CN109136233B - 超量表达转录因子SlBBX20提高番茄花青素的含量 - Google Patents
超量表达转录因子SlBBX20提高番茄花青素的含量 Download PDFInfo
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- CN109136233B CN109136233B CN201710503661.XA CN201710503661A CN109136233B CN 109136233 B CN109136233 B CN 109136233B CN 201710503661 A CN201710503661 A CN 201710503661A CN 109136233 B CN109136233 B CN 109136233B
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Abstract
本发明属于植物基因工程技术领域。具体涉及超量表达转录因子SlBBX20提高番茄花青素的含量。本发明克隆到一个控制番茄叶片花青素合成的关键基因SlBBX20，对其进行了生物学功能鉴定。该基因的cDNA序列如序列表SEQ ID NO：1所示，其编码的蛋白质的序列如SEQ ID NO：3所示。通过转基因功能验证表明，在番茄中超量表达该基因，可以显著调节番茄叶片花青素含量的功能。
Description
技术领域
本发明属于植物基因工程领域。具体涉及超量表达转录因子SlBBX20提高番茄花青素的含量。本发明利用反向遗传学的方法，分离克隆了一个具有提高番茄花青素的含量的功能基因SlBBX20，将该基因在番茄中超量表达，能够显著提高番茄叶片花青素合成与积累。利用本发明可以有效提高番茄叶片中花青素的含量。
背景技术
花青素(Anthocyanin)，又称为花色素，是黄酮类化合物中最主要的组成部分，主要赋予植物果实、叶片、花等组织器官以各种各样的颜色，是一种水溶色素。花青素作为植物次生代谢的重要产物，参与多种多样的生物进程。当植物受到外界环境胁迫，如光照、低温、盐胁迫、氧化胁迫、蔗糖及大量营养元素缺失等，都能诱导花青素的表达以提高植物对外界环境的适应能力。类黄酮化合物形成的起始底物为苯丙氨酸，这一类化合物主要包括花青素、黄酮醇以及原花青素3种(Winkel-Shirley，2001)。近几年，对于类黄酮的研究主要着眼于对人类健康方面的潜在功能，比如抗氧化、抗衰老、抗心血管疾病及抗癌等(Yaoetal，2004)。因此，进一步探讨花青素的合成、调控及生物学功能具有重要的意义。
目前，花青素生物合成途径及其关键酶的研究已经相当清楚(Martens etal2010；Wang et al 2013)，对花青素合成途径相关转录因子的研究也取得了较大进展，主要受四种转录因子家族的调控，分别是MYB、bHLH、WD40-repeat以及bZIP。在拟南芥、玉米、矮牵牛、葡萄、金鱼草、苹果、紫苏、草莓、非洲菊、杨梅等植物中均分离和鉴定出一些花青素合成相关的调控因子。研究表明，在拟南芥花青素合成途径中，MYB转录因子家族基因与bHLH转录因子家族基因及WD40蛋白家族互作，形成一个MYB-bHLH-WD40(MBW)复合体，从而调控花青素的合成，在对其他植物，如：草莓、苹果、葡萄、杨梅等的研究中也发现了类似的调控模式(An et al 2012；Gonzalez et al 2008；Outchkourov et al 2014；Ramsay andGlover2005；Schaart et al 2013)。MYB转录因子家族主要参与苯丙烷代谢途径的调控，其中主要是R2R3-MYB亚家族。对R2R3-MYB(PAP1、PAP2)研究发现R2R3-MYB激活了花青素合成途径上的结构基因，同时苯丙烷代谢途径的起始基因苯丙氨酸裂解酶基因(PAL1、PAL2)也被激活，进而促进花色素的积累(Borevitz etal，2000)。R3-MYB蛋白家族中，MYBL2被认为是花青素合成途径上的一个抑制子(Matsui et al 2008)。有研究表明，在强光照条件下，MYBL2的表达水平明显降低，进而导致花青素大量积累(Rowan et al 2009)。拟南芥中参与调控花青素合成的bHLH家族基因主要有：TT8、EGL3、GL3等，当异位表达EGL3、GL3和TT8时发现三者之间存在功能冗余，在蛋白互作方面可以相互替代(Zhang et al 2003)。在花青素合成生物途径中，WD40并没有很显著的催化特性，TTG1是这一家族的一个重要的与花青素合成相关的转录因子，主要与bHLH和MYB家族的相关转录因子形成一个复合体，进而激活花青素合成途径(Chattopadhyay et al 1998)。bZIP家族转录因子中HY5，HYH是参与花青素合成途径的两个重要转录因子。HY5(elongated hypocotyl 5)，是下游光敏色素、隐花色素，UV-B光感受器的一个组件，参与花青素合成途径(Vandenbussche et al 2014)。HY5是对光信号响应的一个重要转录因子，并且通过影响下游基因的表达进而调控光形态建成(Chattopadhyay et al1998)。因此，HY5在协调光信号及关键基因表达方面起到了一个重要的调节作用(Lee et al 2007)。HY5调控花青素合成的机理主要是通过与早期表达基因及后期表达基因(如CHS、CHI、F3H、F3’H、DFR及LDOX等)的启动子结合进而调控此途径上的结构基因的表达(Zhang et al 2011)。
在番茄中，对花青素积累及调控的研究相对较少，有些研究是通过导入异源物种基因来调控花青素(Butelli et al 2008)。番茄中研究鉴定的与花青素相关的调控因子主要有两个，LeANT1和LeAN2，均为R2R3-MYB型转录因子(Mathews 2003；Petroni andTonelli2011；Sapir et al 2008)。而在对众多植物的研究中发现的锌指蛋白类转录因子能够调控花青素的积累未见报道。
发明内容
本发明的目的在于克服现有技术的缺陷，分离、克隆了一个植物花青素合成的调控基因，被命名为SlBBX20基因，通过在番茄中超量表达SlBBX20基因，能够显著提高番茄花青素的合成与积累，经基因改良后的番茄叶片由绿色变为深紫色。针对这样的结果，申请人推测如果能够充分开发与利用，从番茄叶片中提取天然抗氧化物质花青素，将可能产生较好的经济效益。
申请人发现并鉴定了首个锌指蛋白类转录因子能够调控花青素的积累，超量表达该转录因子后能够强烈调控番茄叶片花青素合成与积累。
本发明的SlBBX20基因是能够调控番茄叶片花青素积累的一个新的转录因子，目前未见有该基因功能的报道。申请人从番茄材料中克隆到该基因的序列，该基因在GenBank核苷酸数据库中登录号为XM＿004231383.2预测为Solanum lycopersicum B-boxzincfinger protein20(LOC101247171)，transcript variant X2，mRNA。SolGenomicsNetwork上预测为Zinc finger protein CONSTANS-LIKE 1。分析其保守域发现含有两个bBOX结构域，基因注释为锌指蛋白20，因此申请人根据该类基因的命名规则，将该基因命名为SlBBX20基因。本发明是这样实现的：
本发明在对利用全基因组关联技术获得的大量候选基因进行功能鉴定时，发现在番茄中超量表达SlBBX20转录因子后，番茄叶片中的花青素显著积累。
本发明所述的植物包括单子叶植物、双子叶植物。
本发明将鉴定克隆的基因命名为SlBBX20，包括该基因的DNA序列、cDNA序列，与该序列具有高度同源性的序列及编码相同功能蛋白质的DNA序列，均属于本发明的保护范围。
本发明所述基因包括完整的或部分的(20个及以上)核苷酸和(6个及以上)氨基酸序列。
本发明可以通过农杆菌介导的遗传转化方法或其他转基因方法把SlBBX20基因转入植物中，通过筛选鉴定并纯化，获得纯合的转基因植株或株系。或与其他基因融合，通过颜色变化作为标记，简单快速的筛选转基因植株均属于本发明的保护范围。
本发明通过在植物体内超量表达该基因可以提高植物花青素的合成与积累，其在该方面的应用均属于本发明的保护范围。
附图说明
序列表SEQ ID NO：1是SlBBX20基因的cDNA序列。序列长度为835bp。
序列表SEQ ID NO：2是SlBBX20基因编码的氨基酸序列和对应的核苷酸序列(上列为核苷酸序列，下列为氨基酸序列)。
序列表SEQ ID NO：3是SlBBX20基因编码的蛋白质序列。
图1：利用pDNOR221和PMV3构建SlBBX20超量表达载体的示意图。
图2：SlBBX20基因在番茄不同组织和器官中特异性表达分析。图2中A图：番茄不同组织中SlBBX20的表达分析。图2中B图：番茄不同花器官中SlBBX20的表达分析。
图3：超量表达SlBBX20基因的植株SlBBX20-OE(B，D)和干涉表达植株SlBBX20-RI(A，C)的表型。
图4：超量表达SlBBX20在筛选培养过程中愈伤组织变成紫色。
图5：超量表达SlBBX20的转基因植株根颜色的变化。图5中A图：超量表达SlBBX20的根变成紫色。图5中B图：对照的根为白色。
图6：超量表达SlBBX20的转基因植株的花和果实颜色比较。图6中A图：超量表达SlBBX20的花萼片变成紫色。图6中B图：对照的花萼片为绿色。图6中C图：超量表达SlBBX20的果实变成深绿色。图6中D图：对照的果实有浅绿的果肩。
图7：超量表达SlBBX20后T0代转基因株系表达量的检测。
图8：超量表达SlBBX20株系叶片花青素的含量与颜色比较。图8中A图：超量表达SlBBX20株系叶片花青素的含量。图8中B图：提取超量表达SlBBX20株系叶片时花青素的颜色。
具体实施方式
以下结合具体实施例对本发明做出更详细的描述。根据以下的描述和这些实施例，本领域技术人员可以确定本发明的基本特征，并且在不偏离本发明精神和范围的情况下，可以对本发明做出各种改变和修改，以使其适用各种用途和条件。
实施例1：SlBBX20基因的克隆与遗传转化
1)、SlBBX20基因的克隆与分析
本发明中克隆的SlBBX20基因是根据全基因组关联分析结果，对大量候选基因进行转基因功能验证的基础上，发现SlBBX20转录因子基因能显著提高番茄合成和积累花青素的能力。根据数据库注释的基因信息(Http：//solgenomics.net/tomato/)，设计扩增SlBBX20基因的引物，其正向引物为SlBBX20For：5｀-CAGAGACTTTAGGTGTGAGCCC-3｀，反向引物为SlBBX20Rev：5｀-GGAAAAAACTACACATCTGGTCG-3｀，通过PCR的方法(具体见表1)，从TS-93(引自中国农科院蔬菜花卉研究所)中扩增出全长SlBBX20基因cDNA序列。扩增方法是先提取番茄材料TS-93的RNA，然后利用反转录试剂盒II 1st Strand cDNASynthesis Kit(购于Vazyme biotech)，按照试剂盒说明书反转录合成SlBBX20基因的cDNA，利用上述设计SlBBX20基因的引物，通过PCR扩增出SlBBX20全长基因，经1％琼脂糖凝胶检测，用回收试剂盒(购自Omega Bio-Tek公司，具体程序见说明书)回收目的片段，克隆至pEASY-Blunt载体(购自北京全式金生物技术有限公司Trans Gen Biotech)，取3μL PCR产物，1μL载体于25℃保温15min，用热击法(参照J.萨姆布鲁克等著，黄培堂等译，分子克隆实验指南，第三版，科学出版社，2002版)转化至大肠杆菌Trans-T1(购自北京全式金生物技术有限公司TransGenBiotech)，涂布于含有100mg/L卡那青霉素的LB固体平板上，37℃培养过夜，挑选单克隆若干，于含Kan 100mg/L的液体LB培养基中，37℃200r/min振荡培养过夜，用通用引物M13正向引物(5＇-TGTAAAACGACGGCCAGT-3＇)与SlBBX20基因反向引物(5｀-GGAAAAAACTACACATCTGGTCG-3｀)检测重组克隆，对经检测正确的阳性重组克隆进行序列测定，测序工作由北京擎科生物科技技术有限公司完成。
利用GENESCAN对该基因序列分析，结果表明，该基因ORF序列全长为612bp，编码203个氨基酸。该基因含有2个内含子，具有两个b-BOX保守结构域。
表1 SlBBX20基因克隆的PCR体系
2)、载体构建
在扩增SlBBX20基因的前后引物前分别加上部分B位点5｀-AAAAAGCAGGCT(attB1)，5｀-AGAAAGCTGGGT(attB2)，随后以番茄材料TS-93(引自中国农科院蔬菜花卉研究所)的cDNA为模板，进行PCR扩增，然后以获得的PCR产物为模板，用全长B位点5｀-GGGGACAAGTTTGTACAAAAAAGCAGGCT(attB1)和5｀-GGGGACCACTTTGTACAAGAAAGCTGGGT(attB2)为引物进行第二轮扩增，扩增产物经1％琼脂糖凝胶检测，用胶回收试剂盒回收目的片段，克隆至pDNOR221载体(载体图见图1)，取1.9μL PCR产物，0.5μL载体质粒，0.6μLBP酶，于25℃保温5h，用热击法转化至大肠杆菌Trans-T1，涂布于含有Kan 100mg/L卡那霉素(Kan)的LB固体平板上，37℃培养过夜，挑选单克隆若干，于含Kan100mg/L的液体LB培养基中，37℃200r/min振荡培养过夜，用通用引物M13正向引物与SlBBX20基因特异的反向引物检测重组克隆菌液，对经检测正确的阳性重组克隆进行序列测定，测序工作由北京擎科生物技术有限公司完成。对测序正确的单克隆摇菌提取质粒(购自Omega Bio-Tek公司，具体程序见说明书)进行LR反应，与表达载体PMV3(见图1)连接，连接体系及条件同BP反应，使用LR酶，连接完成后热激转化至大肠杆菌，涂布于含有壮观霉素的LB固体平板上，37℃培养过夜，挑选单克隆若干，于含Spec 100mg/L的液体LB培养基中，37℃200r/min振荡培养过夜，检测引物用CaMV35S(5’-GACGCACAATCCCACTATCC-3’)和SlBBX20Rev引物的反向检测重组克隆菌液，挑取正确的单克隆摇菌提取质粒，用XbaI和XhoI做两个单酶切检测，酶切体系可通过产品公司相关网站进行查询，酶切条件为37℃，2h，1％的琼脂糖凝胶电泳检测后若结果为XbaI能检测到目的条带，而XhoI检测不到目的条带的为构建成功的质粒(目的片段中不含有这两个酶的酶切位点)。
对获得的重组克隆利用电转化仪在1800V的电压下转化农杆菌C58(购自北京全式金生物技术有限公司Trans Gen Biotech)，用含利福平(Rif)100mg/L，壮观霉素(Spec)50mg/L的LB固体平板筛选，挑选阳性克隆，28℃、150r/min振荡培养过夜，然后用CaMV35S和SlBBX20Rev引物检测重组克隆菌液，确认为阳性的保存后用于进一步的遗传转化。
3)、遗传转化
将A57番茄种子(引自美国番茄遗传资源中心(TGRC)，http：//tgrc.ucdavis.edu/)，经2％的次氯酸钠消毒15min，然后用清水冲洗数遍，将灭菌后的种子播种于1/2MS发芽培养基上(配方见表2，pH＝5.8)，于25±2℃，黑暗条件下培养直至种子发芽，转入光照强度为1800lx，每天16h光照，8h黑暗的光周期条件下培养。切取7-8d苗龄无菌苗子叶预培养2d(培养基为MS 0，pH5.8，配方见表2)。预培养基MS0重悬至OD600≈0.5的农杆菌液浸染3-5min，用灭菌滤纸吸干多余菌液，重新置回预培养基上，黑暗条件下共培养2d，转入到筛选培养基[编号1.0ZR+头孢霉素(Cef)400mg/L+卡那霉素100mg/L](配方见表2)上进行抗性筛选，每两周继代一次，当抗性芽出现后将外植体转入再生培养基0.2ZR+Cef(头孢霉素)200mg/L+Kan 100mg/L(配方见表2)的培养基中。于20～30d后切下抗性芽，转移至生根培养基(配方见表2)中诱导生根，将根系发达的植株移栽到营养钵中，练苗后移栽到温室。
表2番茄遗传转化培养基配方
注：以上培养基中除MS 0外，均含琼脂7.4g/L，培养基配齐后，补充蒸馏水至1L，按照常规方法灭菌。
实施例2：超量表达SlBBX20基因的表型鉴定
1)、SlBBX20基因的组织表达谱分析
利用Q-RT-PCR技术，对SlBBX20基因在番茄不同组织和器官(根，茎，叶，花，果)的表达情况进行了分析，结果表明，在正常生长条件下，SlBBX20基因在所检测的组织和器官中都有表达，且在花中表达量较高(见图2中的A图)，我们对花的不同结构进一步进行表达分析发现，在花瓣中的表达量最高。其他的花器官结构表达量相对较低(见图2中的B图)。
2)、超量表达SlBBX20的表型分析
超量表达SlBBX20后叶片颜色与转基因的受体材料相比发生了明显的变化，即颜色变成深紫色，而干涉植株是绿色(图3)。在SlBBX20超量表达植株遗传转化的初期，即在筛选培养基上愈伤组织已经表现出颜色的变化(图4)，再将有芽点的转化苗移入生根培养基后，叶片颜色的变化已经非常明显，并且随着根系的生长，根系也开始变成紫色(图5)。将SlBBX20超量植株移栽到土壤中开花结果后，发现花瓣也变成了紫色，而果实则变成了深绿色(图6)。
3)、超量表达SlBBX20后的表达量分析
采用Trizol法提取转基因番茄植株叶片的总RNA。对转SlBBX20的T0代阳性植株的表达量进行了检测。结果发现超量表达植株的颜色深浅与表达量是完全正相关的关系，颜色越深的植株的表达量越高，大致上升了100倍以上，而颜色较浅的植株的表达量较低，大都超量表达了十几到几十倍，即表型与表达量高低是一致的(图7)。
4)、超量表达SlBBX20植株的花青素含量检测
从叶片颜色来看，花青素类物质有较大积累。用常规酸化甲醇法提取色素，配制提取液：1％盐酸甲醇(v/v)，采取适量转基因番茄植株叶片组织液氮研磨，称取0.1g鲜样组织于1.5ml离心管中，并记录重量；加入1ml提取液，震荡混匀，4℃避光提取24h，期间晃动，充分提取。12000g离心15min，取200ul上清液用酶标仪在波长530nm(花青素吸收峰)和657nm(叶绿素吸收峰)测吸光值，以提取液作参比，根据公式：(A/g)＝(A530-0.25＊A657)/m计算花青素的对含量(实验设计三个生物学重复，测量吸光值设计三个技术重复)。结果发现，与背景材料A57相比，大部分转基因株系的花青素相对含量有较大提高，有几个株系，如编号为24，27，70，76等株系提高了十几倍之多(图8)。
参考文献
1.An XH，Tian Y，Chen KQ，Wang XF，Hao YJ.The appleWD40proteinMdTTG1interacts with bHLH but not MYB proteins to regulateanthocyaninaccumulation.J Plant Physiol，2012，169：710-717.
2.Borevitz JO，Xia Y，Blount J，Dixon RA，LambC.Activationtaggingidentifies a conserved MYB regulator of phenylpropanoidbiosynthesis.PlantCell，2000，12：2383-2394.
3.Butelli E，Titta L，Giorgio M，Mock HP，Matros A，PeterekS，Schijlen EG，Hall RD，Bovy AG，Luo J，Martin C.Enrichment of tomatofruit with health-promoting anthocyanins by expression of selecttranscriptionfactors.NatBiotechnol，2008，26：1301-1308.
4.Chattopadhyay S，Ang LH，Puente P，Deng XW，WeiN.ArabidopsisbZIPprotein HY5directly interacts with light-responsivepromotersin mediatinglight control of gene expression.PlantCell，1998，10：673-683.
5.Gonzalez A，Zhao M，Leavitt JM，Lloyd AM.Regulation oftheanthocyaninbiosynthetic pathway by the TTG1/bHLH/Mybtranscriptionalcomplex inArabidopsis seedlings.Plant J，2008，53：814-827.
6.Lee J，He K，Stolc V，Lee H，Figueroa P，Gao Y，Tongprasit W，ZhaoH，LeeI，Deng XW.Analysis of transcription factor HY5genomic bindingsites revealeditshierarchical role in light regulation ofdevelopment.Plant Cell，2007，19：731-749.
7.Martens S，Preuss A，Matern U.Multifunctional flavonoiddioxygenases：flavonol and anthocyanin biosynthesis in ArabidopsisthalianaL.Phytochemistry，2010，71：1040-1049
8.Mathews H.Activation Tagging in Tomato IdentifiesaTranscriptionalRegulator of AnthocyaninBiosynthesis，Modification，andTransport.PlantCell，2003，15：1689-1703.
9.Matsui K，Umemura Y，Ohme-Takagi M.AtMYBL2，a protein with asingleMYBdomain，acts as a negative regulator of anthocyaninbiosynthesisinArabidopsis.Plant J，2008，55：954-967.
10.Outchkourov NS，Carollo CA，Gomez-Roldan V，de Vos RC，BoschD，HallRD，Beekwilder J.Control of anthocyanin and non-flavonoidcompounds byanthocyanin-regulating MYB and bHLH transcriptionfactors in Nicotianabenthamianaleaves.Front Plant Sci，2014，5：519.
11.Petroni K，Tonelli C.Recent advances on the regulationofanthocyaninsynthesis in reproductive organs.PlantSci，2011，181：219-229.
12.Ramsay NA，Glover BJ.MYB-bHLH-WD40protein complex andtheevolutionof cellular diversity.Trends Plant Sci，2005，10：63-70.
13.Rowan DD，Cao M，Lin-Wang K，Cooney JM，Jensen DJ，Austin PT，HuntMB，Norling C，Hellens RP，Schaffer RJ，Allan AC.Environmentalregulation ofleafcolour in red 35S：PAP1Arabidopsis thaliana.NewPhytol，2009，182：102-115.
14.Sapir M，Oren-Shamir M，Ovadia R，Reuveni M，Evenor D，TadmorY，NahonS，Shlomo H，Chen L，Meir A，Levin I.Molecular aspects ofAnthocyanin fruittomato inrelation to high pigment-1.JHered，2008，99：292-303
15.Schaart JG，Dubos C，Romero De La Fuente I，van Houwelingen AM，deVosRC，Jonker HH，Xu W，Routaboul JM，Lepiniec L，Bovy AG.Identificationandcharacterization of MYB-bHLH-WD40regulatory complexescontrollingproanthocyanidinbiosynthesis in strawberry(Fragaria xananassa)fruits.NewPhytol，2013，197：454-467.
16.Vandenbussche F，Tilbrook K，Fierro AC，Marchal K，Poelman D，VanDerStraeten D，Ulm R.Photoreceptor-mediated bending towards UV-BinArabidopsis.Mol Plant，2014，7：1041-1052.
17.Wang H，Fan W，Li H，Yang J，Huang J，ZhangP.FunctionalCharacterizationof Dihydroflavonol-4-Reductase inAnthocyanin Biosynthesisof Purple SweetPotato Underlies the DirectEvidence of Anthocyanins Functionagainst AbioticStresses.PLoSONE，2013，8：e78484.
18.Winkel-Shirley B.Flavonoid Biosynthesis.A ColorfulModelforGenetics，Biochemistry，Cell Biology，and Biotechnology.PlantPhysiol.，2001：(126)：485-493.
19.Yao LH，Jiang YM，Shi J，S-N FA，Datta N，SinganusongR，ChenSS.Flavonoids in Food and Their Health Benefits.Plant FoodsforHumanNutrition，2004，59：113-122.
20.Zhang F，Gonzalez A，Zhao M，Payne CT，Lloyd A.A networkofredundantbHLH proteins functions in all TTG1-dependent pathwaysofArabidopsis.Development，2003，130：4859-4869.
21.Zhang H，He H，Wang X，Wang X，Yang X，Li L，Deng XW.Genome-widemappingof the HY5-mediated gene networks in Arabidopsis thatinvolvebothtranscriptional and post-transcriptional regulation.PlantJ，2011，65：346-358。
SEQUENCE LISTING
<110> 华中农业大学
<120> 超量表达转录因子SlBBX20提高番茄花青素的含量
<130>
<141> 2017-06-28
<160> 3
<170> PatentIn version 3.1
<210> 1
<211> 835
<212> DNA
<213> 番茄（Lycopersicon esculentum）
<400> 1
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cccccactaa agtggggata cctcataata tcaaatggag tacctgttcc acagatcaac 720
tctccatcaa cctagcaact tataggacag acttggtaat aagggcataa tatcttttaa 780
gatattaaga aaagagttat ataaactgcc atctgacctt ttcttttggt tacta 835
<210> 2
<211> 609
<212> DNA
<213> 番茄（Lycopersicon esculentum）
<220>
<221> CDS
<222> (1)..(609)
<223>
<400> 2
atg aag att caa tgt gat gtt tgt gat aaa gaa gag gca tca gtt tat 48
Met Lys Ile Gln Cys Asp Val Cys Asp Lys Glu Glu Ala Ser Val Tyr
1 5 10 15
tgt tca gca gat gaa gcc aca ctt tgc caa agc tgt gat tat caa gtg 96
Cys Ser Ala Asp Glu Ala Thr Leu Cys Gln Ser Cys Asp Tyr Gln Val
20 25 30
cat cat gcc aac aag ctt gca agc aaa cat ctt cgt ttt tct cta att 144
His His Ala Asn Lys Leu Ala Ser Lys His Leu Arg Phe Ser Leu Ile
35 40 45
cat cct tcg ttc aaa gat tct cct ctt tgt gac att tgc cag gaa aga 192
His Pro Ser Phe Lys Asp Ser Pro Leu Cys Asp Ile Cys Gln Glu Arg
50 55 60
cgt gca ttg cta ttt tgt aaa gaa gat aga gca ata ctt tgc aaa gaa 240
Arg Ala Leu Leu Phe Cys Lys Glu Asp Arg Ala Ile Leu Cys Lys Glu
65 70 75 80
tgt gac ttg cct ata cac aaa gca aat gaa cac aca aag aaa cac aac 288
Cys Asp Leu Pro Ile His Lys Ala Asn Glu His Thr Lys Lys His Asn
85 90 95
aga ttt ctt cta agt gga gtg cag cta tct tct gat ata ctt gct tct 336
Arg Phe Leu Leu Ser Gly Val Gln Leu Ser Ser Asp Ile Leu Ala Ser
100 105 110
aat tat aat aat aac caa aat tca ata tcc cca gct gga tct gct gca 384
Asn Tyr Asn Asn Asn Gln Asn Ser Ile Ser Pro Ala Gly Ser Ala Ala
115 120 125
agt aat gct ggt aca aat aat ttt aaa gca ctt agt gga aat ttt ggg 432
Ser Asn Ala Gly Thr Asn Asn Phe Lys Ala Leu Ser Gly Asn Phe Gly
130 135 140
atg aag agt aat tcg att tcg agt act aca gaa tcg aca cat aac tat 480
Met Lys Ser Asn Ser Ile Ser Ser Thr Thr Glu Ser Thr His Asn Tyr
145 150 155 160
ttt cat gtt gat tat gta caa gag ggt tct gtt tca act agt agc ata 528
Phe His Val Asp Tyr Val Gln Glu Gly Ser Val Ser Thr Ser Ser Ile
165 170 175
tca gaa tat ttg act gag act ctt cct ggt tgg cat gtt gaa gat ttt 576
Ser Glu Tyr Leu Thr Glu Thr Leu Pro Gly Trp His Val Glu Asp Phe
180 185 190
ctt gaa tat ccc tct tct tct tcc tat gaa ttt 609
Leu Glu Tyr Pro Ser Ser Ser Ser Tyr Glu Phe
195 200
<210> 3
<211> 203
<212> PRT
<213> 番茄（Lycopersicon esculentum）
<400> 3
Met Lys Ile Gln Cys Asp Val Cys Asp Lys Glu Glu Ala Ser Val Tyr
1 5 10 15
Cys Ser Ala Asp Glu Ala Thr Leu Cys Gln Ser Cys Asp Tyr Gln Val
20 25 30
His His Ala Asn Lys Leu Ala Ser Lys His Leu Arg Phe Ser Leu Ile
35 40 45
His Pro Ser Phe Lys Asp Ser Pro Leu Cys Asp Ile Cys Gln Glu Arg
50 55 60
Arg Ala Leu Leu Phe Cys Lys Glu Asp Arg Ala Ile Leu Cys Lys Glu
65 70 75 80
Cys Asp Leu Pro Ile His Lys Ala Asn Glu His Thr Lys Lys His Asn
85 90 95
Arg Phe Leu Leu Ser Gly Val Gln Leu Ser Ser Asp Ile Leu Ala Ser
100 105 110
Asn Tyr Asn Asn Asn Gln Asn Ser Ile Ser Pro Ala Gly Ser Ala Ala
115 120 125
Ser Asn Ala Gly Thr Asn Asn Phe Lys Ala Leu Ser Gly Asn Phe Gly
130 135 140
Met Lys Ser Asn Ser Ile Ser Ser Thr Thr Glu Ser Thr His Asn Tyr
145 150 155 160
Phe His Val Asp Tyr Val Gln Glu Gly Ser Val Ser Thr Ser Ser Ile
165 170 175
Ser Glu Tyr Leu Thr Glu Thr Leu Pro Gly Trp His Val Glu Asp Phe
180 185 190
Leu Glu Tyr Pro Ser Ser Ser Ser Tyr Glu Phe
195 200
Claims (2)
1.超量表达转录因子SlBBX20基因在提高番茄花青素的含量中的应用，其特征在于，该基因的cDNA序列如SEQ ID NO：1所示。
2.超量表达转录因子SlBBX20基因在提高番茄花青素的含量中的应用，该基因编码的蛋白质序列如SEQ ID NO：3所示。
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| CN112195186B (zh) * | 2020-10-06 | 2022-05-27 | 华中农业大学 | SlBBX20基因在调控番茄灰霉病抗性方面的应用 |
| CN112458103B (zh) * | 2021-01-28 | 2022-09-30 | 青岛农业大学 | 一种调控辣椒红素积累的基因CaBBX20及其应用 |
| CN114656547B (zh) * | 2022-04-01 | 2023-05-23 | 四川农业大学 | 草莓FaBBX21转录因子及其编码蛋白与应用 |
| CN116144694B (zh) * | 2022-09-28 | 2024-08-13 | 西南科技大学 | 一种创制高花色素苷含量材料的方法、应用及制备的材料 |
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