海岛棉GbTCP14基因的克隆与表达分析

以海岛棉‘新海21号’为试验材料,根据陆地棉GhTCP14基因序列,设计1对引物,通过RT-PCR技术获得海岛棉GbTCP14核苷酸序列,开放阅读框长1 221bp,编码406个氨基酸,分子式C1892H2950N572O620S16,预测分子量为44.134 6kD,等电点为6.88,氨基酸序列包含1个高度保守的TCP结构域及4个低丰度复杂区。GbTCP14蛋白氨基酸序列与其他物种TCP14氨基酸序列比对表明,海岛棉GbTCP14蛋白与其他植物中的TCP14蛋白共同具有一个高度保守的TCP结构域,序列之间的一致性较高。进化树分析表明,海岛棉GbTCP14基因与木本棉GaTCP14基因分布在同一分支上。实时荧光定量PCR表明,海岛棉GbTCP14基因在开花后第15天时表达量最高,在第5~20天时相对表达量比其他时期较高,第25天时相对表达量最低。在不同组织中花瓣和花萼中表达量较高,根和茎中表达量一般,叶组织中表达量最低。通过酵母系统转录激活分析显示,GbTCP14蛋白不具有转录活性。     

海岛棉GbTCP15基因的克隆与表达分析

根据陆地棉GhTCP15基因序列,设计1对引物,通过PCR技术从海岛棉品种‘新海21号’中克隆一个同源基因,命名为GbTCP15。海岛棉GbTCP15基因具有一个1 056 bp开放阅读框,编码351个氨基酸,预测分子量约为38 057.4 kD,等电点为9.01,序列中含有一个高度保守的TCP结构域。氨基酸序列对比表明,海岛棉GbTCP15蛋白与其他植物中的TCP蛋白有较高的一致性,说明该基因在进化过程中是相当保守的。亚细胞定位结果显示,GbTCP15主要分布在细胞核上,推测GbTCP15在复制和转录的过程中发挥着信号转导以及转录调控等作用。进化树分析表明,海岛棉GbTCP15基因与雷蒙德氏棉GrTCP15基因分布在同一分支上。实时荧光定量PCR表明,海岛棉GbTCP15基因在茎部和15 d纤维中表达量较高。研究表明,GbTCP15转录因子可能参与棉花纤维以及表皮毛的发育。     

海岛棉GbHCT基因克隆及生物信息学分析

克隆海岛棉木质素合成关键酶基因GbHCT全长cDNA序列,分析其在纤维不同发育时期的表达情况。根据海岛棉转录组数据中的HCT序列设计引物,采用RT-PCR技术克隆基因,通过生物信息学方法分析GbHCT的cDNA序列和氨基酸序列。通过分析转录组数据研究GbHCT在纤维不同发育时期的表达。从海岛棉中克隆了1个编码HCT的基因GbHCT,开放阅读框长度为1311bp,编码436个氨基酸,蛋白质分子量为48.58kD,等电点为6.03。GbHCT氨基酸序列中含有HTLGD和DFGWG2个保守基序。GbHCT氨基酸与陆地棉、亚洲棉和雷蒙德氏棉HCT一致性较高,与其他植物中的HCT氨基酸的一致性为55.7%-60.7%。转录组数据分析发现GbHCT在纤维不同发育阶段都有表达,在5DPA的纤维中表达量最高,表明该基因可能参与棉花纤维的发育。     

海岛棉GAUT基因家族的鉴定及其在棉纤维发育中的表达分析

本研究鉴定了海岛棉全基因组中的GAUT基因家族,对其进行了生物信息学及纤维发育过程中的表达分析。结果表明,海岛棉GAUT家族包含37个成员,分为3个亚组,分布在海岛棉18条染色体上。GAUT氨基酸序列保守基序有4个,保守性较强,所有GAUT蛋白均定位在高尔基体膜。根据GAUTs在海岛棉纤维发育不同时期的表达变化,将其分为起始期高表达、纤维伸长期高表达、次生壁增厚期高表达、全时期低表达等4类模式。其中,GAUT05、GAUT06、GAUT07、GAUT23、GAUT24、GAUT26等在纤维发育某个时期具有优势表达的基因可能在纤维发育过程中发挥着重要作用。上述结果为研究海岛棉GAUT基因家族在棉纤维发育中的功能提供了参考依据。     

海岛棉WRKY转录因子的全基因组鉴定及响应黄萎病菌侵染的表达分析

WRKY蛋白是植物中一类重要的转录因子,其在很多生物过程中都发挥有重要作用。目前,关于海岛棉(Gossypium barbadense L.)WRKY转录因子的研究相对较少。本研究利用生物信息学方法在海岛棉基因组中鉴定出180个WRKY转录因子(GbWRKY)。根据基因结构及系统进化关系,可将GbWRKY基因分为3组(I～III),II组又可细分为5个亚组(IIa～IIe)。GbWRKY基因在染色体上分布不均匀。基因重复事件分析表明,全基因组复制及区段重复事件是GbWRKY基因数量增多的主要原因。表达分析发现,有61个GbWRKY基因在黄萎病菌(Verticillium dahliae)侵染条件下呈现差异表达,表明它们在黄萎病菌胁迫应答过程中发挥作用。本研究结果揭示了海岛棉WRKY转录因子的结构、进化及表达特征,为进一步研究棉花WRKY基因的功能提供了理论指导。     

四倍体海岛棉LTR反转录转座子的数量与分布

棉花是重要的纤维作物,在四个栽培棉种中,海岛棉纤维品质最优,了解LTR反转录转座子的数量与分布,可以促进海岛棉基因组的研究。通过综合不同方法挖掘海岛棉基因组中的LTR反转录转座子序列,并进行家族归类和数据分析。结果表明海岛棉A亚组和D亚组共有的LTR反转录转座子家族占全部家族的95%,LTR反转录转座子的Copia超家族和Gypsy超家族的分布特征有明显的不同,但相同超家族在相同亚组染色体上则表现出相似的特征。LTR反转录转座子周边基因的GO注释主要富集在结合活性、催化活性和代谢过程等方面。研究结果揭示了LTR反转录转座子在海岛棉染色体上的分布特征及其周边基因的功能富集。     

马铃薯低氮肥胁迫响应TCP转录因子的鉴定与分析

TCP转录因子是植物特有的一类转录因子,参与植物生物学过程的多个方面。为研究马铃薯TCP转录因子在响应低氮肥胁迫中的作用,该研究以氮肥供应不足(0.05 mmol·L^-1)和氮肥供应充足(7.5 mmol·L^-1)条件下马铃薯的根和叶片构建4个转录组文库进行测序,并对差异表达的TCP转录因子进行分析。结果表明:(1)在4个转录组文库中共鉴定TCP转录因子24个,它们主要分布在2号、3号、6号染色体上。(2)经结构域分析显示,24个TCP 转录因子均具有典型的basic-Helix-Loop-Helix结构域。(3)经系统进化分析显示,马铃薯与拟南芥TCP蛋白可聚集在一起,分属于10个亚类。(4)转录组测序结果显示,在低氮肥胁迫下,大多数TCP转录因子被抑制表达,有3个TCP转录因子在根中显著性差异表达,5个TCP转录因子在叶中特异性表达。(5)根据GO功能注释分析和马铃薯TCP转录因子与拟南芥TCP转录因子的亲缘关系分析推测,这些TCP转录因子参与了马铃薯对低氮肥胁迫的响应。该研究结果为进一步研究马铃薯与其他粮食作物TCP转录因子响应低氮肥胁迫的分子功能奠定了基础。     

海岛棉抗枯萎病基因GbMPK16的克隆及功能验证

研究MAPK基因在海岛棉抗枯萎病途径中的功能,为棉花枯萎病抗性育种提供基因资源。根据前期海岛棉枯萎病抗性转录组测序和表达谱数据分析,从差异表达基因中筛选到一个海岛棉抗枯萎病相关Unigene序列,该序列注释为MAPK相关基因。从海岛棉抗病品种"06-146"中克隆了一个MAPK家族基因GbMPK16,进行生物信息学分析,并在枯萎病菌、乙烯、水杨酸诱导下对该基因进行表达分析,通过VIGS技术进行该基因抗病性功能分析。研究显示GbMPK16基因有1 665 bp的ORF序列,编码554个氨基酸,属于MAPK家族D组,保守区域包括TDY磷酸化部位,GbMPK16蛋白与其他生物MPK16蛋白有较高的一致性。进化树分析表明,GbMPK16与GhMPK16亲缘关系最近。亚细胞定位预测表明,GbMPK16分布于细胞核的可能性46.3%,在细胞质里的可能性40.7%。qRT-PCR表达分析显示,GbMPK16基因在不同抗病海岛棉品种表达量明显高于感病海岛棉品种,乙烯和水杨酸处理后出现表达量上调趋势,并且在抗病材料中该基因表达量均高于感病材料。通过VIGS技术沉默GbMPK16基因后,与对照相比该基因的表达量明显下降,枯萎病侵染15 d后,沉默该基因的植株发病情况高于对照。GbMPK16基因可能在海岛棉抗枯萎病途径中发挥作用。     

胡萝卜DcPAB基因的分离及其结构与功能分析

运用改进的减法杂交技术分离到胡萝卜Poly(A)结合蛋白基因DcPAB .其cDNA编码区长度为 1 977bp ,编码 6 5 8个氨基酸和 1个终止密码子 .基因组转录序列区长度为 4 6 1 6bp ,包含 9个外显子和 8个内含子 .DcPAB在胡萝卜基因组中为单拷贝基因 .该基因在胡萝卜体细胞胚中特异性表达 ,且其表达活性在调控 解调控前后有明显差异 .体外结合实验表明 ,在大肠杆菌中表达并纯化的DcPAB蛋白具有与oligo(A) 2 0 特异性结合的性能 .酵母突变体互补实验进一步证明 ,该基因可以互补PAB基因缺失的酵母突变体的功能缺陷     

人受精促进肽受体基因（TCP11b）的剪接、克隆和差别表达

运用同源比较和PCR法 ,从人睾丸组织中分离了人受精促进肽受体TCP11基因的一个新的剪切体TCP11b ,它编码 5 0 3个氨基酸的蛋白质 ,与TCP11a相比 ,在基因组的 5′端存在复杂的外显子剪接现象。运用荧光原位杂交 (FISH)方法 ,显示该基因定位到人染色体 6p2 1。Northern杂交及多组织RT PCR的结果显示该转录本在正常睾丸中表达 ,而其他组织、无精症患者及胎儿睾丸组织中未见该基因的表达。该结果结合mTcp 11功能的提示 ,TCP11b这种转录本对精子发生和人受精过程可能起重要作用。     

Effects of MULTIFOLIATE-PINNA, AFILA, TENDRIL-LESS and UNIFOLIATA genes on leafblade architecture in Pisum sativum

In order to dissect the genetic regulation of leafblade morphogenesis, 16 genotypes of pea, constructed by combining the wild-type and mutant alleles of MFP, AF, TL and UNI genes, were quantitatively phenotyped. The morphological features of the three domains of leafblades of four genotypes, unknown earlier, were described. All the genotypes were found to differ in leafblade morphology. It was evident that MFP and TL functions acted as repressor of pinna ramification, in the distal domain. These functions, with and without interaction with UNI, also repressed the ramification of proximal pinnae in the absence of AF function. The expression of MFP and TL required UNI function. AF function was found to control leafblade architecture multifariously. The earlier identified role of AF as a repressor of UNI in the proximal domain was confirmed. Negative control of AF on the UNI-dependent pinna ramification in the distal domain was revealed. It was found that AF establishes a boundary between proximal and distal domains and activates formation of leaflet pinnae in the proximal domain.     

Presence of Rhizobium Etli bv . Phaseoli and Rhizobium Gallicum bv . Gallicum in Egyptian Soils

Seven bean rhizobial strains EBRI 2, 3, 21, 24, 26, 27 and 29 identified as Rhizobium etli, and EBRI 32 identified as Rhizobium gallicum, isolated from Egyptian soils and which nodulated Phaseolus vulgaris efficiently, were subjected to hybridization with a nifH probe in order to estimate the copy number of this gene. Seven strains (EBRI 2, 3, 21, 24, 26, 27 and 29) which were only able to nodulate Phaseolus vulgaris, contained three copies of the nifH gene, consistent with their identification as Rhizobium etli bv. phaseoli. Only one strain (EBRI 32) which nodulated both Phaseolus vulgaris and Leucaena leucocephala, had one copy of nifH gene. This confirmed the classification of this strain as Rhizobium gallicum bv. gallicum.     

Prosthecium species with Stegonsporium anamorphs on Acer

Data from microscopic morphology, single-spore cultures, and DNA analyses of teleomorphs and anamorphs support the recognition of five species of Prosthecium with Stegonsporium anamorphs on Acer: P. acerinum sp. nov., the teleomorph of S. acerinum; P. acerophilum comb. nov., formerly known as Dictyoporthe acerophila; P. galeatum comb. nov., originally described as Massaria galeata; P. opalus sp. nov.; and P. pyriforme sp. nov., the teleomorph of S. pyriforme s. str. The morphology of both type specimens and freshly collected material was investigated. The teleomorphs have brown ellipsoidal ascospores with five distosepta and often a longitudinal distoseptum. The anamorphs of all species described here belong to Stegonsporium; their connection to the Prosthecium teleomorphs was demonstrated by morphology and DNA sequences of single spore cultures derived from both ascospores and conidia. The anamorphs and teleomorphs of all five Prosthecium species are described and illustrated by LM images, and a key to these species is provided. As perceived from this work, S. pyriforme is restricted to Europe and does not occur in North America, whereas S. acerinum is restricted to North America, not found in Europe. The host associations given in the literature are revised and evidence is provided that only A. opalus, A. pseudoplatanus, and A. saccharum are confirmed hosts of Prosthecium with Stegonsporium anamorphs. Molecular phylogenetic analyses of tef1, ITS rDNA, and partial nuLSU rDNA sequences confirm that the species with Stegonsporium anamorphs are closely related to P. ellipsosporum, the generic type species. Stilbospora macrosperma is confirmed as the anamorph of P. ellipsosporum by DNA data of single spore isolates obtained from both ascospores and conidia.     

Characterisation of cell wall polysaccharides, arabinogalactans-proteins (AGPs) and phenolics of Cola nitida, Cola acuminata and Garcinia kola seeds

Many Cola plant species are endemic to West and Central Africa. Cola acuminata and Cola nitida are used as masticatory when fresh, while the dried nuts are used for beverages and pharmaceutical purposes in Europe and North America. Garcinia kola seeds, that serve as a substitute for the true kola nuts, are used in African traditional medicine for the treatment of various diseases, including colic, headache and liver cirrhosis. Seeds extracts of G. kola are also known for their anti-inflammatory, antimicrobial and antiviral properties. To gain information on the chemical properties of the kolas, we have isolated and analyzed cell wall polysaccharides, arabinogalactan-proteins and phenolic substances from the seeds of the three kola species. The sugar composition of cell wall material of C. acuminata, C. nitida and G. kola revealed that Gal (up to 30%), Ara, GalA and Glc as the predominant monosaccharides, representing approximately 90% by mol of the total hydrolysable sugar present in this material. In Ammonium oxalate cell wall fraction, GalA was found to be the major sugar present in all kola species. In the alkali-soluble fraction, there were significant differences in the level of Glc and Gal. The level of Glc was high in C. acuminata and C. nitida while the level of Gal and Xyl were high in C. nitida and G. cola. Isolation and quantification of arabinogalactan-proteins demonstrate that G. kola seeds contained four to eight times more of these proteoglycans than the seeds of the other two species. Finally, analysis of soluble phenolic substances shows that caffeine and catechin were largely represented in C. acumina and C. nitida seeds, with caffeine accounting for 50% of all soluble phenolics. These findings indicate that the three Kola seeds are highly enriched in pectins and proteoglycans and that C. acuminata and C. nitida can be used as a possible source of caffeine and catechin.     

中国黑粉菌属和利罗黑粉菌属

黑粉菌属是Roussel 1806年建立的,全世界记载有三百余种,主要寄生于禾本科,是经济作物及牧草的重要致病菌·长期以来,对黑粉菌的邢子使用过各种名称,如厚垣孢子,冬孢子及黑粉孢子等.本文采用黑粉孢子以区别锈菌的冬孢子. 芳’(1979)在《中国真菌总汇》中列出黑粉菌属五十种及一个变型.作者经过显微结构和超显微结构的研究,承认其中二十九种为正确名称,八种及一变型为异名,顶黑粉菌(Ustilago acrearus Berk.)由于错拼而被废弃.埃地黑粉菌(Ustilago emodensis Berk.)被转移至利罗粉菌属(Liroa).另有十一种黑粉菌因缺少标本留待今后订正.自1979年以后,杨信东(1983)增加黑粉菌属二种我国新纪录,K.范基和郭林(1986)描述一新种,四种新纪录.在本文中,作者描述一新种:鸢尾蒜黑粉(Ustilago ixiolirii Guo L) ,孢子堆生在蒴果内,不开裂,黑色,粉末状.黑粉孢子球形,近球形,稀椭圆形, 12.5-21×10-21μm,黑褐色,壁厚1-1.Sμm,纹饰脑状.是迄今生在石蒜科植物上唯一黑粉菌的种,其它几种黑粉菌均属条黑粉菌属.本文增加七种我国新纪录.共计四十九种,寄生于六科四十四属植物,主要是禾本科和蓼科.这仅是黑粉菌属研究的初步报告,在全国范围内大量采集黑粉菌标本后,作者相信会有更多新种和我国新纪录被发现.利罗黑粉菌属(Liroa)是从黑粉菌属(Ustaligo)分出的,此属为单种属.     

Comparative study of Bifidobacteriumanimalis, Escherichiacoli, Lactobacilluscasei and Saccharomycesboulardii probiotic properties

The present work investigates some probiotic properties of four different microorganisms (Bifidobacterium animalis var. lactis BB-12, Escherichia coli EMO, Lactobacillus casei and Saccharomyces boulardii). In vitro and in vivo tests were carried out to compare cell wall hydrophobicity, production of antagonistic substances, survival capacity in the gastrointestinal tract of germ-free mice without pathological consequence, and immune modulation by stimulation of Küpffer cells, intestinal sIgA and IL-10 levels. In vitro antagonism against pathogenic bacteria and yeast was only observed for the probiotic bacteria B. animalis and L. casei. The hydrophobic property of the cell wall was higher for B. animalis and E. coli EMO, and this property could be responsible for a better ability to colonize the gastrointestinal tract of germ-free mice. Higher levels of sIgA were observed mainly for S. boulardii, followed by E. coli EMO and B. animalis, and only S. boulardii induced a significant higher level of IL-10. In conclusion, for a probiotic use, S. boulardii presented better characteristics in terms of immunomodulation, and B. animalis and L. casei for antagonistic substance production. The knowledge of the different probiotic properties could be used to choice the better microorganism depending on the therapeutic or prophylactic application.     

Probing the Drosophila retinal determination gene network in Tribolium (II): The Pax6 genes eyeless and twin of eyeless

The Pax6 genes eyeless (ey) and twin of eyeless (toy) are upstream regulators in the retinal determination gene network (RDGN), which instructs the formation of the adult eye primordium in Drosophila. Most animals possess a singleton Pax6 ortholog, but the dependence of eye development on Pax6 is widely conserved. A rare exception is given by the larval eyes of Drosophila, which develop independently of ey and toy. To obtain insight into the origin of differential larval and adult eye regulation, we studied the function of toy and ey in the red flour beetle Tribolium castaneum. We find that single and combinatorial knockdown of toy and ey affect larval eye development strongly but adult eye development only mildly in this primitive hemimetabolous species. Compound eye-loss, however, was provoked when ey and toy were RNAi-silenced in combination with the early retinal gene dachshund (dac). We propose that these data reflect a role of Pax6 during regional specification in the developing head and that the subsequent maintenance and growth of the adult eye primordium is regulated partly by redundant and partly by specific functions of toy, ey and dac in Tribolium. The results from embryonic knockdown and comparative protein sequence analysis lead us further to conclude that Tribolium represents an ancestral state of redundant control by ey and toy.     

Nomenclatural changes resulting from the transfer of tropical African Sarcostemma to Cynanchum ( Apocynaceae : Asclepiadoideae )

Summary  Four species of tropical African Sarcostemma are transferred to Cynanchum together with two subspecies of S. viminale. In addition, Sarcostemma mulanjense is reduced to subspecific rank under C. viminale.