本发明涉及生物技术领域,具体涉及飞蝗粘蛋白mucin17、其编码基因及其应用。
背景技术:
飞蝗是一种洲际性的农业害虫,主要分布于亚、欧、非、澳四大洲,我国大部分地区都有分布。其主要威胁玉米、谷子等禾本科农作物,在饥饿情况下,还取食大豆、白菜及向日葵等。飞蝗具有跳跃性以及功能强大的前后翅,具有远距离、大规模迁移的生理功能,使其活动空间从平面向立体扩展,行动半径大大提高,从而在觅食、求偶、避敌和扩大分布效率等方面都得到大幅度提高。因此,蝗灾一旦爆发,将会对我国的粮食安全和农业增收造成非常严重的影响。目前对其防治主要依靠化学防治,然而长期施用化学杀虫剂导致一系列问题,如环境污染,抗药性产生和对非靶标生物的危害等。因此,开发新型、可替代化学防治的方法变得极其紧迫。
粘蛋白(mucin)是一种相对分子质量较高的糖蛋白家族,是人类等哺乳动物上皮细胞产生的粘液中的主要成分,广泛地分布于消化道,对消化道粘膜起保护和润滑作用。目前已发现21种粘蛋白家族成员,可分为膜结合型和分泌型两个亚家族。正常生理状况下mucin起着润滑、保护、屏障等作用,参与细胞黏附、信号转导、细胞-细胞间的相互作用,异常表达的mucin参与调控肿瘤细胞的分化、增殖、浸润和转移以及化疗药物抵抗等过程。在昆虫中,肠道粘蛋白(intestinalmucin)主要构成围食膜的成分,如烟草天蛾、棉铃虫等。在脊椎动物和无脊椎动物中,mucin蛋白是一类高度o连接糖基化蛋白,在组织完整性、分泌及分泌小泡形成、生物发育等过程具有重要作用,可作为害虫防治的潜在靶标。
rna干扰(rnai)是一种由双链rna分子引起的特异性转录后基因沉默的现象,自2006年获得诺贝尔奖以来,rnai技术一直跻身于科技前沿。rnai不仅是研究基因功能的有力工具,同时在害虫防治方面也具有极大潜力。通过rna干扰进行害虫防治具有如下特点:1)杀虫具有专一性,对非靶标生物无杀伤作用;2)rna在自然界极易降解,无残留;3)对环境无毒无害,相对安全。因此,国内外学者将其称为第四代杀虫剂。基于rna干扰进行害虫防治的基础技术是筛选靶标序列获得具有高致死作用的dsrna。
技术实现要素:
针对飞蝗迁飞性和跳跃性的特点,本发明基于rna干扰技术,选取粘蛋白基因为对象,利用其重要生物学功能来进行害虫防治,安全高效,是一种环境友好型、对人类无毒无害、特异性的新型分子靶标,在害虫防治领域具有广阔的应用前景。
本发明提供的粘蛋白来源于飞蝗,其名称为mucin17。所述mucin17的基因序列如seqidno:1所示。该基因序列是基于飞蝗整虫转录组数据库,搜索获得的片段通过bioedit软件结合飞蝗基因组序列对其进行拼接,然后经pcr扩增、克隆测序获得。
mucin17基因编码的氨基酸序列是如seqidno:2所示的蛋白质。
本发明提供的一种飞蝗粘蛋白基因dsrna,其核苷酸序列是seqidno:3,是根据seqidno:1通过primerpremier5.0软件设计含有t7启动子的上游引物seqidno:4和下游引物seqidno:5,通过pcr扩增获得seqidno:3,其产物含有t7启动子。产物经纯化后按照t7ribomaxtmexpressrnaisystem(promega)试剂盒说明体外转录合成dsrna。
seqidno:3合成的dsrna在飞蝗防治中的应用:通过微量进样器将seqidno:3合成的dsrna注射进入飞蝗体腔内。结果表明:和对照相比,seqidno:3合成的dsrna可以导致飞蝗蜕皮前及蜕皮中致死(致死率分别为33.3%和8.3%)、翅皱缩不能展开和后足发育畸形(表型率为50%),最终导致死亡。
附图说明
图1:飞蝗粘蛋白基因mucin17序列获得。a:mucin17引物设计与拼接示意图;b:部分测序峰图;c:mucin17基因翻译后氨基酸序列与结构示意图。
图2:飞蝗注射seqidno:3合成的dsrna24小时后,飞蝗粘蛋白基因mucin17的mrna沉默情况,β-actin做为内参基因,其中**,p<0.01。
图3:5龄飞蝗注射seqidno:3合成的dsrna,飞蝗蜕皮前及蜕皮中致死(致死率分别为33.3%和8.3%)、翅皱缩不能展开和后足发育畸形(表型率为50%),最终导致死亡。a:注射dsgfp的对照;b-d:注射seqidno:3合成的dsrna,即dsmucin17;e-f:对照和异常翅的表型图;g-h:对照和异常足的表型图。
具体实施方式
实施例1:飞蝗粘蛋白基因序列获得
1、飞蝗粘蛋白基因序列获得
基于本实验室获得的飞蝗整虫转录组数据库(genbank登录号:gezb00000000),采用生物信息学方法对飞蝗粘蛋白基因(mucin)进行搜索,获得飞蝗粘蛋白基因mucin17部分序列。利用ncbi网站对序列进行blast分析,同时结合飞蝗基因组序列(genbank登录号:avcp000000000)获得其编码序列,利用bioedit软件进行拼接及比对后,获得飞蝗粘蛋白基因mucin17全长开放阅读框序列。根据获得的飞蝗粘蛋白基因mucin17全长开放阅读框序列,采用primerpremier5.0软件设计4对上、下游引物序列且每队引物之间与共同区域衔接,引物设计位置、长度及序列如图1所示(图1中的a),引物送往生工生物工程(上海)有限公司合成。
选取生长健康,大小一致的飞蝗成虫(本实验室养虫室室内饲养),解剖翅组织,并冷冻于液氮中。每3头一个生物学重复,设置3个生物学重复,依照trizol试剂盒(takara公司)提取翅组织总rna。采用m-mlv反转录酶(takara公司)将所提总rna反转录成第一链cdna,并以此作为模板,结合设计的上下游引物(图1中a),通过pcr扩增仪分别扩增获得mucin17基因片段,将得到的产物经gelextractionkit(sigma)试剂盒进行纯化、连接peasy-t3载体(北京全式金生物技术有限公司)并转化到大肠杆菌transt1感受态(北京全式金生物技术有限公司)中,挑取阳性克隆送往上海英潍捷基生物有限公司进行测序、拼接(测序峰图如图1中的b所示),获得其全长开放阅读框序列,其序列为seqidno:1。利用expasy-translatetool(https://web.expasy.org/translate/)将飞蝗粘蛋白基因mucin17序列翻译成氨基酸序列,其序列为seqidno:2。该基因编码2650个氨基酸,smart网站(http://smart.embl-heidelberg.de/)预测其含有两个表皮生长因子结构域(epidermalgrowthfactordomain,egf)、一个钙结合类表皮生长因子结构域(calcium-bindingegf-likedomain,egf_ca)、一个精子蛋白,肠激酶和糖蛋白结构域(spermprotein,enterokinaseandagrindomain,sea)和一个跨膜结构域(transmembranedomain,tm)(图1中的c)。
2、飞蝗粘蛋白基因特异性dsrna合成
1)飞蝗粘蛋白基因dsrna引物的设计
基于飞蝗粘蛋白基因mucin17开放阅读框序列(seqidno:1),采用primerpremier5.0软件设计dsrna引物,其上下游引物序列分别为seqidno:4和seqidno:5。上下游引物均携带t7启动子序列。所有引物均由生工生物工程(上海)有限公司合成。
2)飞蝗粘蛋白基因dsrna的合成
以上述获得的cdna为模板,seqidno:4和seqidno:5作为上下游引物,进行pcr扩增。将扩增的pcr产物,经gelextractionkit(sigma)试剂盒纯化后按照t7ribomaxtmexpressrnaisystem(promega)试剂盒说明体外转录合成dsrna。使用nanodrop2000(thermoscientific)进行定量,使其终浓度达到2μg/μl。保存于-20℃低温冰箱备用。
实施例2:飞蝗粘蛋白基因dsrna致死飞蝗实验
1、飞蝗粘蛋白基因dsrna注射
选取生长健康、大小一致的30头5龄第2天若虫进行实验。将5μl(10μg)seqidno:3的dsrna用25μl规格微量注射器顺着血流方法轻轻注射进入若虫侧腹部的二、三腹节之间。同时选取30头若虫设立为对照组。注射相同体积和浓度的dsgfp至对照组体内。分别将注射后飞蝗置于30℃恒温生化培养箱中饲养(光照:黑暗时间=14h:10h,温度30±2℃,湿度60%),每天饲喂新鲜小麦幼苗和麦麸。
2、飞蝗粘蛋白基因沉默检测
收集注射dsgfp和dsrna24h后的若虫各9头,解剖翅芽组织进行总rna提取,并反转录成第一链cdna,每组设置3个生物学重复,每个生物学重复3头若虫,采用real-timepcr方法分别检测目的基因(mucin17)和管家基因(β-actin)的相对表达量,计算其沉默效率。结果表明,与对照组比较,注射dsrna后,处理组飞蝗粘蛋白基因表达量极显著降低(图2)。
3、注射dsrna后表型观察
5龄若虫经注射dsrna后,注射dsgfp对照组6天后开始蜕皮并全部成功蜕至成虫,蜕至成虫后虫体形态活力正常,并于半天后开始正常进食。注射飞蝗粘蛋白基因dsrna的处理组若虫共30头,与对照组若虫相比,飞蝗蜕皮前及蜕皮中致死(致死率分别为33.3%和8.3%)、翅皱缩不能展开和后足发育畸形(表型率为50%),最终导致死亡(图3)。
序列表
<110>中国农业科学院植物保护研究所
<120>飞蝗粘蛋白mucin17、其编码基因及其应用
<141>2019-02-25
<160>5
<170>siposequencelisting1.0
<210>1
<211>8290
<212>dna
<213>未知(unknown)
<400>1
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aaggaggacgagcaggaaccgcggcaggacggcggcggcgcaggtggcaacgtgctgctg6180
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gcggcgcgcaacgaggtgtgccagcgcgccgcggacggccacccacgttgcctctgcagg6300
ccgggcttcgccaggatcttcccagaccggccctgcaagccgacatacacatatacgcta6360
cggctagtgctgtcgcgccatggcgaccgccccatcaccttctcgcccggcctgggagat6420
acaggcagtgacaccttcgctgagacgtctcgcgctgccgccgaggggctcgaccggctg6480
gtgatgcagtccgacctgcgcgacgtcttccacggtgtctctgttgaggcgttcctgccg6540
cccgacgaaggcagcaccgccggcgttatcgtcaccttccaagtcaggctgtcagacaat6600
accgacgaggagcgactgaaggaggtattccgaaagtcactcagggctaccaactacagc6660
ctgggtggcacggatgtctttgccgacaaggagcgcatccagcacattgaggcagaagac6720
ttcgacgagtgcgcgagccggcagtaccacgactgttcggagaacgcgcactgcttcaac6780
ctgcgcggcacgtacacgtgcagctgccgcgagggcttcgccgacctctccgagaacccg6840
ctgttcccgggcaggctctgctcagcggagctgatcgggtgtgagcggtgccacttccac6900
ggcacgtgcggcgacgaccaccaccaggtgcagtgcgagtgcttccagtggtacgccggg6960
gacacctgccacatcaacctcaaggcgctgctgatcgcactggtgacgatcggctgcgta7020
ctggtggcgcttctgctggtgtgcgtgctgatgacgtgcgcgcgtggaggcagtcgcggt7080
gggcgcacccggggacggtcgcggcgcgaggggccgttggagcggcgcgccatgatccac7140
gactcggccagcgaggccagcggcgaccagacgctgcccaggagccagactccggcagcg7200
gtgttcggcaagaaggcgcgaccggtgtcggcgccgacggcgacggtggtagccgtggtt7260
cccccgcctccagcgggcttcgcccccacgcccgtaccggtgccccctccactgcagccg7320
cccccggagcagcgcgaccgctcgctgacggtgatgatcccgcgcgccaagtaccggccg7380
tcgcccgtggcgcccccgcaccagccactgcccacgctggtctccatgtcgacgttcggc7440
gatgccgagaaacgcgccatcgctcacgagagcaagctgctcgccctgctggagggagcc7500
tccaagcaggatggccagctacagtcccatcatcatcatcatcaacaccagcagcagaca7560
tcaaaacgaaaacctagcaacacaacgagccaagcaacagaggcgtccactcccaggaag7620
aattcatcttcgaggaaaccttcgaatggaaacattccaccaccagcaccgggagccctg7680
gtctcggcgggcttcgaggtgtcggcgactgtcggcaagcctgcgacggactccgacacg7740
gaggcatcggcagtcgtccacaacggttccacgttgcggacgacagacagtgtacaaatc7800
accgttgaagagatgtcggagccatcggtaaaggctgggtctgctgtggaaacgctgacc7860
gtgtccgaagctcgatcttgtgacgagacaacgatacatccaccgacaaagtcgctccac7920
agcaattacagctcaaaacactcatcaaacaacctcaacaatgatgagggccacacaatg7980
gctgagagagacatcggctctacgttcgtgatgccacagtcgcacttgtacacaccagac8040
aggggaagtgacatttcaaatttcgactcactgtgagtaaagccttcagtgcaactgcag8100
aagtttgctacatcacttgcactgcgaggctttcgggagtggagatgcgacacttctgaa8160
cggagctatcatgtgtcgacttgaaacgaaggaaattgtgaaaagaatgtgactgtaaca8220
ggctcaatttcaaagacatattgagtgagttattgtatttaatataatggaggactgaat8280
actgttaact8290
<210>2
<211>2650
<212>prt
<213>未知(unknown)
<400>2
metalametaspglnalaproalavalglnprothrlysthralaglu
151015
proproalavalalaprothrvalglyproaspalaserglnileala
202530
prosermetaspthrleuglnproalaasnvalgluasnilevalthr
354045
glyserthrileilephepheaspglugluaspglyvalaspserasp
505560
serglyprothralaalaalathrproalaleuaspserserthrval
65707580
alaalaleuseraspserargthrthrvalvalasnalaglyalaala
859095
thrvalilepheproaspprogluprothrthralagluproalaasp
100105110
serglnglngluserthrthrvalserthraspaspaspglutyrgly
115120125
thrglyaspglugluasnglyglyileserglnthrprothrserasp
130135140
alaglyproalaalaserthrservalvalthrvalthralaserpro
145150155160
valthrthrthrvalalavalprovalthrlyslysprovalvalgly
165170175
proglyglysergluileglnvalseraspleuleuserleuglyser
180185190
leuglyileasnglyleuasnalaleuglyprovalpheasnalamet
195200205
alaglyleuileglnglyasnleuglnglnglnproaspalaalalys
210215220
argargasnaspservalasnthrglyasnargalaprogluleuval
225230235240
proglyasparggluasptyrprohisprovaltyrargproglnpro
245250255
glnalaaspthrgluproproproargserproiletyrileproval
260265270
glyglyleualaalaalaasphisaspileglualaalaglusergln
275280285
asntyrglualaargilelysileasnalaleuproglyserglylys
290295300
trpglyglyseralagluservalglyglyvalvalileglyargpro
305310315320
thrleuglulysproleuglyaspglyileproileserproglyglu
325330335
valilethralaasnseraspvalilevalglylysproalavalala
340345350
glyproargproprolysilethralaalaglyhisgluhisasphis
355360365
alaleuasphisglyglyilealavalglymetargpropropropro
370375380
hisarglysprotrpleuhishishisglnglnproprometargasp
385390395400
glupheleuglyproproproproproproproproglyserhisval
405410415
glyglygluvalglnglyargproproglyseralaasnhistyrarg
420425430
leuhishisaspargprohisproglyleuasnleuasnhisasnhis
435440445
asnglnasnasnpheasnasnasnglnlysproproalaleulysleu
450455460
argprohisvalvalgluglyleuprotyrlysleugluservalval
465470475480
thrglyserglnalavalleuvalalagluglyglyalaglygluval
485490495
valhisglyserprovalprohisarghisalavalleuproalagly
500505510
alaaspalaalaserleuleuvalaspileargproserglnvalala
515520525
asnvalleuileproargglyserhisthralahisvalpheserala
530535540
proalaproserproleuvalalaaspaspvalpheargaspaspser
545550555560
propheprovalproglualaalaalaaspphevalglyleuaspala
565570575
alaproglnleugluhisglyvalglythrvalilegluvalhispro
580585590
glyaspserprothrgluthrglyglyvalvalalaglyleuglngly
595600605
glyaspileasnvalasnilealaproglyglnglyileservalasp
610615620
valglyproglyglysermetvalhisalaargproaspservalpro
625630635640
glyproglylysgluvalhisargileglnproserglyvalvalleu
645650655
aspalaproargproglnleuglnproglnprohisargproglugln
660665670
proleualahisproproleuprohisargprogluserproleuala
675680685
hisproproproproproproproproproproprosersersergln
690695700
glnserproserargargproprovalthrleutyrargproserthr
705710715720
glythrglyglyserthralaglyglyargprohisglntyrargphe
725730735
provalproglyargproleuargproseraspphemetthrpropro
740745750
proproproprohisleuproleuargproproalaprolysprothr
755760765
proglyvalhisglnhisleuargproglnglnproglyglyproleu
770775780
valileproleuargproaspalaglyargproproargproglyasn
785790795800
asphisphevalhisglyhisasphisgluhisaspleuhisilehis
805810815
glyaspasphisgluleuhisvalhisglyleuglualagluargasp
820825830
histhrthrargleuhisprovalargileglyglyglyglyvalval
835840845
valargproglyproglyprovalaspvalphevalargproaspgly
850855860
gluleuvalservalgluaspaspleuglyvalalaglyserserarg
865870875880
proleuproargproproserleuserprolysproserargargpro
885890895
provalthraspleuphealailetrpproglyaspargtyrasngly
900905910
serargproglnhisglyalaglyproproproproargglnglyval
915920925
glyserargproglyasparggluarggluglngluargproprogln
930935940
glnprothrleuglnhisglyphehisgluvalargprovalalaglu
945950955960
proglyaspvalprothrvalproglyphevalasnglyglnprolys
965970975
proproproaspphehisserglnserhisargglnargpropropro
980985990
ilethrglualaaspasnasnileilevalglyhisvalprohisphe
99510001005
asngluleuhisserprovalproproaspaspleuargprohisval
101010151020
valalavalglygluprothrasnlysvalaspgluproproglyasn
1025103010351040
valasphishisglnleuhishishishisvalalaglygluvalser
104510501055
serprogluileglyvalvalargproglygluargproleuglnser
106010651070
glyileargpheproglyalaglyasphisphegluvalglugluasp
107510801085
thrglnservalglulysseraspproaspserthrglyalaalaser
109010951100
glythraspglyserproaspproproalahisglyseraspserala
1105111011151120
aspseraspgluaspargleuglyserileleuhisglyglythrgln
112511301135
thrgluargproleuvalthrleuproproaspasnserproglyhis
114011451150
asphisvalhisasnthrasnthrglythralathrarggluproglu
115511601165
hisargaspgluthrproserargasngluileproileleuglyval
117011751180
glylysgluargproprohisleuhisvalglyvalserglytrpgln
1185119011951200
ileileglyserglnproaspargproglythrproglnaspglyala
120512101215
serasnglyproaspvalglyseralathrproglyargglypropro
122012251230
hisargvalalaglytyrargproproglytrphisthrglyglyser
123512401245
glyhisleuilevalthrvalglyaspgluserglyprovalargasp
125012551260
argproserglyaspaspseraspvalhistyrglyargprohisval
1265127012751280
serproglyglythrprogluaspalaargprohistrpserglyglu
128512901295
alaproargproproilegluthrprovalvalasparggluthrser
130013051310
glytyrglyvalalaasppheglyglnaspvalproprophelysala
131513201325
metglygluaspvalseralavalvalaspservalleuargpropro
133013351340
serleuasnleuproproalathralavalargprothrglnpropro
1345135013551360
proproproproproproproproargvalthrvalleuthrasphis
136513701375
arghisasnasnlysprohisargglnprovalilevalleuglyarg
138013851390
proalagluphethrglyaspvalalathrhisargprothrglnarg
139514001405
prosermetthrglugluglyargproglyaspglygluproproala
141014151420
aspmetthrgluvalasplysproproproproproproargproala
1425143014351440
valalaaspleugluthralaalaserthralaargprophelysleu
144514501455
argproglnaspaspservalmetglyleualaproproproproarg
146014651470
valthrvalvalglyvalglyasnglyglyglyproproproleugln
147514801485
glyaspserthrhisargprovalleuileglyglyglyarglyspro
149014951500
provalglnthrleuproproproprosergluphegluglnpropro
1505151015151520
proproleuvalthrthrproargargproprophevalvalthrpro
152515301535
prothrproalatyrserseralaprothrthrlysprolysproser
154015451550
thrserthrservalaspaspthrlysaspgluleugluaspgluasp
155515601565
glngluleualagluthrserserglyserlysproproalapropro
157015751580
leuproproleuserproservalthrseralaalaalaprogluleu
1585159015951600
valaspvalthrtyrargproprovaltyrthrthrglnargproarg
160516101615
provalthrproleuaspalathrarglysproproglnvalvalser
162016251630
proserargthrglualaaspmetleugluthrvalservalglyarg
163516401645
provalserglnproleuproproproproproproproproprothr
165016551660
argglualaproproargproproservalalaleualaprothrpro
1665167016751680
valargaspalaalaserthrgluvallysalaserpropropropro
168516901695
proalaalaalaalathrproproalathrleuaspargthrtrpthr
170017051710
thrleualavalaspleugluglyserglnthrargleuproasnarg
171517201725
argargprothralathrproalavalalaprothrseralathrser
173017351740
seraspthrserserasnthrglylysvalthrservalvalvalthr
1745175017551760
serglyserlysthrvalleuphevallysprothralaalaargleu
176517701775
prothrglnthrthrlysserthrprothrproargproargproarg
178017851790
proargprothrthrgluthrglyglyleupheaspasppheaspasp
179518001805
valalaaspgluaspaspglugluglusergluthrglythralagly
181018151820
glyserargglylysvalaspleuglyvalalaserserasnglnglu
1825183018351840
lysvalvalvalglyseralaproalaserthrleutyrvalthrhis
184518501855
thrhisthrleuthrvalthrthrthrgluthrglnvalvalserthr
186018651870
alaglyargproprovalthrargthrvalvalleuthrlysthrgln
187518801885
thrserthrvalvalaspthrvalthrgluthrglnthrleuleuarg
189018951900
prothrservaltyralathrvalthrthrthrvalseralailearg
1905191019151920
prophethrgluglyvalserserthrserthrvalserthrvalthr
192519301935
thrproalaprothrthrglyserglnpropropropropropropro
194019451950
alaglythrglyaspargproglyaspargthralaglyalaglyasp
195519601965
thrphephevalvalvalthraspprohishislysleupropropro
197019751980
serglyalaproleuproproglyileileglutyrgluvalasplys
1985199019952000
gluaspgluglngluproargglnaspglyglyglyalaglyglyasn
200520102015
valleuleuthralaglyvalglyglyserglyalaalaalathrser
202020252030
alacysargproglucysseralaalaargasngluvalcysglnarg
203520402045
alaalaaspglyhisproargcysleucysargproglyphealaarg
205020552060
ilepheproaspargprocyslysprothrtyrthrtyrthrleuarg
2065207020752080
leuvalleuserarghisglyaspargproilethrpheserprogly
208520902095
leuglyaspthrglyseraspthrphealagluthrserargalaala
210021052110
alagluglyleuaspargleuvalmetglnseraspleuargaspval
211521202125
phehisglyvalservalglualapheleuproproaspgluglyser
213021352140
thralaglyvalilevalthrpheglnvalargleuseraspasnthr
2145215021552160
aspglugluargleulysgluvalphearglysserleuargalathr
216521702175
asntyrserleuglyglythraspvalphealaasplysgluargile
218021852190
glnhisileglualagluasppheaspglucysalaserargglntyr
219522002205
hisaspcyssergluasnalahiscyspheasnleuargglythrtyr
221022152220
thrcyssercysarggluglyphealaaspleusergluasnproleu
2225223022352240
pheproglyargleucysseralagluleuileglycysgluargcys
224522502255
hisphehisglythrcysglyaspasphishisglnvalglncysglu
226022652270
cyspheglntrptyralaglyaspthrcyshisileasnleulysala
227522802285
leuleuilealaleuvalthrileglycysvalleuvalalaleuleu
229022952300
leuvalcysvalleumetthrcysalaargglyglyserargglygly
2305231023152320
argthrargglyargserargarggluglyproleugluargargala
232523302335
metilehisaspseralaserglualaserglyaspglnthrleupro
234023452350
argserglnthrproalaalavalpheglylyslysalaargproval
235523602365
seralaprothralathrvalvalalavalvalproproproproala
237023752380
glyphealaprothrprovalprovalproproproleuglnpropro
2385239023952400
progluglnargaspargserleuthrvalmetileproargalalys
240524102415
tyrargproserprovalalaproprohisglnproleuprothrleu
242024252430
valsermetserthrpheglyaspalaglulysargalailealahis
243524402445
gluserlysleuleualaleuleugluglyalaserlysglnaspgly
245024552460
glnleuglnserhishishishishisglnhisglnglnglnthrser
2465247024752480
lysarglysproserasnthrthrserglnalathrglualaserthr
248524902495
proarglysasnserserserarglysproserasnglyasnilepro
250025052510
proproalaproglyalaleuvalseralaglyphegluvalserala
251525202525
thrvalglylysproalathraspseraspthrglualaseralaval
253025352540
valhisasnglyserthrleuargthrthraspservalglnilethr
2545255025552560
valgluglumetsergluproservallysalaglyseralavalglu
256525702575
thrleuthrvalserglualaargsercysaspgluthrthrilehis
258025852590
proprothrlysserleuhisserasntyrserserlyshisserser
259526002605
asnasnleuasnasnaspgluglyhisthrmetalagluargaspile
261026152620
glyserthrphevalmetproglnserhisleutyrthrproasparg
2625263026352640
glyseraspileserasnpheaspserleu
26452650
<210>3
<211>430
<212>dna
<213>未知(unknown)
<400>3
cggcaaaggacacggggggacacgggccaagacgggcgcggaagccacggccgcccgacg60
ccgacgcagacgacgaagcacccacgccgcgaccgaggccgagaccccgcccaccaccga120
gaccggaggacggacgaccgacgacgcgcggacgaggacgacgaggaggaacggagacgg180
gcacggccggcgggcgcggggcaaggggaccgggcgcgcagcccaaccaggagaaggggg240
gcggccggcgcccgccccaccccacgcacccacacgcacacaccacagaacgacacggag300
acgcagggggagcacagccggccggccgccgggacgcgcacggggccaccaagacgcaga360
cgcaacagcgggacacagcaccgagacccagacgcgcccggccgaccagcgcagccaccg420
caccaccacg430
<210>4
<211>45
<212>dna
<213>未知(unknown)
<400>4
gcgtaatacgactcactataggcggcaaagtgacatctgtggtgg45
<210>5
<211>43
<212>dna
<213>未知(unknown)
<400>5
gcgtaatacgactcactataggcgtggtggtgacggtggcata43