白菜CesA基因家族鉴定及表达模式分析

为探究CesA基因家族在白菜生长发育及纤维素合成过程中的作用机制,该文通过生物信息学的方法,以白菜的全基因组序列为研究区域,进行理化特征、基因结构、进化特征、保守基序及结构域、顺式作用元件和组织表达等鉴定分析。结果表明:(1)共鉴定出16个编码纤维素合成酶亚基的CesA基因,该家族成员所编码蛋白的理论等电点为4.76～9.12,相对分子量为17.76～122.67 kD,长度为153～1 089 aa。(2)15个基因不均匀地分布于白菜的7条染色体上,Bra036008定位于scaffold上。(3)大部分成员包含4～14个外显子,1～11个保守基序。(4)该家族具有保守的DDD-QXXRW 保守功能域。(5)该家族编码蛋白主要分布在质膜上,二级结构以无规则卷曲与α-螺旋为主,多数成员都含有CesA蛋白典型的N端、C端和跨膜区。(6)CesA基因在茎中表达量相对较高,其中Bra011865、Bra023952和Bra029874在茎、叶、花中显著表达。该研究结果为后续深入研究CesA基因功能以及白菜生长发育研究奠定了基础。     

烟草TCP家族成员鉴定及表达分析

TCP家族作为植物特有的转录因子,在植物发育的不同方面发挥着重要作用。为筛选烟草中TCP家族成员,本研究通过全基因组同源比对,鉴定烟草与拟南芥TCP家族同源序列。利用生物信息学的方法分析其理化性质、系统进化关系、顺式作用元件等;筛选AtTCP3/AtTCP4的同源基因,并利用RT-qPCR检测在20% PEG6000处理下的基因表达量变化。结果表明烟草中含有TCP家族成员63个,其氨基酸序列长度范围为89−596 aa,蛋白亲水性(grand average of hydropathicity, GRAVY)范围为−1.147−0.125,等电点(isoelectric point, pI)范围为4.42−9.94,内含子个数为0−3,亚细胞定位均位于细胞核。保守结构域和系统进化关系分析结果表明,烟草TCP家族可分为PCF、CIN和CYC/TB1这3个亚家族且每个亚家族具有稳定序列。基因启动子区顺式作用元件结果表明,TCP家族基因含有低温顺式作用元件(LTR)及多种胁迫及代谢调控相关的元件(MYB、MYC)等顺式作用元件。基因表达模式分析表明,AtTCP3/AtTCP4同源基因(NtTCP6、NtTCP28、NtTCP30、NtTCP33、NtTCP42、NtTCP57和NtTCP63)在20% PEG6000处理下表达量显著上调表达/下调表达,并发现NtTCP30和NtTCP57基因对干旱胁迫响应较为明显。研究结果剖析了烟草基因组中的TCP家族,为烟草抗旱基因功能研究及品种培育提供了候选基因。     

茄子TCP基因家族全基因组的鉴定与分析

TCP (teosinte branched1/cincinnata/proliferating cell factor)转录因子是植物特有转录因子家族,在植物整个生长发育过程中都有着很重要作用。目前,在茄子(Solanum melongena L.)中还没有关于TCP转录因子的相关报道。本研究利用生物信息学方法在茄子基因组数据库中鉴定出分布于11条染色体上的29个茄子TCP家族基因(eggplant TCP,SmTCP)。研究结果显示,该家族所有成员均含有编码TCP保守结构域的序列。这些成员氨基酸长度范围为201–538 aa,外显子数为1或2。亚细胞定位显示,有3个SmTCP (SmTCP02/03/21)蛋白位于细胞质中,其他SmTCP蛋白都位于细胞核中。系统发育树和序列特征分析均将29个SmTCP基因分成ClassⅠ(PCF)和ClassⅡ(CIN和CYC/TB1)两大类。共线性分析发现,17对(21个)SmTCP基因存在共线性关系,且这些存在共线性关系的基因都属于片段复制。基因表达模式分析显示,29个SmTCP基因家族成员在15个组织或器官中都有表达,但是不同成员的表达模式存在差异,其中CIN亚家族的4个基因(SmTCP18/19/20/25)在3个不同生长期的叶片中均较高表达。SmTCP启动子区域的顺式作用元件分析共发现4类顺式作用元件。本研究从多个角度分析了SmTCP基因分子基础,研究了TCP转录因子对茄子生长发育的影响,为茄子分子育种提供理论参考。     

多胺对盐胁迫下黄瓜SOS2基因家族表达的影响

该研究基于黄瓜基因组数据库,利用生物信息学和荧光定量PCR等方法,对黄瓜SOS2基因家族进行全基因组鉴定,并对其表达特性进行系统分析。结果表明：(1)在黄瓜基因组中,共鉴定到 15个SOS2基因(CsSOS2 1~CsSOS2 15),且不均匀分布在5条染色体上;亚细胞定位显示SOS2蛋白主要位于细胞质膜。(2)系统进化分析显示,CsSOS2 2和CsSOS2 6与AtSOS2亲缘关系更近。(3)顺式作用元件预测显示,SOS2基因启动子序列主要含有干旱诱导和防御应激响应元件。(4)结构分析显示,SOS2蛋白所含保守基序的排列顺序完全一致,且主要含有STKc和NAF保守结构域,这些结构可能在基因响应盐胁迫过程中起调控作用。(5)荧光定量试验表明,SOS2基因家族主要在黄瓜的根和叶片响应盐胁迫时上调表达,其中,盐胁迫处理1 d时有5个基因上调表达,并随处理时间延长盐胁迫下的基因表达有所下调;增施多胺显著上调了CsSOS2 1~CsSOS2 5、CsSOS2 8、CsSOS2 9、CsSOS2 12和CsSOS2 15在不同组织中的表达,说明盐胁迫下多胺能诱导黄瓜SOS2基因家族的表达,进而参与植物耐盐分子网络的调控。     

辣椒疫霉MAPK基因家族的鉴定及生物信息学分析

[目的]筛选鉴定辣椒疫霉MAPK基因家族的成员,探讨其基因结构、功能域分布和进化关系。[方法]以辣椒疫霉基因组数据库为平台,搜索MAPK家族成员,并利用生物信息学手段对其基因结构、蛋白功能域及系统进化进行分析。[结果]辣椒疫霉MAPK家族包含16个基因,且在基因组中随机分布,每个基因都含有高度保守的激酶结构域;其中5个不含内含子,4个为非典型磷酸化唇序列,3个含有PH、WW、EF-H等与细胞信号转导相关的功能域;在进化关系上,辣椒疫霉MAPK较真菌相对独立。[结论]首次对辣椒疫霉MAPK基因家族进行鉴定和生物信息学分析,该家族共有16个成员,含有激酶结构域、磷酸化唇序列及信号转导相关蛋白功能域,进化上独立于真菌。     

香蕉ADA1家族成员鉴定及其在生物和非生物胁迫下的表达分析

Spt-Ada-Gcn5-乙酰转移酶(Spt-Ada-Gcn5-acetyltransferase, SAGA)是高度保守的辅助转录起始复合物,转录接头蛋白-激活的改变/缺失亚基1 (alteration/deficiency in activation 1, ADA1),也称作组蛋白H2A功能互作因子1 (histone H2A functional interactor 1, HFI1),它是SAGA核心模块中的一个亚基,在植物的生长发育和抗逆性方面发挥着重要的作用。为了解香蕉ADA1的分子特性,本研究基于香蕉基因组数据库,对香蕉ADA1基因家族成员进行鉴定,分析其基本理化性质、系统进化、选择压力、启动子顺式作用元件及生物与非生物胁迫下的表达等。结果显示,香蕉A、B及阿宽蕉基因组中分别有10、6、7个ADA1家族成员;成员均为不稳定的亲水性蛋白,均保守地含有SAGA-Tad1结构域,MaADA1和MbADA1均可与SAGA核心模块中的SAGA相关因子11 (SAGA-associated factor 11, Sgf11)互作;系统发育显示香蕉ADA1基因家族成员可划分为3个亚族,进化过程中大多受纯化选择;香蕉ADA1基因家族成员的基因结构差异性较大;香蕉ADA1基因家族成员含有多个响应激素的作用元件;MaADA1-1可能对香蕉在低温胁迫下的抗性起着重要的作用,MaADA1均响应香蕉枯萎病菌胁迫。本研究表明,ADA1基因家族成员在香蕉中高度保守,并可能响应生物与非生物胁迫。     

Dof基因家族调节植物生长发育功能的研究进展

Dof（DNA binding with one finger）蛋白是一类植物特异性转录因子,通常含有200～400个氨基酸和2个主要结构域。该家族成员的N 末端为高度保守的单锌指Dof结构域,具有与DNA和蛋白质相互作用的双重功能,其C末端的氨基酸序列则较为多变,是Dof蛋白重要的特异转录调控结构域。研究表明,Dof蛋白作为转录激活物或阻遏物参与了多方面的植物生长发育过程。随着基因组测序技术的发展,已有大量的Dof基因从植物基因组数据库中鉴定出来。该文对近年来国内外有关Dof基因家族的结构特点、全基因组鉴定、蛋白互作以及生物学功能等方面的研究进展进行综述,以期为Dof转录因子的深入研究提供参考。     

糜子黄、黑种皮遗传及转录组学分析

同源异型域-亮氨酸拉链(homedomain-leucine zipper,HD-Zip)转录因子广泛参与植物的生长发育和抗胁迫过程。该研究通过生物信息学方法对青稞HD-Zip基因家族进行全基因组分析鉴定,并采用qRT-PCR技术分析非生物胁迫下该基因的表达特性,为深入探讨青稞HD-Zip转录因子的生物学功能及其在高原作物抗逆育种中的应用奠定基础。结果表明：(1)成功从青稞基因组中共鉴定出41个HD-Zip基因家族成员,依次命名为i>HvvHD-ZipⅠ-1~Ⅳ-13,且这些基因在7条染色体上呈不均匀分布。(2)理化性质分析发现,HvvHD-Zip蛋白包含197～885个不等的氨基酸残基;分子量范围在19 914.36～94 014.87 Da;亚细胞定位表明HvvHD-Zip蛋白都位于细胞核。(3)根据多序列比对、系统进化、基因结构和保守基序差异将其聚为4个亚家族,各亚家族分类特征与系统聚类结果一致。(4)顺式作用元件预测分析发现,i>HvvHD-Zip基因启动子中含有11种植物激素和胁迫响应元件。(5)qRT-PCR结果显示,HvvHD-Zip Ⅰ、Ⅱ、Ⅳ亚家族基因对各胁迫响应明显;与根组织相比,多数i>HvvHD-Zip基因在叶组织中响应明显(上调或下调);与冷和盐胁迫相比,i>HvvHD-Zip各基因对旱胁迫响应较强。     

垫状卷柏SpLEA1基因克隆及干旱胁迫下的表达分析

晚期胚胎发育丰富蛋白(late embryogenesis abundant,LEA),广泛存在于生物体内,与植物抗逆性密切相关,可在干旱胁迫下保护植物细胞,减少植物损伤。垫状卷柏(Selaginella pulvinata)是一种在干旱胁迫下生存能力极强的蕨类植物,具有很强的恢复能力。为探究垫状卷柏SpLEA1基因在耐旱植物中的分子机制与表达特征,该研究以高耐旱性植物垫状卷柏为实验材料,基于转录组测序结果,采用RT-PCR技术获得SpLEA1基因cDNA序列,采用HiTail-PCR技术获得启动子序列,利用生物信息学对序列进行了分析,并采用qRT-PCR技术分析了SpLEA1基因在干旱胁迫下的表达模式。结果表明:(1)垫状卷柏SpLEA1全长为476 bp,开放阅读框(ORF)为279 bp,共编码92个氨基酸,通过在线工具预测到蛋白分子量为9 491.46 Da, 等电点为5.45,蛋白结构预测分析表明该蛋白为亲水性蛋白,含有10个磷酸化位点,二级结构以α-螺旋和无规则卷曲为主。(2)预测到SpLEA1蛋白的保守结构域为Lea-5,来源于LEA1家族。基于系统发生树和遗传距离矩阵,发现垫状卷柏SpLEA1与来自鹰嘴豆(Cicer arietinum )和红车轴草(Trifolium pratense)的Lea-5蛋白同源性较高。(3)对启动子序列进行顺式作用元件的预测分析发现SpLEA1基因启动子含有5类激素响应元件和与干旱胁迫响应有关的功能元件。(4)在自然干旱处理下SpLEA1基因表达上调,并在12 h时达到峰值,在24 h干旱后进行复水处理,表达量显著下调。综上所述,SpLEA1基因在垫状卷柏中很可能参与了干旱胁迫响应机制的相关调控。该结果为进一步研究垫状卷柏SpLEA1基因在干旱胁迫下的功能及其表达调控机制提供了参考。     

葡萄Alfin like转录因子家族的鉴定和表达分析

Alfin like (AL) 转录因子家族对高盐、低温、干旱等非生物胁迫反应具有重要的调控作用。该研究采用同源比对的方法检索鉴定葡萄AL转录因子家族基因、分析其生物信息学特性,并采用qRT PCR方法分析非生物胁迫下AL基因的表达特征,以探究葡萄AL基因在非生物逆境胁迫中的功能。 结果表明：(1)在葡萄中共鉴定出6个AL基因家族成员,分别命名为VvAL1~VvAL6,且6个成员分别分布在6条染色体上。(2)葡萄中AL转录因子具有高度保守的DUF3594结构域和PHD结构域,各家族成员均含有5个外显子和4个内含子;上游启动子区域分析发现大量植物激素与非生物胁迫响应相关的顺式作用元件。(3)基因芯片表达模式分析显示,盐、干旱、ABA胁迫以及低温(5 ℃)处理均显著影响葡萄AL家族基因(VvAL1~VvAL6)的表达。(4)qRT PCR检测显示,不同胁迫处理下AL基因在葡萄叶片中的表达水平不同;ABA处理下葡萄AL转录因子家族基因表达量较对照均显著下调,但在PEG处理下差异不显著;在盐胁迫处理下,VvAL2、VvAL4、VvAL5基因的表达量均显著上调,分别是对照的23倍、8.5倍和10.5倍,而VvAL1和VvAL6基因的表达量均显著下调,分别是对照的33倍和25倍。研究发现,葡萄AL转录因子家族与植物激素和非生物胁迫密切相关,尤其是该家族基因强烈响应高盐胁迫。     

棉花DUR3基因的鉴定及进化分析

植物DUR3同源蛋白属于钠离子/溶质共运蛋白家族的尿素高亲和力运输蛋白，在植物体对外源尿素的主动吸收及内源尿素的再分配过程中具有重要作用。为明确棉花DUR3基因的结构和进化情况，基于生物信息学的方法，从全基因组水平鉴定陆地棉和雷蒙德氏棉的DUR3基因，并对基因结构、跨膜结构域、基序分布、进化关系等进行分析。结果表明：(1)从陆地棉A亚组和D亚组染色体各鉴定出1个DUR3基因，从雷蒙德氏棉基因组鉴定出1个DUR3基因。这3个棉花DUR3同源蛋白同其他植物DUR3同源蛋白一样，具有15个跨膜结构域，具有3个位置一致、高度保守的基序。(2)基因结构分析表明，双子叶植物DUR3基因的外显子个数明显多于单子叶植物，这3个棉花DUR3基因的外显子个数亦是如此。(3)根据物种间种属亲缘关系，对不同物种DUR3氨基酸序列构建的进化树显示，棉花的同双子叶植物的聚在一起。(4)DUR3直系同源基因和旁系同源基因的K_a/K_s比值普遍均大于1,说明这些基因在进化过程中主要受到正向选择的作用。该研究结果为深入研究棉花DUR3同源蛋白提供了理论基础。     

Genome-Wide Identification and Analysis of Genes Encoding Proteolytic Enzymes in Pineapple

Pineapple, Ananas comosus, is an economically important fruit crop. Recently its genome was completely sequenced and a total of 27,024 protein coding genes were predicted. Using a set of well evaluated bioinformatics tools we have predicted the protein subcellular locations and comparatively analyzed the protein conserved domains of the predicted proteomes in pineapple, Oryza sativa (rice), Sorghum bicolor (sorghum), and Brachypodium distachyson. Our analysis revealed that ~24–26 % of proteins were located in nucleus, 17–21 % in cytosol, 9–11 % in chloroplast, and 8–11 % proteins were secreted in these monocot plants. The secretomes in the four species were analyzed comparatively and a large number of secreted glycosyl hydrolases were identified. As pineapple proteolytic enzymes, knowns as bromelains, have been used for medical treatments, we focused on genome-wide identification and analysis of pineapple genes encoding proteases. A total of 512 pineapple genes encoding putative proteolytic enzymes were identified, with 152 secreted, 74 localized in cytosol, 67 in nucleus, 60 in chloroplast, 18 in mitochondria, and the remaining in other subcellular locations. The top large protease families in pineapple were papain family cysteine protease (62 genes), peptidase S8 family (56 genes), aspartyl protease family (38 genes), and serine carboxypeptidase (33 genes). Gene expression analysis revealed that among 512 protease genes 432 were expressed in various tissues and 72 genes were differentially expressed. The highly expressed protease genes were identified including 7 papain family cysteine proteases. The protease genes with the predicted protein subcellular locations will facilitate the efforts for examining their biological roles in pineapple growth and development and for expressing the recombinant proteases for medical use. The information of protein subcellular location of all plant species can be accessed at the PlantSecKB website (http://proteomics.ysu.edu/secretomes/plant.php).     

Pineapple translation factor SUI1 and ribosomal protein L36 promoters drive constitutive transgene expression patterns in Arabidopsis thaliana

The availability of a variety of promoter sequences is necessary for the genetic engineering of plants, in basic research studies and for the development of transgenic crops. In this study, the promoter and 5′ untranslated regions of the evolutionally conserved protein translation factor SUI1 gene and ribosomal protein L36 gene were isolated from pineapple and sequenced. Each promoter was translationally fused to the GUS reporter gene and transformed into the heterologous plant system Arabidopsis thaliana. Both the pineapple SUI1 and L36 promoters drove GUS expression in all tissues of Arabidopsis at levels comparable to the CaMV35S promoter. Transient assays determined that the pineapple SUI1 promoter also drove GUS expression in a variety of climacteric and non-climacteric fruit species. Thus the pineapple SUI1 and L36 promoters demonstrate the potential for using translation factor and ribosomal protein genes as a source of promoter sequences that can drive constitutive transgene expression patterns.     

Three different genes encode the iron-sulfur subunit of succinate dehydrogenase in Arabidopsis thaliana

The iron-sulfur protein is an essential component of mitochondrial complex II (succinate dehydrogenase, SDH), which is a functional enzyme of both the citric acid cycle and the respiratory electron transport chain. This protein is encoded by a single-copy nuclear gene in mammals and fungi and by a mitochondrial gene in Rhodophyta and the protist Reclinomonas americana. In Arabidopsis thaliana, the homologous protein is now found to be encoded by three nuclear genes. Two genes (sdh2-1 andsdh2-2) likely arose from a relatively recent duplication event since they have similar structures, encode nearly identical proteins and show similar expression patterns. Both genes are interrupted by a single intron located at a conserved position. Expression was detected in all tissues analysed, with the highest steady-state mRNA levels found in flowers and inflorescences. In contrast, the third gene (sdh2-3) is interrupted by 4 introns, is expressed at a low level, and encodes a SDH2-3 protein which is only 67% similar to SDH2-1 and SDH2-2 and has a different N-terminal presequence. Interestingly, the proteins encoded by these three genes are probably functional because they are highly conserved compared with their homologues in other organisms. These proteins contain the cysteine motifs involved in binding the three iron-sulfur clusters essential for electron transport. Furthermore, the three polypeptides are found to be imported into isolated plant mitochondria.     

小麦锌指蛋白基因TaLOL2的克隆及特征分析

通过电子克隆与RT-PCR相结合的方法,在条锈菌诱导的小麦叶片中克隆获得1个新的LSD1型锌指蛋白基因TaLOL2,并用qRT-PCR技术分析了其转录表达特征。结果显示:(1)小麦锌指蛋白基因TaLOL2的cDNA全长1 095bp,编码179个氨基酸。(2)TaLOL2含有3个典型的zf-LSD1型(CxxCxRxxLMYxxGASxVxCxxC)保守结构域,与水稻、拟南芥、大麦等植物LSD1型锌指蛋白序列具有高度相似性,其中与水稻OsLOL2相似度达86.0%。(3)进化树分析表明,TaLOL2与水稻、拟南芥和大麦中部分含有3个保守zf-LSD1锌指结构的基因亲缘关系较近,而与其它包含不同数目的zf-LSD1锌指结构的基因亲缘关系较远。(4)qRT-PCR定量分析表明,TaLOL2在条锈菌侵染前期呈上调表达,在亲和及非亲和反应中差异表达。研究表明,TaLOL2参与了条锈菌诱导的小麦抗病防卫反应,很可能作为正调控因子参与了小麦-条锈菌非亲和互作中对条锈菌的抗性信号途径。     

Analysis of the <Emphasis Type="Italic">MAT1-2-1</Emphasis> gene of <Emphasis Type="Italic">Colletotrichum lindemuthianum</Emphasis>

A single MAT1-2-1 gene was identified from a mating pair of the filamentous ascomycete Colletotrichum lindemuthianum. The MAT1-2-1 genes from both mating partners carried an open reading frame (ORF) of 870 bp encoding a putative protein of 290 amino acids that includes the highly conserved high mobility group (HMG) domain typical of the fungal MAT1-2-1 genes. Three introns were confirmed within the C. lindemuthianum ORF, two of which were found to be conserved relative to a previously reported MAT1-2-1 gene from C. gloeosporioides. The amino acid sequence of the HMG domain from C. lindemuthianum MAT1-2-1 was also compared with those from other ascomycetes. These results suggest that although the MAT1-2-1 genes are highly conserved among ascomycetes, the mechanism which defines mating partners in the genus Colletotrichum is distinct to the idiomorph system described for other members of this phylum.     

Genome-Wide Identification,Evolution and Expression Analyses of <i>GA2ox</i> Gene Family in <i>Brassica napus</i> L.

Gibberellin 2-oxidases (GA2ox) are important enzymes that maintain the balance of bioactive GAs in plants. GA2ox genes have been identified and characterized in many plants, but these genes were not investigated in Brassica napus. Here, we identified 31 GA2ox genes in B. napus and 15 of these BnaGA2ox genes were distributed in the A and C subgenomes. Subcellular localization predictions suggested that all BnaGA2ox proteins were localized in the cytoplasm, and gene structure analysis showed that the BnaGA2ox genes contained 2–4 exons. Phylogenetic analysis indicated that BnGA2ox family proteins in monocotyledons and dicotyledons can be divided into four groups, including two C₁₉-GA2ox and two C₂₀-GA2ox clades. Group 4 is a C₂₀-GA2ox Class discovered recently. Most BnaGA2ox genes had a syntenic relationship with AtGA2ox genes. BnaGA2ox genes in the C subgenome had experienced stronger selection pressure than genes in the A subgenome. BnaGA2ox genes were highly expressed in specific tissues such as those involved in growth and development, and most of them were mainly involved in abiotic responses, regulation of phytohormones and growth and development. Our study provided a valuable evolutionary analysis of GA2ox genes in monocotyledons and dicotyledons, as well as an insight into the biological functions of GA2ox family genes in B. napus.     

小立碗藓扩展蛋白基因家族的鉴定与生物信息学分析

扩展蛋白(Expansins,EXP)是一类基因家族,几乎参与了植物发育的全过程,从种子萌发到果实成熟都有扩展蛋白的参与。该研究利用生物信息学的方法对小立碗藓(Physcomitrella patens) Expansin基因家族成员进行鉴定,分析了其基因结构、染色体定位以及系统发生关系。结果表明:小立碗藓基因组中含有Expansin A(EXPA) 32个、Expansin-like A(EXLA) 6个,并未发现Expansin-like B(EXLB)及Expansin B(EXPB)。扩展蛋白氨基酸序列长度在228～290 aa之间,编码蛋白质具有两个保守的结构域Pollen_allerg_1和DPBB_1。蛋白质亚细胞定位预测结果表明:运用CELLO在线工具预测发现小立碗藓中约4/5的EXP家族基因定位于细胞外;而Euk-mPLoc预测结果则显示小立碗藓EXP基因家族成员全定位于细胞外。基因结构分析表明,小立碗藓中约68%Expansin基因有含有1～3个内含子。以上结果可为深入研究小立碗藓扩展蛋白基因的分子进化与生物学功能奠定基础。