CN106967696A - 一种蒿属植物花粉的过敏原及其应用 - Google Patents
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Abstract
本发明公开了一种蒿属植物花粉的过敏原及其应用。一种蒿属植物花粉的过敏原,它为蒿属植物的半乳糖氧化酶,氨基酸序列为SEQ ID No.1、SEQ ID No.2、SEQ ID No.3、SEQ ID No.4、SEQ ID No.5、SEQ ID No.6或SEQ ID No.7。本发明一种蒿属植物花粉的过敏原为新发现的一种过敏原,为蒿属植物的半乳糖氧化酶,是蒿属植物中发现的第7种过敏原。该过敏原可被用于制备治疗或预防过敏症的药物,或者用于制备过敏原检测的试剂盒。
Description
技术领域
本发明涉及生物技术相关技术领域,特别是涉及一种蒿属植物花粉的过敏原及其应用。
背景技术
Ⅰ型过敏症也称Ⅰ型超敏反应性疾病,有关的疾病包括特应性疾病、变应性鼻炎、过敏性结膜炎、特应性皮炎和过敏性哮喘(外源性)和一些荨麻疹、胃肠道食物反应和全身性过敏反应等。Ⅰ型过敏症在世界范围内普遍存在,患特应性疾病的患者对吸入或摄入的物质(过敏原)产生由IgE抗体介导的超敏反应,而这些物质对无特应性疾病的人是无害的。除特应性皮炎外,通常IgE抗体会介导超敏反应。
引发Ⅰ型过敏症的物质称为过敏原,是蛋白、糖蛋白或多肽。从粘膜摄入后,这些过敏原与结合在致敏个体的肥大细胞表面上的IgE抗体反应。如果两个IgE抗体通过过敏原彼此交联,就会导致效应细胞释放介质(如组胺、***素)和细胞因子,从而产生相应的临床症状。
过敏原的命名是根据物种的属和种国际免疫学会联合会(I.U.I.S)下属的过敏原命名专门委员会(Allergen Nomenclature Sub-committee)(http://www.allergen.org)制定了***的命名规则。命名主要根据过敏原来源物种的拉丁名,所在属名的前三个字母加种的1-2个字母,然后加过敏原代号,如北艾(Artemesia vugaris)的过敏原为Art v 1~Art v 6,黄花蒿(Artemesia annua)类似的过敏原为Art an 1~Art an 6。
蒿花粉是中国北方和西部主要的吸入类致敏原,引起夏秋季过敏人群的鼻炎,结膜炎、皮疹、哮喘等过敏性疾病。目前欧洲用北艾已经鉴定出6种过敏原蛋白(Art v 1~Artv 6)。体外诊断主要是用花粉的提取物。蛋白组分在国外有科研应用性两种(Art v 1和Artv 3)。国内外的一些血清免疫蛋白研究在1996年就发现有一个60~70kDa蛋白是潜在过敏原,但一直不清楚是哪种蛋白以及具体的氨基酸序列。我国蒿属(Artemisia)过敏源的主要种类为黄花蒿(A.annua),艾蒿(A.arygi),茵陈蒿(A.capillaries),裂叶莲蒿(A.gmelinii),野艾蒿(A.lavandulifolia),大籽蒿(A.sieversiana),欧洲蒿属植物种类主要为北艾(A.vulgaris),仅在我国宁夏等西北部有少量分布,目前进入我国蒿诊断试剂市场的国外品牌赛默飞世尔科技公司的试剂ImmunoCAP所用的材料就是北艾。
发明内容
本发明提供了一种蒿属植物花粉的过敏原及其应用。
一种蒿属植物花粉的过敏原,它为蒿属植物的半乳糖氧化酶,氨基酸序列为SEQID No.1、SEQ ID No.2、SEQ ID No.3、SEQ ID No.4、SEQ ID No.5、SEQ ID No.6或SEQ IDNo.7。所述蒿属植物为植物分类学上属于蒿属的植物,包括黄花蒿(A.annua),艾蒿(A.arygi),茵陈蒿(A.capillaries),裂叶莲蒿(A.gmelinii),野艾蒿(A.lavandulifolia),大籽蒿(A.sieversiana),北艾(A.vulgaris)等。所述的7种序列分别为上述列举的7种蒿属植物各自半乳糖氧化酶的氨基酸序列,序列间具有高度保守性,序列相似性(Identity)达到94.1%。各物种来源的半乳糖氧化酶均具有相似的致敏性。
本发明又提供了编码所述过敏原的DNA分子,核苷酸序列为SEQ ID No.8、SEQ IDNo.9、SEQ ID No.10、SEQ ID No.11、SEQ ID No.12、SEQ ID No.13或SEQ ID No.14。所述的DNA分子即为编码上述7种氨基酸序列的基因,各基因之间具有高度的保守性,序列相似性(Identity)达到92.2%。
本发明又提供了包含所述DNA分子的重组表达载体。所述重组表达载体可以为各种用于蛋白表达的质粒。
本发明又提供了包含所述DNA分子或包含所述重组表达载体的宿主生物体。所述宿主生物体可以为用于原核表达的大肠杆菌、用于真核表达的酵母、昆虫细胞、哺乳动物细胞(如CHO细胞)等各种蛋白表达***。
本发明还提供了所述过敏原或所述DNA分子在制备治疗或预防过敏症的药物中的应用。
本发明还提供了一种用于治疗或预防过敏症的药物,包含所述过敏原或所述DNA分子中的至少一种。
优选的,所述药物为疫苗。
有效治疗过敏症的传统方法是特异性免疫治疗或脱敏,即以递增的剂量对过敏患者皮下注射天然过敏原提取物。
本发明还提供了所述过敏原或所述DNA分子在制备过敏原检测的试剂盒中的应用。
本发明还提供了一种用于过敏原检测的试剂盒,包含所述过敏原或所述DNA分子中的至少一种。
对于过敏症患者,一个重要的手段是检查出引起过敏的过敏原,通过有针对性的减少与过敏原的接触可有效预防过敏的发生。本发明所述过敏原或所述DNA分子可以用于单独或者与其他过敏原一起使用,制备成用于筛查过敏症患者的过敏原的试剂盒。
本发明一种蒿属植物花粉的过敏原为新发现的一种过敏原,为蒿属植物的半乳糖氧化酶,是蒿属植物中发现的第7种过敏原。该过敏原可被用于制备治疗或预防过敏症的药物,或者用于制备过敏原检测的试剂盒。
附图说明
图1为7种蒿的花粉蛋白提取物检测图,其中,图A为电泳图,图B为免疫印迹图,泳道1~7分别为黄花蒿、艾蒿、茵陈蒿、裂叶莲蒿、野艾蒿、大籽蒿、北艾。
图2为黄花蒿花粉提取物与15例单个血清的蒿花粉过敏阳性血清作用的免疫印迹图,其中S为黄花蒿花粉蛋白提取物,p49~60、88~90为实验组,N为阴性对照组。
图3为实施例3中纯化后的黄花蒿半乳糖氧化酶蛋白的电泳图。
图4为实施例3中纯化后的黄花蒿半乳糖氧化酶蛋白总体蛋白质分子量质谱图。
图5为实施例3中肽段匹配序列图。
图6为皮试实验结果图,其中1:组胺阳性对照(10μg/mL);2:黄花蒿花粉蛋白提取物(1μg/mL);3:艾蒿花粉蛋白提取物(1μg/mL);4:大籽蒿花粉蛋白提取物(1μg/mL);5:纯化天然半乳糖氧化酶nArt an 7(0.25μg/mL);6:纯化天然半乳糖氧化酶nArt an 7(0.5μg/mL)。
具体实施方式
材料
黄花蒿、艾蒿、茵陈蒿、裂叶莲蒿、大籽蒿:采集自山西省;
野艾蒿:采集自浙江省;
北艾:购自德国默克公司。
蒿花粉过敏阳性血清:来源于浙江大学医学院附属第二医院以及科研合作的山西省太原市第三人民医院临床病例,伦理批号No.2011-R-1,患者都同意并签有知情同意书。
实施例1
提取蒿属(Artemisia)7个代表性种(黄花蒿、艾蒿、茵陈蒿、裂叶莲蒿、野艾蒿、大籽蒿、北艾)花粉中的蛋白。蛋白提取纯化方法:2g花粉溶在35mL PBS(pH 7)中,在4度摇床过夜12小时,10000g离心10分钟(4度)后经过0.22μm Millipore膜过滤获得花粉蛋白提取物。
7种蒿属植物的花粉蛋白提取物,每种50μg,分别用12%浓度的SDS PAGE电泳分离,然后将蛋白转膜到PVDF膜上,5%脱脂奶粉封闭。蒿花粉过敏阳性血清35例混合为血清池进行孵育。结合的IgE用羊抗人IgE HRP(1∶3000),加底物ECL化学发光检测(BioRadChemiDoc),SDS PAGE胶染色和免疫印迹的检测结果如图1所示,共鉴定出三条一致的反应蛋白条带,其中10和25kD对应为已知的非特异脂质转移蛋白(nsLTP)和类似防护蛋白,还有一条大小在60~70kD的为未知蛋白。
使用15例单个血清的蒿花粉过敏阳性血清对黄花蒿花粉蛋白提取物进行进一步验证,免疫印迹检测结果如图2所示,结果与混合血清池检测结果一致。
实施例2
黄花蒿新鲜冰冻花粉提取mRNA,反转录为cDNA,RNA-seq测序使用IlluminaHiseq2000。通过SOAPdenovo序列组装进行基因注释。花粉中蛋白全谱采用胰肽酶解肽段后用Q-EXACTIVE(ThermoFisher,USA)测定质谱。肽段质谱匹配用Mascot,设置参数为20ppm和0.6Da。通过转录组RNA-seq、总蛋白质谱相结合的方法,鉴定出黄花蒿花粉中有2000多种蛋白。
将实施例1中经SDS PAGE鉴定分离到的分子量为60~70kD的蛋白条带割胶,进行蛋白酶解质谱测试,参照转录组和蛋白组鉴定的所有可能蛋白氨基酸序列,获得一种注释为半乳糖氧化酶的蛋白为候选,等电点为4.9,酸性蛋白。
实施例3
黄花蒿花粉蛋白提取物(按实施例1所述方法提取)50mL,在AKTA蛋白纯化***上用阴离子交换柱(Explorer 100,GE公司)和蛋白分子筛(Super 75 10/300GL型,GE公司)分离纯化目标蛋白,获得大小约为62kDa的蛋白(图3)。
纯化所得目标蛋白质谱分析,总体蛋白质分子量质谱为61.95kDa(图4),与理论计算的相差1‰,胰肽酶解肽段质谱匹配的氨基酸覆盖度(下划线部分)达到整个氨基酸序列长度的62%(图5)。对应氨基酸序列,得到编码基因全长SEQ ID No.8。
实施例4
将从黄花蒿花粉蛋白提取物中按实施例3方法分离纯化的天然半乳糖氧化酶蛋白(Art an7)进行生物素标记,纯化半乳糖氧化酶蛋白结合在链霉亲和素ImmunoCAPs(Phadia,Uppsala,Sweden)上。
用ImmunoCAP 100进行测试血清中特异IgE。测试100个蒿过敏患者血清,IgE滴度大于0.35kUA/L为阳性,结果如表1所示,检测结果阳性率超过95%。
表1
注:症状缩写:AS——哮喘;C——结膜炎;R——鼻炎;U——皮疹。N1~N5为阴性对照血清。
实施例5
通过PCR引物克隆测序以及蒿花粉的RNA-seq,获得7种蒿准确的半乳糖氧化酶氨基酸序列。
艾蒿、茵陈蒿、裂叶莲蒿、野艾蒿、大籽蒿、北艾花粉中的半乳糖氧化酶编码基因用引物(上游引物:ATGGCTTCATCAATTAAAACA;下游引物:TAACCCAAGTGTAGCTGATT)进行克隆测序,获得基因序列SEQ ID No.9~14(已去除其中编码信号肽的序列)。
实施例6
将黄花蒿、艾蒿、大籽蒿的花粉蛋白提取物(实施例1方法制备),以及从黄花蒿花粉蛋白提取物中按实施例3方法分离纯化的半乳糖氧化酶蛋白进行体内试验(皮试),皮试方法如下:将经过浓度稀释的半乳糖氧化酶蛋白,选择0.5μg/mL进行皮下注射0.1mL;黄花蒿、艾蒿、大籽蒿的花粉蛋白提取物稀释到1μg/mL皮下注射0.1mL;阳性对照为浓度为10μg/mL的组胺皮下注射0.1mL;阴性对照为皮下注射0.1mL变应原溶媒。均使用变应原溶媒进行稀释(太原制药厂)。皮试的丘斑直径达到组胺阳性对照的1/3即认定为阳性。皮试对象为表2中的P54病例。
试验结果如图6和表2所示,发现黄花蒿、艾蒿、大籽蒿的花粉蛋白提取物以及经分离纯化所得的黄花蒿半乳糖氧化酶蛋白均可以引发过敏反应,且分离纯化所得的黄花蒿半乳糖氧化酶蛋白所引起的过敏反应较强烈。阴性对照结果无丘斑产生,仍然为正常皮肤样,图片未展示。
进一步在7个蒿过敏病例(P101-107)和一个阴性对照病例进行黄花蒿提取物和纯化蛋白的皮试,新发现的组分半乳糖氧化酶IgE值较低时(小于0.68),皮试显现阴性(表2)。
表2
注:症状缩写:AS——哮喘;C——结膜炎;R——鼻炎;U——皮疹。N6为阴性对照血清。
SEQUENCE LISTING
<110> 杭州艾乐吉生物科技有限公司
<120> 一种蒿属植物花粉的过敏原及其应用
<130>
<160> 16
<170> PatentIn version 3.3
<210> 1
<211> 571
<212> PRT
<213> 黄花蒿(A. annua)
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Ser Gly Asn Leu Trp Pro Asn Gly Asn Leu Met Ala Thr Gly Gly Thr
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Phe Ser Gly Asp Lys Ala Ile Arg Val Ile Ala Asn Asp Asp Pro Lys
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Gly Asp Phe Thr Thr Lys Ile Gly Ala Leu Ala Asp Thr Arg Trp Tyr
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Pro Lys Arg Phe Asp Leu Pro Phe Met Gln Gln Thr Thr Glu Pro Pro
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Leu Gly Pro Gly Arg Pro Val Glu Asn Asn Leu Tyr Pro Phe Leu Phe
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Leu Leu Pro Asp Gly Asn Ile Phe Leu Phe Ala Asn Asn Arg Ala Ile
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Thr Phe Glu Pro Ala Thr Gly Lys Thr Val Lys Glu Phe Pro Val Leu
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Pro Gly Gly Ser Arg Asn Tyr Pro Pro Ser Gly Ser Ala Ala Leu Phe
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Pro Leu Lys Leu Thr Ala Asp Asn Ala Pro Val Ile Pro Glu Ile Val
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Ile Cys Gly Gly Asn Gln Pro Asn Ala Tyr Glu Leu Val Asp Ala Arg
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His Val Thr Glu Lys Gln Phe Leu Pro Ala Leu Gln Asp Cys Asn Arg
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Ile Gln Pro Met Ala Ala Asp Ala Ala Trp Ile Pro Glu Gln Asn Met
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Pro Ser Pro Arg Thr Met Gly Asp Leu Ile His Leu Ala Asn Gly Asp
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370 375 380
Ala Thr Glu Ala Asn Leu Thr Pro Val Leu Tyr Thr Pro Tyr Lys Pro
385 390 395 400
Met Gly Gln Arg Phe Lys Glu Leu Thr Pro Thr Thr Ile Ala Arg Met
405 410 415
Tyr His Ser Cys Ser Ala Leu Leu Pro Asp Thr Arg Val Leu Val Ala
420 425 430
Gly Ser Asn Met His Gln Phe Tyr Thr Phe Asp Thr Glu Phe Pro Thr
435 440 445
Glu Leu Arg Val Glu Lys Phe Ser Pro Pro Tyr Leu Asp Pro Ala Leu
450 455 460
Glu Thr Glu Arg Thr Gln Ile Asp Pro Ala Asn Thr Asp Ala Val Leu
465 470 475 480
Lys Tyr Gly Lys Pro Phe Lys Ile Thr Ala Ala Leu Met Glu Lys Gln
485 490 495
Pro Leu Val Leu Gly Glu Val Lys Val Thr Leu Leu Tyr Pro Pro Phe
500 505 510
Thr Thr His Gly Phe Ser Gln Asn Gln Arg Met Ile Val Pro Ala Ile
515 520 525
Thr Ser Val Gln Asn Gly Val Ile Thr Ala Val Ala Pro Pro Ser Gly
530 535 540
Gln Ile Ala Pro Pro Gly Tyr Tyr Ile Met Phe Val Ser His Leu Gly
545 550 555 560
Ile Pro Gly Ala Gly Ile Trp Val His Ile Asp
565 570
<210> 2
<211> 571
<212> PRT
<213> 艾蒿(A. arygi)
<400> 2
Ala Pro Asp Ile Thr Asp Gly Gly Asp Lys Pro Gly Pro Gln Val Asp
1 5 10 15
Asp Gly Gly Gly Asp Lys Pro Val Pro Gly Asn Asn Asp Gly Ala Ser
20 25 30
Asp Tyr Ala Lys Pro Ala Phe Glu Pro Glu Phe Met Gly Ala Trp Val
35 40 45
Ile Asp Asn Pro Asn Ala Gly Val Ala Ala Met Gln Leu Gln Leu Met
50 55 60
Pro Asn Asp Gln Ile Val Trp Phe Asp Thr Thr Ser Leu Gly Ala Ser
65 70 75 80
Gly Tyr Lys Leu Pro Glu Gly Thr Pro Cys Pro Asn Asn Pro Asp Ala
85 90 95
Asn Asn Gln Pro Asp Cys Tyr Val His Gly Ile Ala Tyr Asp Trp Lys
100 105 110
Thr Ser Lys Tyr Arg Pro Leu Thr Leu Gln Gly Asp Ala Trp Cys Ser
115 120 125
Ser Gly Asn Leu Trp Pro Asn Gly Asn Leu Met Ala Thr Gly Gly Thr
130 135 140
Phe Ser Gly Asp Lys Ala Ile Arg Val Ile Ala Asn Asp Asp Pro Asn
145 150 155 160
Gly Asp Phe Thr Thr Lys Ile Gly Ala Leu Ala Asp Thr Arg Trp Tyr
165 170 175
Ser Ser Asn Gln Val Leu Pro Asp Gly Ser Ser Val Val Leu Gly Gly
180 185 190
Arg Asp Ser Tyr Ser Tyr Glu Ile Val Pro Pro Gln Met Glu Phe Lys
195 200 205
Pro Arg Arg Phe Asp Leu Pro Phe Met Gln Gln Thr Thr Glu Pro Pro
210 215 220
Leu Gly Pro Gly Arg Pro Val Glu Asn Asn Leu Tyr Pro Phe Leu Phe
225 230 235 240
Leu Leu Pro Asp Gly Asn Ile Phe Leu Phe Ala Asn Asn Arg Ala Ile
245 250 255
Thr Phe Glu Pro Ala Thr Gly Lys Thr Val Lys Glu Phe Pro Val Leu
260 265 270
Pro Gly Gly Ser Arg Asn Tyr Pro Pro Ser Gly Ser Ala Ala Leu Phe
275 280 285
Pro Leu Lys Leu Thr Ala Asp Asn Ala Pro Val Ile Pro Glu Ile Val
290 295 300
Ile Cys Gly Gly Asn Lys Pro Asn Ala Tyr Glu Leu Val Asp Ala Arg
305 310 315 320
His Val Thr Glu Lys Gln Phe Leu Pro Ala Leu Gln Asp Cys Asn Arg
325 330 335
Ile Gln Pro Met Ala Ala Asp Ala Ala Trp Ile Pro Glu Gln Asn Met
340 345 350
Pro Ser Pro Arg Thr Met Gly Asp Leu Ile His Leu Ala Asn Gly Asp
355 360 365
Met Leu Met Leu Asn Gly Ala Lys Lys Gly Thr Ser Gly Trp Glu Asp
370 375 380
Ala Thr Glu Ala Asn Leu Thr Pro Val Leu Tyr Thr Pro Tyr Lys Pro
385 390 395 400
Met Gly Gln Arg Phe Lys Glu Leu Ser Pro Thr Thr Ile Ala Arg Met
405 410 415
Tyr His Ser Cys Ser Ala Leu Leu Pro Asp Thr Arg Val Leu Val Ala
420 425 430
Gly Ser Asn Met His Gln Phe Tyr Thr Phe Asp Thr Glu Phe Pro Thr
435 440 445
Glu Leu Arg Val Glu Lys Phe Ser Pro Pro Tyr Leu Asp Pro Ala Leu
450 455 460
Glu Pro Glu Arg Thr Gln Ile Asp Pro Ala Asn Thr Asp Val Val Leu
465 470 475 480
Lys Tyr Gly Lys Pro Phe Lys Ile Thr Ala Ala Leu Met Glu Lys Gln
485 490 495
Pro Leu Val Leu Gly Glu Val Lys Val Thr Leu Leu Tyr Pro Pro Phe
500 505 510
Thr Thr His Gly Phe Ser Gln Asn Gln Arg Met Ile Val Pro Ala Ile
515 520 525
Thr Ser Val Glu Asn Gly Val Ile Thr Ala Ile Ala Pro Pro Ser Gly
530 535 540
Glu Ile Ala Pro Pro Gly Tyr Tyr Ile Met Phe Val Ser His Leu Gly
545 550 555 560
Ile Pro Gly Ala Gly Ile Trp Val His Ile Asp
565 570
<210> 3
<211> 571
<212> PRT
<213> 茵陈蒿(A. capillaries)
<400> 3
Ala Pro Asp Ile Thr Asp Gly Gly Asp Lys Pro Gly Pro Leu Ile Asp
1 5 10 15
Asp Gly Gly Gly Asp Lys Pro Val Pro Gly Asn Asn Asp Gly Ala Ser
20 25 30
Asp Tyr Ala Lys Pro Ala Ile Glu Pro Glu Phe Met Gly Glu Trp Val
35 40 45
Ile Asp Asn Pro Asn Ala Gly Val Ala Ala Met Gln Leu Gln Leu Met
50 55 60
Pro Asn Asp Gln Ile Val Trp Phe Asp Thr Thr Ser Leu Gly Ala Ser
65 70 75 80
Gly Tyr Lys Leu Pro Glu Gly Thr Pro Cys Pro Ile Asn Pro Asp Ala
85 90 95
Asn Asn Gln Pro Asp Cys Tyr Ala His Gly Ile Ala Tyr Asp Trp Lys
100 105 110
Thr Ser Lys Tyr Arg Pro Leu Thr Leu Gln Gly Asp Ala Trp Cys Ser
115 120 125
Ser Gly Asn Leu Trp Pro Asn Gly Asn Leu Met Ala Thr Gly Gly Thr
130 135 140
Phe Ser Gly Asp Lys Ala Ile Arg Val Ile Ala Asn Asp Asp Pro Asn
145 150 155 160
Gly Asp Phe Thr Thr Lys Ile Gly Ala Leu Ala Asp Thr Arg Trp Tyr
165 170 175
Ser Ser Asn Gln Val Leu Pro Asp Gly Ser Ala Val Val Leu Gly Gly
180 185 190
Arg Asp Ser Tyr Ser Tyr Glu Ile Val Pro Pro Gln Met Glu Phe Lys
195 200 205
Pro Lys Arg Phe Asp Leu Pro Phe Leu Gln Gln Thr Thr Glu Pro Pro
210 215 220
Leu Gly Pro Gly Arg Pro Val Glu Asn Asn Leu Tyr Pro Phe Gln Phe
225 230 235 240
Leu Leu Pro Asp Gly Asn Ile Phe Leu Phe Ala Asn Asn Arg Ala Ile
245 250 255
Thr Phe Glu Pro Ala Thr Gly Lys Thr Val Lys Glu Phe Pro Val Leu
260 265 270
Pro Gly Gly Ser Arg Asn Tyr Pro Pro Ser Gly Ser Ala Ala Leu Phe
275 280 285
Pro Leu Lys Leu Thr Ala Asp Asn Ala Pro Val Val Pro Glu Ile Val
290 295 300
Ile Cys Gly Gly Asn Gln Pro Asn Ala Tyr Glu Leu Val Asp Ala Arg
305 310 315 320
His Val Thr Glu Lys Lys Phe Leu Pro Ala Leu Gln Asp Cys Asn Arg
325 330 335
Ile Gln Pro Met Ala Ala Asp Ala Ala Trp Ile Pro Glu Gln Asn Met
340 345 350
Pro Ser Pro Arg Thr Met Gly Asp Leu Leu His Leu Ala Asn Gly Asp
355 360 365
Met Leu Met Leu Asn Gly Ala Lys Lys Gly Thr Ser Gly Trp Glu Asp
370 375 380
Ala Thr Glu Ala Asn Leu Thr Pro Val Leu Tyr Thr Pro Tyr Lys Pro
385 390 395 400
Met Gly Gln Arg Phe Lys Glu Leu Ser Pro Thr Thr Ile Ala Arg Met
405 410 415
Tyr His Ser Cys Ser Ala Leu Leu Pro Asp Thr Arg Val Leu Val Ala
420 425 430
Gly Ser Asn Met His Gln Phe Tyr Thr Phe Asp Thr Glu Phe Pro Thr
435 440 445
Glu Leu Arg Val Glu Lys Phe Ser Pro Pro Tyr Leu Asp Pro Ala Leu
450 455 460
Glu Pro Glu Arg Thr Leu Ile Asp Pro Ala Asn Thr Asp Val Val Leu
465 470 475 480
Lys Tyr Gly Lys Pro Phe Lys Ile Thr Ala Ala Leu Met Glu Lys Gln
485 490 495
Pro Leu Val Leu Gly Glu Val Lys Val Thr Leu Leu Tyr Pro Pro Phe
500 505 510
Thr Thr His Gly Phe Ser Gln Asn Gln Arg Met Ile Val Pro Ala Ile
515 520 525
Thr Gly Val Glu Asn Gly Val Ile Thr Ala Val Ala Pro Pro Ser Gly
530 535 540
Glu Ile Ala Pro Pro Gly Tyr Tyr Ile Met Phe Val Ser His Leu Gly
545 550 555 560
Ile Pro Gly Ala Gly Ile Trp Val His Ile Asp
565 570
<210> 4
<211> 571
<212> PRT
<213> 裂叶莲蒿(A. gmelinii)
<400> 4
Ala Pro Asp Ile Thr Asp Gly Gly Asp Lys Pro Gly Pro Glu Ile Asp
1 5 10 15
Asp Gly Gly Gly Asp Lys Pro Val Pro Gly Asn Asn Asp Gly Ala Ser
20 25 30
Asp Tyr Ala Lys Pro Ala Ile Glu Pro Glu Phe Met Gly Ala Trp Val
35 40 45
Ile Asp Asn Pro Asn Ala Gly Val Ala Ala Met Gln Leu Gln Leu Met
50 55 60
Pro Asn Asp Gln Ile Val Trp Phe Asp Thr Thr Ser Leu Gly Pro Ser
65 70 75 80
Gly Tyr Lys Leu Pro Glu Gly Thr Pro Cys Pro Ile Asn Pro Asp Ala
85 90 95
Asn Asn Gln Pro Asp Cys Tyr Ala His Gly Ile Ala Tyr Asp Trp Lys
100 105 110
Thr Ser Lys Tyr Arg Pro Leu Thr Leu Gln Gly Asp Ala Trp Cys Ser
115 120 125
Ser Gly Asn Leu Trp Pro Asn Gly Asn Leu Met Ala Thr Gly Gly Thr
130 135 140
Phe Ser Gly Asp Lys Ala Ile Arg Val Ile Ala Asn Asp Asp Pro Lys
145 150 155 160
Gly Asp Phe Thr Thr Lys Ile Gly Ala Leu Ala Asp Thr Arg Trp Tyr
165 170 175
Ser Ser Asn Gln Val Leu Pro Asp Gly Ser Ser Val Val Leu Gly Gly
180 185 190
Arg Asp Ser Tyr Ser Tyr Glu Ile Val Pro Pro Gln Met Glu Phe Lys
195 200 205
Pro Lys Arg Phe Asp Leu Pro Phe Met Gln Gln Thr Thr Glu Pro Pro
210 215 220
Leu Gly Pro Gly Arg Pro Val Glu Asn Asn Leu Tyr Pro Phe Leu Phe
225 230 235 240
Leu Leu Pro Asp Gly Asn Ile Phe Leu Phe Ala Asn Asn Arg Ala Ile
245 250 255
Thr Phe Glu Pro Ala Thr Gly Lys Thr Val Lys Glu Phe Pro Val Leu
260 265 270
Pro Gly Gly Ser Arg Asn Tyr Pro Pro Ser Gly Ser Ala Ala Leu Phe
275 280 285
Pro Leu Lys Leu Thr Ala Asp Asn Ala Pro Val Ile Pro Glu Ile Val
290 295 300
Ile Cys Gly Gly Asn Gln Pro Asn Ala Tyr Glu Leu Val Asp Ala Arg
305 310 315 320
His Val Thr Glu Lys Gln Phe Leu Pro Ala Leu Gln Asp Cys Asn Arg
325 330 335
Ile Gln Pro Met Ala Ala Asp Ala Ala Trp Ile Pro Glu Gln Asn Met
340 345 350
Pro Ser Pro Arg Thr Met Gly Asp Leu Ile His Leu Ala Asn Gly Asp
355 360 365
Met Leu Met Leu Asn Gly Ala Lys Lys Gly Thr Ser Gly Trp Glu Asp
370 375 380
Ala Thr Glu Ala Asn Leu Thr Pro Val Leu Tyr Thr Pro Tyr Lys Pro
385 390 395 400
Met Gly Gln Arg Phe Lys Glu Leu Thr Pro Thr Thr Ile Ala Arg Met
405 410 415
Tyr His Ser Cys Ser Ala Leu Leu Pro Asp Thr Arg Val Leu Val Ala
420 425 430
Gly Ser Asn Met His Gln Phe Tyr Thr Phe Asp Thr Glu Phe Pro Thr
435 440 445
Glu Leu Arg Val Glu Lys Phe Ser Pro Pro Tyr Leu Asp Pro Ala Leu
450 455 460
Glu Thr Glu Arg Thr Gln Ile Asp Pro Ala Asn Thr Asp Val Val Leu
465 470 475 480
Lys Tyr Gly Lys Pro Phe Lys Ile Thr Ala Ala Leu Met Glu Lys Gln
485 490 495
Pro Leu Val Leu Gly Glu Val Lys Val Thr Leu Leu Tyr Pro Pro Phe
500 505 510
Thr Thr His Gly Phe Ser Gln Asn Gln Arg Met Ile Val Pro Ala Ile
515 520 525
Thr Ser Val Glu Asn Gly Val Ile Thr Ala Val Ala Pro Pro Ser Gly
530 535 540
Gln Ile Ala Pro Pro Gly Tyr Tyr Ile Met Phe Val Ser His Leu Gly
545 550 555 560
Ile Pro Gly Ala Gly Ile Trp Val His Ile Asp
565 570
<210> 5
<211> 571
<212> PRT
<213> 野艾蒿(A. lavandulifolia)
<400> 5
Ala Pro Asp Ile Thr Asp Gly Gly Asp Lys Pro Gly Pro Gln Val Asp
1 5 10 15
Asp Gly Gly Gly Asp Lys Pro Val Pro Gly Asn Asn Asp Gly Ala Ser
20 25 30
Asp Tyr Ala Lys Pro Ala Phe Glu Pro Glu Phe Met Gly Ala Trp Val
35 40 45
Ile Asp Asn Pro Asn Ala Gly Val Ala Ala Met Gln Leu Gln Leu Met
50 55 60
Pro Asn Asp Gln Ile Val Trp Phe Asp Thr Thr Ser Leu Gly Ala Ser
65 70 75 80
Gly Tyr Lys Leu Pro Glu Gly Thr Pro Cys Pro Asn Asn Pro Asp Ala
85 90 95
Asn Asn Gln Pro Asp Cys Tyr Val His Gly Ile Ala Tyr Asp Trp Lys
100 105 110
Thr Ser Lys Tyr Arg Pro Leu Thr Leu Gln Gly Asp Ala Trp Cys Ser
115 120 125
Ser Gly Asn Leu Trp Pro Asn Gly Asn Leu Met Ala Thr Gly Gly Thr
130 135 140
Phe Ser Gly Asp Lys Ala Ile Arg Val Ile Ala Asn Asp Asp Pro Asn
145 150 155 160
Gly Asp Phe Thr Thr Lys Ile Gly Ala Leu Ala Asp Thr Arg Trp Tyr
165 170 175
Ser Ser Asn Gln Val Leu Pro Asp Gly Ser Ser Val Val Leu Gly Gly
180 185 190
Arg Asp Ser Tyr Ser Tyr Glu Ile Val Pro Pro Gln Met Glu Phe Lys
195 200 205
Pro Arg Arg Phe Asp Leu Pro Phe Met Gln Gln Thr Thr Glu Pro Pro
210 215 220
Leu Gly Pro Gly Arg Pro Val Glu Asn Asn Leu Tyr Pro Phe Leu Phe
225 230 235 240
Leu Leu Pro Asp Gly Asn Ile Phe Leu Phe Ala Asn Asn Arg Ala Ile
245 250 255
Thr Phe Glu Pro Ala Thr Gly Lys Thr Val Lys Glu Phe Pro Val Leu
260 265 270
Pro Gly Gly Ser Arg Asn Tyr Pro Pro Ser Gly Ser Ala Ala Leu Phe
275 280 285
Pro Leu Lys Leu Thr Ala Asp Asn Ala Pro Val Ile Pro Glu Ile Val
290 295 300
Ile Cys Gly Gly Asn Lys Pro Asn Ala Tyr Glu Leu Val Asp Ala Arg
305 310 315 320
His Val Thr Glu Lys Gln Phe Leu Pro Ala Leu Gln Asp Cys Asn Arg
325 330 335
Ile Gln Pro Met Ala Ala Asp Ala Ala Trp Ile Pro Glu Gln Asn Met
340 345 350
Pro Ser Pro Arg Thr Met Gly Asp Leu Ile His Leu Ala Asn Gly Asp
355 360 365
Met Leu Met Leu Asn Gly Ala Lys Lys Gly Thr Ser Gly Trp Glu Asp
370 375 380
Ala Thr Glu Ala Asn Leu Thr Pro Val Leu Tyr Thr Pro Tyr Lys Pro
385 390 395 400
Met Gly Gln Arg Phe Lys Glu Leu Ser Pro Thr Thr Ile Ala Arg Met
405 410 415
Tyr His Ser Cys Ser Ala Leu Leu Pro Asp Thr Arg Val Leu Val Ala
420 425 430
Gly Ser Asn Met His Gln Phe Tyr Thr Phe Asp Thr Glu Phe Pro Thr
435 440 445
Glu Leu Arg Val Glu Lys Phe Ser Pro Pro Tyr Leu Asp Pro Ala Leu
450 455 460
Glu Pro Glu Arg Thr Gln Ile Asp Pro Ala Asn Thr Asp Val Val Leu
465 470 475 480
Lys Tyr Gly Lys Pro Phe Lys Ile Thr Ala Ala Leu Met Glu Lys Gln
485 490 495
Pro Leu Val Leu Gly Glu Val Lys Val Thr Leu Leu Tyr Pro Pro Phe
500 505 510
Thr Thr His Gly Phe Ser Gln Asn Gln Arg Met Ile Val Pro Ala Ile
515 520 525
Thr Ser Val Glu Asn Gly Val Ile Thr Ala Ile Ala Pro Pro Ser Gly
530 535 540
Glu Ile Ala Pro Pro Gly Tyr Tyr Ile Leu Phe Val Ser His Leu Gly
545 550 555 560
Ile Pro Gly Ala Gly Ile Trp Val His Ile Asp
565 570
<210> 6
<211> 572
<212> PRT
<213> 大籽蒿(A. sieversiana)
<400> 6
Ala Pro Asp Ile Thr Asp Gly Ala Gly Asp Lys Pro Gly Pro Gln Val
1 5 10 15
Asp Asp Gly Gly Gly Asp Lys Pro Val Pro Gly Asn Asn Asp Gly Ala
20 25 30
Ser Asp Tyr Ala Lys Pro Ala Ile Glu Ser Glu Phe Met Gly Gln Trp
35 40 45
Val Ile Asp Asn Pro Asn Ala Gly Val Ala Ala Met Gln Leu Gln Leu
50 55 60
Met Pro Asn Asp Gln Ile Val Trp Phe Asp Thr Thr Ser Leu Gly Ala
65 70 75 80
Ser Gly Tyr Lys Leu Pro Glu Gly Thr Pro Cys Pro Ile Asn Pro Asp
85 90 95
Ala Asn Asn Gln Pro Asp Cys Tyr Val His Gly Ile Ala Tyr Asp Trp
100 105 110
Lys Thr Ser Lys Tyr Arg Pro Leu Thr Leu Gln Gly Asp Ala Trp Cys
115 120 125
Ser Ser Gly Asn Leu Trp Pro Asn Gly Asn Leu Met Ala Thr Gly Gly
130 135 140
Thr Phe Ser Gly Asp Lys Ala Ile Arg Val Ile Pro Asn Asp Asp Pro
145 150 155 160
Asn Gly Asp Phe Thr Thr Lys Ile Gly Ala Leu Ala Asp Thr Arg Trp
165 170 175
Tyr Ser Ser Asn Gln Val Leu Pro Asp Gly Ser Ser Val Val Leu Gly
180 185 190
Gly Arg Asp Ser Tyr Ser Tyr Glu Ile Val Pro Pro Gln Met Glu Phe
195 200 205
Lys Pro Lys Arg Phe Asp Leu Pro Phe Met Gln Gln Thr Thr Glu Pro
210 215 220
Pro Leu Gly Pro Gly Arg Pro Val Glu Asn Asn Leu Tyr Pro Phe Leu
225 230 235 240
Phe Leu Leu Pro Asp Gly Asn Ile Phe Leu Phe Ala Asn Asn Arg Ala
245 250 255
Ile Thr Phe Glu Pro Ala Thr Gly Lys Thr Val Lys Glu Phe Pro Val
260 265 270
Leu Pro Gly Gly Ser Arg Asn Tyr Pro Pro Ser Gly Ser Ala Ala Leu
275 280 285
Phe Pro Leu Lys Leu Thr Ala Asp Asn Ala Pro Val Val Pro Glu Ile
290 295 300
Val Ile Cys Gly Gly Asn Gln Pro Asn Ala Tyr Glu Leu Val Asp Ala
305 310 315 320
Arg His Val Thr Glu Lys Gln Phe Leu Pro Ala Leu Gln Asp Cys Asn
325 330 335
Arg Ile Lys Pro Met Ala Ala Asp Ala Ala Trp Ile Pro Glu Gln Asn
340 345 350
Met Pro Ser Pro Arg Thr Met Gly Asp Leu Ile His Leu Ala Asn Gly
355 360 365
Asp Met Leu Met Leu Asn Gly Ala Lys Lys Gly Thr Ser Gly Trp Glu
370 375 380
Asp Ala Thr Glu Pro Asn Leu Thr Pro Val Leu Tyr Thr Pro Tyr Lys
385 390 395 400
Pro Met Gly Gln Arg Phe Lys Glu Leu Arg Pro Thr Thr Ile Ala Arg
405 410 415
Met Tyr His Ser Cys Ser Ala Leu Leu Pro Asp Thr Arg Val Leu Val
420 425 430
Ala Gly Ser Asn Met His Gln Phe Tyr Thr Phe Asp Thr Glu Phe Pro
435 440 445
Thr Glu Leu Arg Val Glu Lys Phe Ser Pro Pro Tyr Leu Asp Pro Ala
450 455 460
Leu Glu Leu Glu Arg Ser Gln Ile Asp Pro Ala Asn Thr Asp Val Val
465 470 475 480
Leu Lys Tyr Gly Lys Pro Phe Lys Ile Thr Ala Gly Leu Met Glu Lys
485 490 495
Gln Pro Leu Val Leu Gly Glu Val Lys Val Thr Leu Leu Tyr Pro Pro
500 505 510
Phe Thr Thr His Gly Phe Ser Gln Asn Gln Arg Met Ile Val Pro Ala
515 520 525
Ile Thr Ser Val Glu Asn Gly Val Ile Thr Ala Ile Ala Pro Pro Ser
530 535 540
Gly Glu Ile Ala Pro Pro Gly Tyr Tyr Ile Met Phe Val Ser His Leu
545 550 555 560
Gly Ile Pro Gly Ala Gly Ile Trp Val His Ile Asp
565 570
<210> 7
<211> 571
<212> PRT
<213> 北艾(A. vulgaris)
<400> 7
Ala Pro Asp Ile Thr Asp Gly Gly Asp Lys Pro Gly Pro Gln Val Asp
1 5 10 15
Asp Gly Gly Gly Asp Lys Pro Val Pro Gly Asn Asn Asp Gly Ala Ser
20 25 30
Asp Tyr Ala Lys Pro Ala Phe Glu Pro Glu Phe Met Gly Ala Trp Val
35 40 45
Ile Asp Asn Pro Asn Ala Gly Val Ala Ala Met Gln Leu Gln Leu Met
50 55 60
Pro Asn Asp Gln Ile Val Trp Phe Asp Thr Thr Ser Leu Gly Ala Ser
65 70 75 80
Gly Tyr Lys Leu Pro Glu Gly Thr Pro Cys Pro Asn Asn Pro Asp Ala
85 90 95
Asn Asn Gln Pro Asp Cys Tyr Val His Gly Ile Ala Tyr Asp Trp Lys
100 105 110
Thr Ser Lys Tyr Arg Pro Leu Thr Leu Gln Gly Asp Ala Trp Cys Ser
115 120 125
Ser Gly Asn Leu Trp Pro Asn Gly Asn Leu Met Ala Thr Gly Gly Thr
130 135 140
Phe Ser Gly Asp Lys Ala Ile Arg Val Ile Ala Asn Asp Asp Pro Asn
145 150 155 160
Gly Asp Phe Thr Thr Lys Ile Gly Ala Leu Ala Asp Thr Arg Trp Tyr
165 170 175
Ser Ser Asn Gln Val Leu Pro Asp Gly Ser Ser Val Val Leu Gly Gly
180 185 190
Arg Asp Ser Tyr Ser Tyr Glu Ile Val Pro Pro Gln Met Glu Phe Lys
195 200 205
Pro Arg Arg Phe Asp Leu Pro Phe Met Gln Gln Thr Thr Glu Pro Pro
210 215 220
Leu Gly Pro Gly Arg Pro Val Glu Asn Asn Leu Tyr Pro Phe Leu Phe
225 230 235 240
Leu Leu Pro Asp Gly Asn Ile Phe Leu Phe Ala Asn Asn Arg Ala Ile
245 250 255
Thr Phe Glu Pro Ala Thr Gly Lys Thr Val Lys Glu Phe Pro Val Leu
260 265 270
Pro Gly Gly Ser Arg Asn Tyr Pro Pro Ser Gly Ser Ala Ala Leu Phe
275 280 285
Pro Leu Lys Leu Thr Ala Asp Asn Ala Pro Val Ile Pro Glu Ile Val
290 295 300
Ile Cys Gly Gly Asn Lys Pro Asn Ala Tyr Glu Leu Val Asp Ala Arg
305 310 315 320
His Val Thr Glu Lys Gln Phe Leu Pro Ala Leu Gln Asp Cys Asn Arg
325 330 335
Ile Gln Pro Met Ala Ala Asp Ala Ala Trp Ile Pro Glu Gln Asn Met
340 345 350
Pro Ser Pro Arg Thr Met Gly Asp Leu Ile His Leu Ala Asn Gly Asp
355 360 365
Met Leu Met Leu Asn Gly Ala Lys Lys Gly Thr Ser Gly Trp Glu Asp
370 375 380
Ala Thr Glu Ala Asn Leu Thr Pro Val Leu Tyr Thr Pro Tyr Lys Pro
385 390 395 400
Met Gly Gln Arg Phe Lys Glu Leu Ser Pro Thr Thr Ile Ala Arg Met
405 410 415
Tyr His Ser Cys Ser Ala Leu Leu Pro Asp Thr Arg Val Leu Val Ala
420 425 430
Gly Ser Asn Met His Gln Phe Tyr Thr Phe Asp Thr Glu Phe Pro Thr
435 440 445
Glu Leu Arg Val Glu Lys Phe Ser Pro Pro Tyr Leu Asp Pro Ala Leu
450 455 460
Glu Pro Glu Arg Thr Gln Ile Asp Pro Ala Asn Thr Asp Val Val Leu
465 470 475 480
Lys Tyr Gly Lys Pro Phe Lys Ile Thr Ala Ala Leu Met Glu Lys Gln
485 490 495
Pro Leu Val Leu Gly Glu Val Lys Val Thr Leu Leu Tyr Pro Pro Phe
500 505 510
Thr Thr His Gly Phe Ser Gln Asn Gln Arg Met Ile Val Pro Ala Ile
515 520 525
Thr Ser Val Glu Asn Gly Val Ile Thr Ala Ile Ala Pro Pro Ser Gly
530 535 540
Glu Ile Ala Pro Pro Gly Tyr Tyr Ile Met Phe Leu Ser His Leu Gly
545 550 555 560
Ile Pro Gly Ala Gly Ile Trp Val His Ile Asp
565 570
<210> 8
<211> 1713
<212> DNA
<213> 黄花蒿(A. annua)
<400> 8
ccggatataa ccgatggtgg cgacaaacct gggcccgaaa tcgatgatgg tggtggtgac 60
aaacctgtac ccggaaacaa cgatggtgca tcagattatg caaagccagc attcgagcct 120
gaatttatgg gtgcatgggt aattgacaat cctaacgctg gggttgctgc catgcaactc 180
cagttaatgc ctaatgatca gatcgtatgg tttgatacca cgtcactcgg ggcttcagga 240
tataaattac ctgagggaac tccttgccct attaacccag atgcaaataa tcagcctgat 300
tgttatgctc atggcatcgc ttatgactgg aagacctcaa aatacaggcc gcttacgtta 360
caaggtgatg catggtgctc gtccggaaac ttgtggccaa atggtaacct aatggctacg 420
ggaggtacct tcagtggtga taaagctatt cgagtgatcg ctaacgatga tcctaaagga 480
gactttacca ccaaaattgg tgctcttgct gatactagat ggtactcatc taaccaggtc 540
ttgccagacg gaagttcagt cgtgttggga ggcagggatt cctacagcta cgaaatcgta 600
ccaccacaaa tggaatttaa acccaagaga tttgatctcc ctttcatgca acaaacaact 660
gaacctcctt taggaccagg aagaccagtg gagaacaacc tgtacccgtt ccttttcctc 720
cttcctgatg gaaacatttt cctctttgct aacaatcgag ccatcacctt cgaaccagct 780
actggtaaaa ccgttaagga gttccccgtg ttacctggtg gctctcgtaa ctatccacct 840
tcaggatcgg ccgctctttt cccattgaag ctcactgctg ataatgcccc ggtcattcct 900
gagattgtaa tttgcggtgg taaccaacca aatgcatatg aactcgttga cgcaaggcat 960
gtcaccgaaa agcagttcct acctgccctc caagattgta atagaatcca acccatggca 1020
gctgatgcag cgtggattcc cgaacaaaac atgccatcac caagaacaat gggtgatttg 1080
atacatcttg ccaatggtga tatgttgatg cttaatggtg cgaaaaaagg tacatcaggt 1140
tgggaagatg cgacagaggc taaccttaca cctgttttat atacaccata caaacctatg 1200
ggacaaagat ttaaagaatt gactccaaca acaattgcta gaatgtatca ttcttgctcg 1260
gctttattgc ccgacactag agtcttggtc gcgggtagca acatgcacca gttctacacg 1320
ttcgacacgg agtttcctac agaattgagg gtagagaaat tctcaccacc ttacttggac 1380
ccagccctag aaacggaaag aacccagatc gatccagcta atacagatgc ggtcttgaag 1440
tacggaaaac catttaagat tacagctgct cttatggaaa aacagcccct ggtgcttggt 1500
gaggttaagg taaccttgtt atatccacca ttcaccactc atggtttctc tcaaaaccaa 1560
aggatgattg taccggcaat aaccagtgtt cagaacggtg taatcacagc cgttgcacct 1620
ccttctggac agattgcacc accaggatac tacattatgt ttgtgagtca ccttggtatt 1680
cctggtgcgg gtatttgggt tcacatcgac taa 1713
<210> 9
<211> 1713
<212> DNA
<213> 艾蒿(A. arygi)
<400> 9
ccagatataa ccgatggtgg cgacaaacct gggcctcaag tcgatgatgg tggtggtgac 60
aaacctgtac ccggaaacaa cgatggtgca tctgattatg caaagccagc attcgagcct 120
gaatttatgg gtgcatgggt aattgacaat cctaacgctg gtgttgctgc catgcagctc 180
cagttgatgc ctaatgatca aatcgtatgg tttgatacca cgtcacttgg ggcttcgggg 240
tataaattac ctgagggaac tccttgccct aataacccag atgcaaataa tcagcctgat 300
tgttatgttc atggcatcgc ttatgactgg aagacctcaa aatacaggcc gcttacgtta 360
caaggtgatg catggtgctc gtccggaaac ttgtggccaa atggtaacct aatggcgaca 420
ggcggcacct tcagtggtga taaagctatt cgagtgatcg ctaacgatga tcctaatgga 480
gactttacca ccaaaattgg tgctcttgct gatactagat ggtactcatc taaccaggtc 540
ttgcctgacg gaagttcagt cgtgttggga ggcagggatt cctacagcta cgaaatcgta 600
ccaccacaaa tggaatttaa accccggagg tttgatctcc ctttcatgca acaaacaact 660
gaacctcctt taggaccagg aagaccggtc gagaacaacc tgtacccatt ccttttcctc 720
cttcctgatg gaaacatttt cctctttgct aacaatcgtg ccatcacctt cgaaccagct 780
actggaaaaa ctgttaagga gttccccgtg ttacctggtg gctctcgcaa ctatccacct 840
tcaggatcgg ccgctctttt cccattgaaa ctcacggctg ataatgcccc ggtcattcct 900
gagattgtaa tttgcggtgg taacaaacca aatgcatatg aactcgttga cgcaaggcat 960
gtcaccgaaa agcagttcct acctgccctc caagattgta atagaatcca acccatggca 1020
gctgatgcag cgtggatccc cgaacaaaac atgccatcac caagaaccat gggtgatttg 1080
atacatcttg ccaatggtga tatgttgatg cttaatggtg cgaaaaaagg tacatcaggt 1140
tgggaagatg cgacagaggc taaccttaca cctgttttat atacaccata caaacctatg 1200
ggacaaagat ttaaagaatt gagtccaaca acaattgcta gaatgtatca ttcttgctcg 1260
gctttattgc ccgacactag agtcttggtc gcgggtagca acatgcacca gttctacacg 1320
tttgacaccg agtttcctac agaattgagg gttgagaaat tctcaccacc ttacttggac 1380
ccagccctag aaccagaaag aacccagatc gatccagcta atacggatgt ggtcctgaag 1440
tacggaaaac cattcaagat cacagctgct cttatggaaa aacaacccct ggtgcttggt 1500
gaggttaagg taaccttgtt atatccacca ttcaccactc atggtttctc tcaaaaccaa 1560
aggatgattg taccggcaat aaccagtgtt gagaacggtg taatcacagc cattgcaccg 1620
ccttctggag agattgctcc accaggatac tacattatgt ttgtgagtca cctcggtatt 1680
cctggtgcgg gtatttgggt tcacatcgac taa 1713
<210> 10
<211> 1713
<212> DNA
<213> 茵陈蒿(A. capillaries)
<400> 10
ccagatataa ccgatggtgg cgacaaacct gggcctttaa tcgatgatgg tggtggtgac 60
aaacctgtac ctggaaacaa tgatggtgca tctgattatg caaagcctgc gatcgagcct 120
gaatttatgg gcgaatgggt aattgacaat cctaatgctg gtgttgctgc catgcaactc 180
cagttgatgc ctaatgatca gatcgtatgg tttgatacca cgtcacttgg ggcttcggga 240
tataaattac ctgagggaac tccttgccct attaacccag atgcaaataa ccagcctgat 300
tgttatgctc atggcatcgc ttatgactgg aagacctcaa aatacaggcc gcttacgttg 360
caaggtgatg catggtgctc gtccggaaac ttgtggccaa atggtaacct aatggctacg 420
ggaggtacct tcagtggtga taaagctatt cgagtgatcg ctaatgatga tcctaatgga 480
gactttacca ccaaaattgg tgctcttgct gatactagat ggtactcatc taaccaggtc 540
ttgccagatg gaagtgcagt tgtgttggga ggcagggatt cctacagcta cgaaatcgta 600
ccaccacaaa tggaatttaa acccaagaga tttgatctcc ctttcttgca acaaacaact 660
gaacctccct taggaccagg aagaccggtc gagaacaacc tgtacccgtt ccaattcctt 720
cttcctgatg gaaacatttt cctcttcgct aacaatcgtg ccatcacctt cgaaccagct 780
actggaaaaa ccgttaagga gttccccgtg ttacctggtg gctctcgcaa ctatccacct 840
tcaggatcgg ccgctctttt cccattgaag ctcacggctg ataatgcccc ggtcgttcct 900
gagattgtaa tttgcggtgg taaccagcca aatgcatatg aactcgttga cgcaaggcat 960
gtcaccgaaa agaagttcct acctgccctc caagattgta atagaatcca acccatggca 1020
gctgatgcag cgtggatccc cgaacaaaac atgccatcac caagaaccat gggtgatttg 1080
ctacatcttg ccaatggtga tatgctgatg cttaatggtg cgaaaaaagg tacatcaggt 1140
tgggaagatg cgacagaggc taaccttaca cctgttttat atacaccata caaacctatg 1200
ggacaaagat ttaaagaatt gagtccaaca acaattgcta gaatgtatca ttcttgctcg 1260
gctttattgc ccgacactag agtcttggtc gcaggtagca acatgcacca gttctacaca 1320
ttcgacactg agtttcctac agaattgagg gttgagaaat tctcaccacc ttacttggac 1380
ccagccctag aaccagaaag aaccctgatc gatccagcta atacggatgt ggtcttgaag 1440
tacggaaaac catttaagat tacagctgct cttatggaaa aacaacccct ggtgcttggt 1500
gaggttaagg taaccttgtt atatccacca ttcaccactc atggtttctc tcaaaaccaa 1560
aggatgattg taccagcaat aaccggtgtt gagaacggtg taatcacagc cgttgcacct 1620
ccttctggag agattgctcc accaggatac tacattatgt ttgtgagtca cctcggtatt 1680
cctggtgcgg gtatttgggt tcacatcgac taa 1713
<210> 11
<211> 1713
<212> DNA
<213> 裂叶莲蒿(A. gmelinii)
<400> 11
ccggatataa ccgatggtgg cgacaaacct gggcccgaaa tcgatgatgg tggtggtgac 60
aaacctgtac ccggaaacaa cgatggtgca tctgattatg caaagccagc aatcgagcct 120
gaatttatgg gtgcatgggt aattgacaat cctaacgctg gggttgctgc catgcaactc 180
cagttgatgc ctaatgatca gatcgtatgg tttgatacca cgtcacttgg gccttcagga 240
tataaattac ctgagggaac tccttgccct attaacccag atgcaaataa tcagcctgat 300
tgttatgctc atggcatcgc ttatgactgg aagacctcaa aatacaggcc gcttacgtta 360
caaggtgatg catggtgctc gtccggaaac ttgtggccaa atggtaacct aatggctacg 420
ggaggtacct tcagtggtga taaagctatt cgagtgatcg ctaacgatga tcctaaagga 480
gactttacca ccaaaattgg tgctcttgct gatactagat ggtactcatc taaccaggtc 540
ttgccagatg gaagttcagt cgtgttggga ggcagggatt cctacagcta cgaaatcgta 600
ccaccacaaa tggaatttaa acccaagaga tttgatctcc ctttcatgca acaaacaact 660
gaacctcctt taggaccagg aagaccggtg gagaacaacc tgtacccgtt ccttttcctc 720
cttcctgatg gaaacatttt cctctttgct aacaatcgag ccatcacctt cgaaccagct 780
actggtaaaa ccgttaagga gttccccgtg ttacctggtg gctctcgcaa ctatccacct 840
tcaggatcgg ccgctctttt cccattgaag ctcacggctg ataatgcccc ggtcattcct 900
gagattgtaa tttgcggtgg taaccaacca aatgcatatg aactcgttga cgcaaggcat 960
gtcactgaaa agcagttcct acctgccctc caagattgta atagaatcca acccatggca 1020
gctgatgcag cgtggattcc cgaacaaaac atgccatcac caagaaccat gggtgatttg 1080
atacatcttg ccaatggtga tatgttgatg cttaatggtg cgaaaaaagg tacatcaggt 1140
tgggaagatg cgacagaggc taaccttaca cctgttttat atacaccata caaacctatg 1200
ggacaaagat ttaaagaatt gactccaaca acaattgcta gaatgtatca ttcttgctcg 1260
gctttattgc ccgacactag agtcttggtc gcgggtagca acatgcacca gttctacacg 1320
ttcgacacgg agtttcctac agaattgagg gttgagaaat tctcaccacc ttacttggac 1380
ccagccctag aaacggaaag aacccagatc gatccagcta atacagatgt ggtcttgaag 1440
tacggaaaac cgtttaagat tacagctgct cttatggaaa aacagcccct ggtgcttggt 1500
gaggttaagg taaccttgtt atatccacca ttcaccactc atggtttctc tcaaaaccaa 1560
aggatgattg taccggcaat aaccagtgtt gagaacggtg taatcacagc cgtcgcacct 1620
ccttctggac agattgctcc accaggatac tacattatgt ttgtgagtca ccttggtatt 1680
cctggtgcgg gtatttgggt tcacatcgac taa 1713
<210> 12
<211> 1713
<212> DNA
<213> 野艾蒿(A. lavandulifolia)
<400> 12
ccagatataa ccgatggtgg cgacaaacct gggcctcaag tcgatgatgg tggtggtgac 60
aaacctgtac ccggaaacaa cgatggtgca tctgattatg caaagccagc attcgagcct 120
gaatttatgg gtgcatgggt aatcgacaat cctaacgctg gtgttgctgc catgcaactc 180
cagttgatgc ctaatgatca aatcgtatgg tttgatacca catcacttgg ggcttcgggg 240
tataaattac ctgagggaac tccttgccct aataacccag atgcaaataa tcagcctgat 300
tgttatgttc atggcatcgc ttatgactgg aagacctcaa aatacaggcc gcttacgtta 360
caaggtgatg catggtgctc gtccggaaac ttgtggccaa atggtaacct aatggcgaca 420
ggcggcacct tcagtggtga taaagctatt cgagtgatcg ctaacgatga tcctaatgga 480
gactttacca ccaaaattgg tgcacttgct gatactagat ggtactcatc taaccaggtc 540
ttgcctgacg gaagttcagt cgtgttggga ggcagggatt cctacagcta cgaaatcgta 600
ccaccacaaa tggaatttaa accccggagg tttgatctcc ctttcatgca acaaacaact 660
gaacctcctt taggaccagg aagaccggtg gagaacaacc tgtacccatt ccttttcctc 720
cttcctgatg gaaacatttt cctctttgct aacaatcgtg ccatcacctt cgaacctgct 780
actggaaaaa ctgttaagga gttccccgtg ttacctggtg gctctcgcaa ctatccacct 840
tcaggatcgg ccgctctttt cccattgaaa ctcacggctg ataatgcccc ggtcattcct 900
gagattgtaa tttgcggtgg taacaaacca aatgcatatg aactcgttga cgcaaggcat 960
gtcaccgaaa agcagttcct acctgccctc caagattgta atagaatcca acccatggca 1020
gctgatgcag cgtggatccc cgaacaaaac atgccatcac caagaaccat gggtgatttg 1080
atacatcttg ccaatggtga tatgttgatg cttaatggtg cgaaaaaagg tacatcaggt 1140
tgggaagatg cgacagaggc taaccttaca cctgttttat atacaccata caaacctatg 1200
ggacaaagat ttaaagaatt gagtccaaca acaattgcta gaatgtatca ttcttgctca 1260
gctttattgc ccgacactag agtcttggtc gcgggtagca acatgcatca gttctacacg 1320
tttgacaccg agtttcctac agaattgagg gttgagaaat tctcaccacc ttacttggat 1380
ccagccctag aaccagaaag aacccagatc gatccagcta atacggatgt ggtcttgaag 1440
tacggaaaac cattcaagat cacagctgct cttatggaaa aacaacccct ggtgcttggt 1500
gaggttaaag taaccttgtt atatccacca ttcaccactc atggtttctc tcaaaaccaa 1560
aggatgattg taccggcaat aaccagtgtt gagaacggtg taatcacagc cattgcaccg 1620
ccttctggag agattgctcc accaggatac tacattttgt ttgtgagtca cctcggtatt 1680
cctggtgcgg gtatttgggt tcacatcgac taa 1713
<210> 13
<211> 1716
<212> DNA
<213> 大籽蒿(A. sieversiana)
<400> 13
ccagatataa ccgatggtgc aggtgacaaa ccagggcccc aagtcgatga tggtggtggt 60
gacaaacctg tacccggaaa caacgatggt gcatctgatt acgcaaagcc agcaatcgag 120
tctgaattta tgggtcaatg ggtaattgac aatcctaacg ctggtgttgc tgccatgcaa 180
ctccagttga tgcctaatga tcagatcgta tggtttgata ccacgtcact tggggcttcg 240
ggatataaat tacctgaggg aactccttgc cctattaacc cagatgcaaa taaccagcct 300
gattgttatg ttcatggcat cgcttatgac tggaagacct caaaatacag gccgcttacg 360
ttacaaggtg atgcatggtg ctcgtcagga aacttgtggc caaatggtaa cctaatggct 420
acgggaggta ccttcagtgg tgataaagct attcgagtga tccctaacga tgatcctaat 480
ggagacttta ccaccaaaat tggtgctctt gctgatacaa gatggtattc atctaaccag 540
gtcttgccag acggaagttc agtcgtgttg ggaggcaggg attcctacag ctacgaaatc 600
gtaccaccac aaatggaatt caaacccaag agatttgatc tcccgttcat gcaacaaaca 660
actgaacctc ctttaggacc aggaagaccg gtcgagaaca acctgtaccc gttccttttc 720
ctccttcctg atggaaacat tttcctcttt gctaacaatc gtgccatcac cttcgaacca 780
gctacaggaa aaaccgtcaa ggagttcccc gtgttacctg gtggctctcg caactatcca 840
ccttcaggat cggcagctct tttcccattg aagctcacgg ctgataatgc cccggtcgtt 900
cctgagattg taatttgcgg tggtaaccaa ccaaatgcat atgaactcgt tgacgcaagg 960
catgtcaccg aaaagcagtt cctgcctgcc ctccaagatt gtaatagaat caaacccatg 1020
gcagctgatg cagcgtggat ccccgaacaa aacatgccat caccaagaac catgggtgat 1080
ttgatacatc ttgccaatgg tgatatgttg atgcttaatg gtgcgaaaaa aggtacttca 1140
ggttgggaag atgcgacaga gcctaaccta acacctgttt tatatacacc atacaaacct 1200
atgggacaaa gatttaaaga actgagacca acaacaattg ctagaatgta tcattcttgc 1260
tcggctttat tgcccgacac tagagtcttg gtcgcgggta gcaacatgca ccagttctat 1320
acgtttgaca ctgagtttcc tacagaattg agggttgaga aattctcacc accttacttg 1380
gacccagccc tagaactgga aagatcccag atcgatccag ctaatacgga tgttgtctta 1440
aagtacggaa aaccatttaa gattacagct ggtcttatgg aaaaacaacc cctggtgctt 1500
ggtgaggtta aggtaacctt gctatatcca ccattcacca ctcatggttt ctctcaaaac 1560
caaaggatga ttgttccggc aataaccagt gttgagaatg gtgtaatcac agccattgca 1620
cctccttctg gagagattgc tccaccagga tactacatta tgtttgtgag tcacctcggt 1680
attcctggtg cgggtatttg ggttcacatc gactaa 1716
<210> 14
<211> 1713
<212> DNA
<213> 北艾(A. vulgaris)
<400> 14
ccagatataa ccgatggtgg cgataaacct gggcctcaag tcgatgatgg tggtggtgac 60
aaacctgtac ccggaaacaa cgatggtgca tctgattatg caaagccagc attcgagcct 120
gaatttatgg gtgcatgggt aatcgacaat cctaacgctg gtgttgctgc catgcaactc 180
cagttgatgc ctaatgatca aatcgtatgg tttgatacca catcacttgg ggcttcgggg 240
tataaattac ctgagggaac tccttgccct aataacccag atgcaaataa tcagcctgat 300
tgttatgttc atggcatcgc ttatgactgg aagacctcaa aatacaggcc gcttacgtta 360
caaggtgatg catggtgctc gtccggaaac ttgtggccaa atggtaacct aatggcgaca 420
ggcggcacct tcagtggtga taaagctatt cgagtgatcg ctaacgatga tcctaatgga 480
gactttacca ccaaaattgg tgcacttgct gatactagat ggtactcatc taaccaggtc 540
ttgcctgacg gaagttcagt cgtgttggga ggcagggatt cctacagcta cgaaatcgta 600
ccaccacaaa tggaatttaa accccggagg tttgatctcc ctttcatgca acaaacaact 660
gaacctcctt taggaccagg aagaccggtc gagaacaacc tgtacccatt ccttttcctc 720
cttcctgatg gaaacatttt cctctttgct aacaatcgtg ccatcacctt cgaacctgct 780
actggaaaaa ctgttaagga gttccccgtg ttacctggtg gctctcgcaa ctatccacct 840
tcaggatcgg ccgctctttt cccattgaaa ctcacggctg ataatgcccc ggtcattcct 900
gagattgtaa tttgcggtgg taacaaacca aatgcatatg aactcgttga cgcaaggcat 960
gtcaccgaaa agcagttcct acctgccctc caagattgta atagaatcca acccatggca 1020
gctgatgcag cgtggatccc cgaacaaaac atgccatcac caagaaccat gggtgatttg 1080
atacatcttg ccaatggtga tatgttgatg cttaatggtg cgaaaaaagg tacatcaggt 1140
tgggaagatg cgacagaggc taaccttaca cctgttttat atacaccata caaacctatg 1200
ggacaaagat ttaaagaatt gagtccaaca acaattgcta gaatgtatca ttcttgctcg 1260
gctttattgc ccgacactag agtcttggtc gcgggtagca acatgcatca gttctacacg 1320
tttgacaccg agtttcctac agaattgagg gttgagaaat tctcaccacc ttacttggac 1380
ccagccctag aaccagaaag aacccagatc gatccagcta atacggatgt ggtcttgaag 1440
tacggaaaac cattcaagat cacagctgct cttatggaaa aacaacccct ggtgcttggt 1500
gaggttaagg taaccttgtt atatccacca ttcaccactc atggtttctc tcaaaaccaa 1560
aggatgattg taccggcaat aaccagtgtt gagaacggtg taatcacagc cattgcaccg 1620
ccttcgggag agattgctcc accaggatac tacattatgt ttctgagtca cctcggtatt 1680
cctggtgcgg gtatttgggt tcacatcgac taa 1713
<210> 15
<211> 21
<212> DNA
<213> 人工序列
<400> 15
atggcttcat caattaaaac a 21
<210> 16
<211> 20
<212> DNA
<213> 人工序列
<400> 16
taacccaagt gtagctgatt 20
Claims (9)
1.一种蒿属植物花粉的过敏原,其特征在于,它为蒿属植物的半乳糖氧化酶,氨基酸序列为SEQ ID No.1、SEQ ID No.2、SEQ ID No.3、SEQ ID No.4、SEQ ID No.5、SEQ ID No.6或SEQ ID No.7。
2.编码如权利要求1所述过敏原的DNA分子,其特征在于,核苷酸序列为SEQ ID No.8、SEQ ID No.9、SEQ ID No.10、SEQ ID No.11、SEQ ID No.12、SEQ ID No.13或SEQ IDNo.14。
3.包含如权利要求2所述DNA分子的重组表达载体。
4.包含如权利要求2所述DNA分子或包含如权利要求3所述重组表达载体的宿主生物体。
5.如权利要求1所述过敏原或如权利要求2所述DNA分子在制备治疗或预防过敏症的药物中的应用。
6.一种用于治疗或预防过敏症的药物,其特征在于,包含如权利要求1所述过敏原或如权利要求2所述DNA分子中的至少一种。
7.如权利要求6所述的药物,其特征在于,所述药物为疫苗。
8.如权利要求1所述过敏原或如权利要求2所述DNA分子在制备过敏原检测的试剂盒中的应用。
9.一种用于过敏原检测的试剂盒,其特征在于,包含如权利要求1所述过敏原或如权利要求2所述DNA分子中的至少一种。
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CN109061184A (zh) * | 2018-08-10 | 2018-12-21 | 杭州艾乐吉生物科技有限公司 | 一种蒿属植物花粉过敏原组合、应用及试剂盒 |
CN112710837A (zh) * | 2020-11-03 | 2021-04-27 | 浙江大学 | 一种定量蒿属植物花粉中nsLTP过敏原的方法 |
CN112710837B (zh) * | 2020-11-03 | 2022-05-17 | 浙江大学 | 一种定量蒿属植物花粉中nsLTP过敏原的方法 |
CN112725320A (zh) * | 2021-01-06 | 2021-04-30 | 中国医学科学院北京协和医院 | 黄花蒿花粉果胶裂合酶及其应用 |
CN116554288A (zh) * | 2022-01-29 | 2023-08-08 | 浙江大学 | 一种芒果果实的过敏原及其应用 |
CN114685636A (zh) * | 2022-03-04 | 2022-07-01 | 首都医科大学附属北京同仁医院 | 蒿属植物花粉过敏原、应用及试剂盒 |
CN114685636B (zh) * | 2022-03-04 | 2024-03-01 | 首都医科大学附属北京同仁医院 | 蒿属植物花粉过敏原、应用及试剂盒 |
CN114933642A (zh) * | 2022-06-30 | 2022-08-23 | 吉林大学中日联谊医院 | Art v 1重组蛋白及其应用 |
CN114933642B (zh) * | 2022-06-30 | 2023-09-12 | 吉林大学中日联谊医院 | Art v 1重组蛋白及其应用 |
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