | Novel microbial enzymes and their use |
| JP2012055208A (en) * | 2010-09-07 | 2012-03-22 | Tokyo Univ Of Agriculture & Technology | Plant cell accumulating l-dopa, and use thereof |
-
1996
- 1996-12-17 JP JP8337127A patent/JPH10174586A/en active Pending
Cited By (6)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| WO2001048210A1 (en) * | 1999-12-23 | 2001-07-05 | Fudan University | A novel polypeptide, tyrosinase 10 and the polynucleotide encoding the polypeptide |
| EP1848798A1 (en) * | 2005-02-10 | 2007-10-31 | Valtion Teknillinen Tutkimuskeskus | Novel microbial enzymes and their use |
| EP1848798A4 (en) * | 2005-02-10 | 2008-08-27 | Valtion Teknillinen | Novel microbial enzymes and their use |
| AU2006212149B2 (en) * | 2005-02-10 | 2010-03-04 | Valtion Teknillinen Tutkimuskeskus | Novel microbial enzymes and their use |
| US7910344B2 (en) | 2005-02-10 | 2011-03-22 | Valtion Teknillinen Tutkimuskeskus | Proteins having tyrosinase activity |
| JP2012055208A (en) * | 2010-09-07 | 2012-03-22 | Tokyo Univ Of Agriculture & Technology | Plant cell accumulating l-dopa, and use thereof |
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