罗汉果全基因组Survey分析

罗汉果是广西特有药用及甜料植物,其主要成分之一甜苷V作为天然、非糖甜味剂,具有广阔的开发前景,但罗汉果目前完全来自于栽培,适生区狭窄,连作障碍严重,加之含量低导致甜苷V生产成本居高不下,严重限制了其应用。为了减少盲目性,在大规模全基因组深度测序之前,先做低覆盖度的基因组Survey测序,评价基因组的大小及复杂程度,以确定适合该植物全基因组的测序研究策略。该研究采用第二代高通量测序技术(Illumina Hiseq TM 2000)首次测定了罗汉果基因组大小,并利用生物信息学方法估计罗汉果杂合率、重复序列和GC含量等基因组信息。结果表明:(1)获得了18.1 Gb罗汉果基因组测序数据,基因组大小估计为344.95 Mb左右,测序深度为52×;(2)从K-mer分布曲线发现罗汉果基因组有明显的杂合峰,杂合率达1.5%,基因组高杂合导致组装的结果中Contig N50和Scaffold N50的长度比预期的要短很多,还造成GC平均深度及含量分布明显异常,存在一个低深度分布区域。基因组主峰后面有微弱的重复峰,说明罗汉果存在较多的重复序列;(3)由于罗汉果存在高杂合率和重复序列较多的特点,该基因组测序分析仅采用全基因组鸟枪法(WGS)策略不合适,为了更好地对全基因组进行序列拼接和组装,可尝试结合采用Fosmid-to-Fosmid或BAC-to-BAC策略。该研究结果对于揭示罗汉果产量、有效成分含量、发育及抗病虫的分子机制,以及通过分子育种来提高甜苷V含量和降低生产成本具有重要意义,为全基因组测序策略的选择提供了依据。     

樟树全基因组调查

樟树是我国特有的珍贵用材和经济树种,富含多糖多酚、萜类等次生代谢物质,是香精香料、油脂化工和医药等的重要原料树种。本研究采用高通量测序技术(Illumina HiseqTM2000)首次测定了樟树基因组大小,并利用生物信息方法估计樟树杂合率、重复序列情况和GC含量等基因组信息,为全基因组测序策略的选择提供依据。主要结论如下:(1)樟树基因组大小粗略估计为760 Mb左右;(2)樟树基因组有较高的杂合率和一定的重复,杂合率约为0.65%;(3)由于樟树杂合率较高,全基因组鸟枪法策略不适合该基因组测序分析,可尝试使用BAC-to-BAC策略或fosmid策略,有利于樟树基因组的序列拼接和组装。     

基于线粒体CO Ⅰ和Cyt b基因的粉蝶亚科及黄粉蝶亚科(粉蝶科)部分类群的分子系统发生

测定了粉蝶科的粉蝶亚科和黄粉蝶亚科14属共24种线粒体COⅠ和Cyt b基因部分序列,并从GenBank中下载了2种粉蝶的同源序列,以眼蝶科的2个物种为外类群,运用NJ法、贝叶斯法分别重建了分子系统树,探讨了它们的系统发生关系。基因序列分析结果表明,经比对和处理后的序列总长度为1111bp,其中变异位点478个,简约位点382个,碱基T、C、A、G平均含量为39.9%、16.9%、30.9%、12.3%,A＋T含量和C＋G含量分别为70.8%和29.2%。分子系统树显示：黄粉蝶亚科不是单系群,但其中迁粉蝶属和豆粉蝶属在不同的分析方法中均聚合在一起。粉蝶亚科形成一个独立的支系,其中,襟粉蝶族为并系群;粉蝶族的粉蝶属、飞龙粉蝶属和云粉蝶属具有较近的亲缘关系。     

基于Survey分析的濒危植物四合木基因组研究

评价濒危植物四合木（Tetraena mongolica）基因组的大小及复杂程度,开展基因组研究可揭示四合木的超旱生机制,进一步挖掘其特色基因资源。为更好破解四合木的全基因组信息,采用第二代高通量测序技术的基因组Survey分析技术开展四合木基因组大小估测研究,并利用生物信息学方法估计了四合木杂合率、重复序列和GC含量等基因组信息。结果表明：四合木基因组大小为1 079.25 Mb,修正后的基因组大小为1 065.84Mb,杂合率为0.76%,重复序列比例为75.25%,GC含量为33.57%。在经过四合木基因组初步组装后,获得3502 126条contigs,总计682 Mb,其N50为187 bp,推测四合木基因组属于同源四倍体复杂基因组,全基因组测序组装难度较大。由于四合木的高杂合率,后续可采用第三代高通量测序技术（单分子测序）同时结合染色质区域捕获技术,有望最终获得高质量的四合木全基因图谱。     

中国龙船花基因组Survey分析

中国龙船花Ixora chinensis是一种兼备药用与观赏价值的传统中药材,解析中国龙船花的基因组特征信息,可为其全基因组测序和药效成分生物合成的分子机制研究奠定理论基础。本研究利用Survey基因组测序技术,使用K-mer分析方法对中国龙船花基因组特征信息展开评估,获得基因组重复度、杂合度以及GC含量等信息。Survey高通量测序获得Raw Data 47.63 Gb,过滤后Clean data 45.62 Gb。K-mer分析表明,基因组大小588.35 Mb,杂合度1.5598%,重复度64.49%,GC含量35.71%,中国龙船花基因组呈现高杂合度、高重复度、基因组庞大的特征。     

中国灰姑娘绢粉蝶一新亚种(鳞翅目,粉蝶科)

记述中国灰姑娘绢粉蝶Aporia potanini Alphéraky 1新亚种,灰姑娘绢粉蝶白色亚种Aporia potanini huanghaoi ssp.nov..模式标本分别保存于北京自然博物馆昆虫标本室、中国科学院动物研究所昆虫标本馆和中国农业大学昆虫标本室.     

中国迁粉蝶属(鳞翅目,粉蝶科)基于NDⅠ和COⅠ基因部分序列的系统发育关系

测定了中国迁粉蝶属3种4亚种以及迁粉蝶指名亚种5个生态型的线粒体COⅠ基因和NDⅠ基因部分序列,以灵奇尖粉蝶为外群,分别采用最大简约法、最大似然法和贝叶斯法构建分子系统树,探讨了中国迁粉蝶属3种4亚种间的系统发育关系,并首次明确了迁粉蝶指名亚种5个生态型的亲缘关系。结果显示在测得的COⅠ基因661bp序列和NDⅠ基因472bp序列中,有变异位点227个,简约位点119个,A+T含量(75.5%)明显偏高。迁粉蝶属4亚种分子系统关系为:(镉黄迁粉蝶指名亚种+((梨花迁粉蝶指名亚种+梨花迁粉蝶海南亚种)+迁粉蝶指名亚种)),迁粉蝶指名亚种5个生态型分子系统关系为:(无纹型+((有纹型+红角型)+(银斑型+血斑型)))。     

黄唇鱼(Bahaba flavolabiata)的全基因组测序和基因组特征研究

黄唇鱼(Bahaba flavolabiata)为国家二级重点保护野生动物、IUCN(世界自然保护联盟)红色名录的极度濒危物种(CR)。基于其样本数量极其有限,全基因组研究可以提供大量与重要性状相关的功能基因和分子标记,从而揭示其重要生命现象的遗传机制。采用二代测序技术于2018年5月完成了黄唇鱼基因组精细图的测序,分析结果表明,测序得到约202 Gb的高质量数据,总测序深度约为317×;组装得到的基因组大小为637.43 Mb,Contig N50约为88 Kb,Scaffold N50约为4.65 Mb;重复序列约142.72 Mb,占比22.39%,预测得到23743个基因、920个t RNA、85个rRNA、176个假基因;98.46%的基因可以注释到NR、GO等数据库中;有67个基因家族是黄唇鱼所特有的。本研究从单碱基错误率、核心基因完整性及二代Reads比对分析3个方面对黄唇鱼基因组精细图的组装结果进行了评估,结果显示所组装的基因区的完整性较好。黄唇鱼基因组序列图谱的绘制完成,对于黄唇鱼自然资源的保护和种质资源挖掘具有极其重要的科学意义。     

黄毛纺蚋线粒体基因组全序列测定与分析

利用PCR步移法对黄毛纺蚋的线粒体基因组全序列进行了测定和分析。黄毛纺蚋线粒体基因组全长15904 bp(Gen Bank序列号KP793690),包括13个蛋白编码基因、22个tRNA基因、2个rRNA基因以及长度为939 bp的非编码区。A、T、C、G碱基含量分别为39.1%、35.8%、10.4%、14.7%。9个蛋白编码基因和14个tRNA基因在J链编码,其余4个蛋白编码基因和8个tRNA基因在N链编码,基因排列顺序与其它已知双翅目昆虫相同。13个蛋白编码基因中除COI以TTG作为起始密码外,其余蛋白质基因均以ATN作为起始密码子,终止密码子多数为典型的TAA、TAG,只有COI和ND4L以单独的T作为终止密码子。在所测得的22个tRNA基因中,除tRNASer(AGN)缺少DHU臂外,其余tRNA均能形成典型的三叶草结构。     

Sexual Isolation Between Two Sympatric Types of the Butterfly Eurema hecabe (L.)

In Japan, the butterfly Eurema hecabe (L.) consists of two separate types. These types differ in wing fringe color (brown or yellow), which is linked with expression of seasonal wing morphs and with ability for host-plant use. It was recently found that these two types occur together on the subtropical Okinawa-jima Island (one of the Ryukyu Islands). In the present work we studied whether sexual isolation may exist between these sympatric types of E. hecabe on Okinawa-jima, using caged conditions. When living females (days 0, 1, and 7) were presented to males of the same or different type, females of the brown type received males of the same type and responded with a refusal posture to those of the yellow type. Females of the yellow type received males of the same type at all ages tested. However, when they were exposed to males of the brown type, the proportion of females showing a refusal posture changed with female age. The proportion was high at day 7 but relatively low at day 0 or day 1. When model females of the brown and yellow types were simultaneously presented to males of each type, males courted not only the model females of the same type, but also those of the different type. The results suggest that (1) sexual isolation exists between these types of E. hecabe on Okinawa-jima, although it is not perfect; (2) this isolation is mediated by female recognition but not male recognition; and (3) the sexual isolation system between two types is asymmetrical, depending on the female age after adult eclosion.     

Naturally occurring single and double infection with Wolbachia strains in the butterfly Eurema hecabe: transmission efficiencies and population density dynamics of each Wolbachia strain

Wolbachia belonging to Alphaproteobacteria are transovarially transmitted bacteria responsible for reproductive alterations in a wide range of arthropods. In natural populations of the butterfly Eurema hecabe, there are two different types of Wolbachia-infected individuals. Individuals singly infected with Wolbachia strain wHecCI exhibit strong cytoplasmic incompatibility, whereas those doubly infected with wHecCI and wHecFem exhibit feminization. Here, we examined the infection frequencies and population densities of each Wolbachia strain in different host tissues (ovary, testis, fat body, midgut, Malpighian tubule and leg), and the cost of infection in offspring produced by single-infected and double-infected mothers of E. hecabe. The vertical transmission rate of wHecCI was nearly 100%, and that of wHecFem was c. 80%. The wHecCI densities were 10(3)-10(4)-fold higher than the wHecFem densities. In most tissues, the wHecCI densities were significantly higher in offspring of single-infected mothers than in offspring of double-infected mothers. In offspring of double-infected mothers, however, the wHecCI densities were not affected by the presence of wHecFem, suggesting a lack of interaction between the wHecCI and wHecFem densities. The offspring development time was dependent on the infection status of the mothers. These results imply that the maternal infection status affects the Wolbachia densities and fitness of offspring.     

Courtship Solicitation by Females of the Barred Sulphur Butterfly (<Emphasis Type="Italic">Eurema daira</Emphasis>) (Lepidoptera: Pieridae)

Virgin females of the subtropical pierid butterfly Eurema daira were observed to actively solicit male courtship. The resulting interactions involved both aggressive contact with male individuals and elaborate female posturing. Cage-based trials showed that there was a direct relationship between the frequency of courtship solicitation attempts and female age. Virgin females under 3 days old showed little or no interest in initiating courtship. By contrast, 23.2 percent of the virgin females over three days old actively solicited available males with peak solicitation behavior exhibited from five day old individuals.     

Female mating biases for bright ultraviolet iridescence in the butterfly Eurema hecabe (Pieridae)

Exaggerated male-limited coloration is widespread among butterflies,yet convincing demonstrations of intraspecific mating preferencesfor signal brightness and/or chromaticity are relatively rarein this group. Here, I couple behavioral experiments involvingmanipulations of ambient light environments and male reflectancepatterns with observation of wild mating patterns to investigatevisual mating biases in the large grass yellow (Eurema hecabe).Males in this species possess exaggerated, limited-view ultraviolet(UV) iridescence across most of their dorsal wing surface thathas putative sexual signaling function. In the first experiment,conducted in small (0.7–m³) cages, individuals were significantlyless likely to copulate when the UV portion of natural ambientillumination (i.e., 300–400 nm) was strongly reduced.In 2 subsequent experiments, conducted under full-spectrum sunlightin small and large (5 x 6 x 4 m) cages, males with their UVsignal artificially dulled by 25% consistently copulated withfewer, and smaller, females than sham-control individuals. Importantly,the manipulated levels of UV brightness in these experimentsfall well within the naturally occurring bounds of variationin male UV reflectance. These findings therefore unanimouslysupport the presence of a UV signal–based female bias.In apparent contrast, comparison of 161 in-copula and 188 free-flyingmales from a high-density field assemblage revealed that copulatingmales were significantly older and henceforth actually possessed(subtly) less UV bright wings. Copulating male UV brightnesswas, however, positively related to the size of their mate,which echoes the experimental findings and may represent a signatureof mutual mate choice. I discuss these results in light of thefull complexities of the butterfly mating system and the potentialsignaling value of iridescent coloration in butterflies andanimals generally.     

Biological Implications of Isochore Boundaries in the Human Genome

Abstract

The human genome is composed of large sequence segments with fairly homogeneous GC content, namely isochores, which have been linked to many important functions; biological implications of most isochore boundaries, however, remain elusive, partly due to the difficulty in determining these boundaries at high resolution. Using the segmentation algorithm based on the quadratic divergence, we re-determined all 79 boundaries of previously identified human isochores at single-nucleotide resolution, and then compared the boundary coordinates with other genome features. We found that 55.7% of isochore boundaries coincide with termini of repeat elements; 45.6% of isochore boundaries coincide with termini of highly conserved sequences based on alignment of 17 vertebrate genomes, i.e., the highly conserved genome sequence switches to a less or non-conserved one at the isochore boundary; some isochore boundaries coincide with abrupt change of CpG island distribution (note that one boundary can associate with more than one genome feature). In addition, sequences around isochore boundaries are highly conserved. It seems reasonable to deduce that the boundaries of all the isochores studied here would be replication timing sites in the human genome. These results suggest possible key roles of the isochore boundaries and may further our understanding of the human genome organization.     

The Decline of Isochores in Mammals: An Assessment of the GC ContentVariation Along the Mammalian Phylogeny

Whether isochores, the large-scale variation of the GC content in mammalian genomes, are being maintained has recently been questioned. It has been suggested that GC-rich isochores originated in the ancestral amniote genome but that whatever force gave rise to them is no longer effective and that isochores are now disappearing from mammalian genomes. Here we investigated the evolution of the GC content of 41 coding genes in 6 to 66 species of mammals by estimating the ancestral GC content using a method which allows for different rates of substitution between sites. We found a highly significant decrease in the GC content during early mammalian evolution, as well as a weaker but still significant decrease in the GC content of GC-rich genes later in at least three groups of mammals: primates, rodents, and carnivores. These results are of interest because they confirm the recently suggested disappearance of GC-rich isochores in some mammalian genomes, and more importantly, they suggest that this disappearance started very early in mammalian evolution.This article contains online supplementary material.     

Genome evolution in the genus Sorghum (Poaceae)

BACKGROUND AND AIMS: The roles of variation in DNA content in plant evolution and adaptation remain a major biological enigma. Chromosome number and 2C DNA content were determined for 21 of the 25 species of the genus Sorghum and analysed from a phylogenetic perspective. METHODS: DNA content was determined by flow cytometry. A Sorghum phylogeny was constructed based on combined nuclear ITS and chloroplast ndhF DNA sequences. KEY RESULTS: Chromosome counts (2n = 10, 20, 30, 40) were, with few exceptions, concordant with published numbers. New chromosome numbers were obtained for S. amplum (2n = 30) and S. leiocladum (2n = 10). 2C DNA content varies 8.1-fold (1.27-10.30 pg) among the 21 Sorghum species. 2C DNA content varies 3.6-fold from 1.27 pg to 4.60 pg among the 2n = 10 species and 5.8-fold (1.52-8.79 pg) among the 2n = 20 species. The x = 5 genome size varies over an 8.8-fold range from 0.26 pg to 2.30 pg. The mean 2C DNA content of perennial species (6.20 pg) is significantly greater than the mean (2.92 pg) of the annuals. Among the 21 species studied, the mean x = 5 genome size of annuals (1.15 pg) and of perennials (1.29 pg) is not significantly different. Statistical analysis of Australian species showed: (a) mean 2C DNA content of annual (2.89 pg) and perennial (7.73 pg) species is significantly different; (b) mean x = 5 genome size of perennials (1.66 pg) is significantly greater than that of the annuals (1.09 pg); (c) the mean maximum latitude at which perennial species grow (-25.4 degrees) is significantly greater than the mean maximum latitude (-17.6) at which annual species grow. CONCLUSIONS: The DNA sequence phylogeny splits Sorghum into two lineages, one comprising the 2n = 10 species with large genomes and their polyploid relatives, and the other with the 2n = 20, 40 species with relatively small genomes. An apparent phylogenetic reduction in genome size has occurred in the 2n = 10 lineage. Genome size evolution in the genus Sorghum apparently did not involve a 'one way ticket to genomic obesity' as has been proposed for the grasses.     

Analysis of nucleotide distribution in the genome of Streptomyces coelicolor A3(2) using the Z curve method

The nucleotide distribution of all 33 527 open reading frames (ORFs) (≥300 bp) in the genome of Streptomyces coelicolor A3(2) has been analyzed using the Z curve method. Each ORF is mapped onto a point in a 9-dimensional space. To visualize the distribution of mapping points, the points are projected onto the principal plane based on principal component analysis. Consequently, the distribution pattern of the 33 527 points in the principal plane shows a flower-like shape, in which there are seven distinct regions. In addition to the central region, there are six petal-like regions around the center, one of which corresponds to 7172 coding sequences. The central region and the remaining five petal-like regions correspond to the intergenic sequences and out-of-frame non-coding ORFs, respectively. It is shown that selective pressure produces a remarkable bias of the G+C content among three codon positions, resulting in the interesting phenomenon observed. A similar phenomenon is also observed for other bacterial genomes with high genomic G+C content, such as Pseudomonas aeruginosa PA01 (G+C=66.6%). However, for the genomes of Bacillus subtilis (G+C=43.5%) and Clostridium perfringens (G+C=28.6%), no similar phenomenon was observed. The finding presented here may be useful to improve the gene-finding algorithms for genomes with high G+C content. A set of supplementary materials including the plots displaying the base distribution patterns of ORFs in 12 prokaryotes is provided on the website http://tubic.tju.edu.cn/highGC/.     

甘薯属耐盐植物马鞍藤基因组大小及特征分析

马鞍藤(Ipomoea pes-caprae(L.)R. Br.)是甘薯的近缘野生种之一,为热带、亚热带海滩生长的多年生藤蔓植物,作为沿海地区的园林绿化植物之一,具有较强的耐盐性。马鞍藤基因组学的研究可为耐盐甘薯种质创制提供信息参考。本试验调查了马鞍藤基因组大小和特征概况,为后续全基因组精细图谱绘制打下基础。研究以已知基因组大小的三裂叶薯(Ipomoea triloba L.)为对照,运用流式细胞术对马鞍藤基因组大小进行初步预估;使用二代高通量测序技术(Illumina Hiseq2500)对马鞍藤基因组进行survey评估,测序深度30×,利用生物信息学方法估算马鞍藤GC含量(即鸟嘌呤和胞嘧啶所占比例)、杂合率、重复序列等基因组概况。结果表明:流式细胞术估算马鞍藤基因组大小为1012.704±17.37 Mb;经全基因组Survey测定获得马鞍藤有效数据为21.71 Gb,基因组大小经修正后估算为1041.65 Mbp;通过K-mer分布曲线估算马鞍藤基因组中重复序列所占比率为74.52%,杂合率为0.99%;经初步组装后,GC平均深度及含量分布存在异常,出现分层的现象,这可能与马鞍藤基因组杂合率较高有关。本试验首次报导甘薯属耐盐植物马鞍藤基因组大小及特征信息,为马鞍藤进一步的全基因组深度测序和甘薯耐盐基因挖掘打下基础。