Identification of a coordinate regulator of interleukins 4, 13, and 5 by cross-species sequence comparisons

Long-range regulatory elements are difficult to discover experimentally; however, they tend to be conserved among mammals, suggesting that cross-species sequence comparisons should identify them. To search for regulatory sequences, we examined about 1 megabase of orthologous human and mouse sequences for conserved noncoding elements with greater than or equal to 70% identity over at least 100 base pairs. Ninety noncoding sequences meeting these criteria were discovered, and the analysis of 15 of these elements found that about 70% were conserved across mammals. Characterization of the largest element in yeast artificial chromosome transgenic mice revealed it to be a coordinate regulator of three genes, interleukin-4, interleukin-13, and interleukin-5, spread over 120 kilobases.     

Evolutionary discrimination of mammalian conserved non-genic sequences (CNGs)

Analysis of the human and mouse genomes identified an abundance of conserved non-genic sequences (CNGs). The significance and evolutionary depth of their conservation remain unanswered. We have quantified levels and patterns of conservation of 191 CNGs of human chromosome 21 in 14 mammalian species. We found that CNGs are significantly more conserved than protein-coding genes and noncoding RNAS (ncRNAs) within the mammalian class from primates to monotremes to marsupials. The pattern of substitutions in CNGs differed from that seen in protein-coding and ncRNA genes and resembled that of protein-binding regions. About 0.3% to 1% of the human genome corresponds to a previously unknown class of extremely constrained CNGs shared among mammals.     

A comparison of whole-genome shotgun-derived mouse chromosome 16 and the human genome

The high degree of similarity between the mouse and human genomes is demonstrated through analysis of the sequence of mouse chromosome 16 (Mmu 16), which was obtained as part of a whole-genome shotgun assembly of the mouse genome. The mouse genome is about 10% smaller than the human genome, owing to a lower repetitive DNA content. Comparison of the structure and protein-coding potential of Mmu 16 with that of the homologous segments of the human genome identifies regions of conserved synteny with human chromosomes (Hsa) 3, 8, 12, 16, 21, and 22. Gene content and order are highly conserved between Mmu 16 and the syntenic blocks of the human genome. Of the 731 predicted genes on Mmu 16, 509 align with orthologs on the corresponding portions of the human genome, 44 are likely paralogous to these genes, and 164 genes have homologs elsewhere in the human genome; there are 14 genes for which we could find no human counterpart.     

哺乳动物BMP4基因A型启动子的比较分析

以鸭嘴兽BMP4基因组序列为模型,采用比较基因组学方法分析哺乳动物BMP4基因A型启动子的结构特点。该研究分离出一段长112bp的进化上高度保守的核心启动子序列以及多个转录因子结合位点和CpG岛结构,并且在BMP4核心启动子区域发现了2种具有重要生物学功能的回文结构,为进一步研究该型启动子的转录调控功能提供了理论基础。同时以鸭嘴兽基因组为参照模型,也为今后研究其他哺乳动物进化上保留下来的基因提供更多新信息。     

Evolutionarily assembled cis-regulatory module at a human ciliopathy locus

Neighboring genes are often coordinately expressed within cis-regulatory modules, but evidence that nonparalogous genes share functions in mammals is lacking. Here, we report that mutation of either TMEM138 or TMEM216 causes a phenotypically indistinguishable human ciliopathy, Joubert syndrome. Despite a lack of sequence homology, the genes are aligned in a head-to-tail configuration and joined by chromosomal rearrangement at the amphibian-to-reptile evolutionary transition. Expression of the two genes is mediated by a conserved regulatory element in the noncoding intergenic region. Coordinated expression is important for their interdependent cellular role in vesicular transport to primary cilia. Hence, during vertebrate evolution of genes involved in ciliogenesis, nonparalogous genes were arranged to a functional gene cluster with shared regulatory elements.     

Human genome ultraconserved elements are ultraselected

Ultraconserved elements in the human genome are defined as stretches of at least 200 base pairs of DNA that match identically with corresponding regions in the mouse and rat genomes. Most ultraconserved elements are noncoding and have been evolutionarily conserved since mammal and bird ancestors diverged over 300 million years ago. The reason for this extreme conservation remains a mystery. It has been speculated that they are mutational cold spots or regions where every site is under weak but still detectable negative selection. However, analysis of the derived allele frequency spectrum shows that these regions are in fact under negative selection that is much stronger than that in protein coding genes.     

两个大白菜FT基因的生物信息学解析

本文利用比较基因组学的方法从大白菜的基因组中解析出两个成花素基因brft1和brft2,并对这两个基因的结构、顺式调控元件以及遗传进化进行了分析。结果表明,brft1和brft2都含有4个外显子,编码区大小均为528 bp,二者预测编码蛋白的序列一致性为90.29%, 说明这两个基因序列存在某种程度的分化。进化分析表明这两个基因应为拟南芥ft和tsf基因的直系同源基因。另外,这两个基因都含有响应激素和逆境信号的顺式调控元件,而且不尽相同,说明这两个基因的表达调控有所不同,可能在功能上存在一定分化。本研究为进一步揭示大白菜ft基因在开花和逆境响应中的功能奠定了基础。     

The Release 5.1 annotation of Drosophila melanogaster heterochromatin

The repetitive DNA that constitutes most of the heterochromatic regions of metazoan genomes has hindered the comprehensive analysis of gene content and other functions. We have generated a detailed computational and manual annotation of 24 megabases of heterochromatic sequence in the Release 5 Drosophila melanogaster genome sequence. The heterochromatin contains a minimum of 230 to 254 protein-coding genes, which are conserved in other Drosophilids and more diverged species, as well as 32 pseudogenes and 13 noncoding RNAs. Improved methods revealed that more than 77% of this heterochromatin sequence, including introns and intergenic regions, is composed of fragmented and nested transposable elements and other repeated DNAs. Drosophila heterochromatin contains "islands" of highly conserved genes embedded in these "oceans" of complex repeats, which may require special expression and splicing mechanisms.     

Comparative genome and proteome analysis of Anopheles gambiae and Drosophila melanogaster

Comparison of the genomes and proteomes of the two diptera Anopheles gambiae and Drosophila melanogaster, which diverged about 250 million years ago, reveals considerable similarities. However, numerous differences are also observed; some of these must reflect the selection and subsequent adaptation associated with different ecologies and life strategies. Almost half of the genes in both genomes are interpreted as orthologs and show an average sequence identity of about 56%, which is slightly lower than that observed between the orthologs of the pufferfish and human (diverged about 450 million years ago). This indicates that these two insects diverged considerably faster than vertebrates. Aligned sequences reveal that orthologous genes have retained only half of their intron/exon structure, indicating that intron gains or losses have occurred at a rate of about one per gene per 125 million years. Chromosomal arms exhibit significant remnants of homology between the two species, although only 34% of the genes colocalize in small "microsyntenic" clusters, and major interarm transfers as well as intra-arm shuffling of gene order are detected.     

美国红鱼和大黄鱼β-珠蛋白基因的克隆和遗传进化分析

根据已发表的其他脊椎动物β-珠蛋白基因序列设计并合成1对特异性引物,通过RT-PCR方法首次克隆到美国红鱼和大黄鱼β-珠蛋白cDNA,其开放读码框均由447个碱基组成,编码148个氨基酸的多肽,预测的相对分子量分别为16 333和16 148 Da,两者核苷酸序列和推导的氨基酸序列同源性分别为90.4%与81.2%.将推导的美国红鱼和大黄鱼β-珠蛋白氨基酸序列与其他鱼类及人β-珠蛋白氨基酸序列进行同源性比较,其同源性在35.5%～70.3%,它们与同属真口鱼纲的真鲷、黄尾鲱鱼、鲤鱼、虹鳟等的同源性较高,而与肉鳍鱼纲的肺鱼及板鳃纲的鲨鱼的同源性较低;二级结构预测表明,美国红鱼和大黄鱼β-珠蛋白存在类似哺乳动物的8个α-螺旋区,在近端与远端与血红素结合的组氨酸高度保守.     

Construction and Characterization of a cDNA Library from the Pulp of Coconut （Cocos nucifera L.）

To investigate the gene expression profile of endosperm development, a cDNA library was constructed and characterized from the pulp of coconut at different developmental stages. The constructed cDNA library incorporated approximately 1 × 10^7 clones in total, and the size of the insertion fragments ranged from 800 to 2 000 bp. Sequencing results of 100 randomly picked clones showed that the recombination rate was 96%. In subsequent sequence analysis, 41 clones （41%） were homologous to known function proteins, and 23 clones showed high amino acid identity （more than 80%） with the corresponding genes of different plants. Semi-quantitative RT-PCR indicated that oleosin and globulin genes are pulpspecific expression, and have differential expression level in different developmental stage. Clone 29, recognized as homologous to KIAA1239 protein （Homo sapiens）, was observed to occur nine times, indicating that this gene may be over-expressed during the endosperm development stage. However, the homologous protein was found only in mammals, and the detailed function is still unknown. Elucidation of the functional characterization of these genes will be carried out immediately.     

Accelerated evolution of conserved noncoding sequences in humans

Changes in gene regulation likely influenced the profound phenotypic divergence of humans from other mammals, but the extent of adaptive substitution in human regulatory sequences remains unknown. We identified 992 conserved noncoding sequences (CNSs) with a significant excess of human-specific substitutions. These accelerated elements were disproportionately found near genes involved in neuronal cell adhesion. To assess the uniqueness of human noncoding evolution, we examined CNSs accelerated in chimpanzee and mouse. Although we observed a similar enrichment near neuronal adhesion genes in chimpanzee, the accelerated CNSs themselves exhibited almost no overlap with those in human, suggesting independent evolution toward different neuronal phenotypes in each species. CNSs accelerated in mouse showed no bias toward neuronal cell adhesion. Our results indicate that widespread cis-regulatory changes in human evolution may have contributed to uniquely human features of brain development and function.     

Mobile elements: drivers of genome evolution

Mobile elements within genomes have driven genome evolution in diverse ways. Particularly in plants and mammals, retrotransposons have accumulated to constitute a large fraction of the genome and have shaped both genes and the entire genome. Although the host can often control their numbers, massive expansions of retrotransposons have been tolerated during evolution. Now mobile elements are becoming useful tools for learning more about genome evolution and gene function.     

福美双诱发肉鸡胫骨软骨发育不良早期生长板消减cDNA文库的构建

 【目的】为应用cDNA芯片技术筛选肉鸡胫骨软骨发育不良(TD)时序性差异表达基因,本研究构建了早期生长板消减 cDNA文库。【方法】将7日龄AV肉鸡随机分为两组,对照组(control,C)饲以基础日粮,试验组(thiram diet-fed,T)饲以基础日粮添加福美双100 mg?kg-1,2 d后继续饲以基础日粮,诱发肉鸡TD。应用抑制消减杂交技术(SSH)构建正反向消减cDNA文库,用PCR验证两个文库中克隆的插入片段,随机抽取100个阳性克隆测序,对所测序列同源性分析和功能预测。【结果】从两个文库共获得2 227个有效阳性克隆,插入片段大小主要分布在200—1 000bp之间;所测序列中有97条ESTs与GenBank中的鸡基因序列同源性达到99%,且非冗余度达到68.7%;此外,有3条ESTs未找到同源序列。这些基因具有构成细胞外基质,参与软骨内骨化和骨的重构,调节骨发育,行使信号转导、血管发生,参与电子传递及能量代谢等功能。【结论】成功构建了消减 cDNA文库,将为进一步点制cDNA芯片,筛选不同阶段的差异表达基因奠定了基础,也为肉鸡TD机理研究提供新线索。     

Identification and Bioinformatics Analysis of SnRK2 and CIPK Family Genes in Sorghum

Through bioinformatic data mining, 10 SnRK2 and 31 CIPK genes were identified from sorghum genome. They are unevenly distributed in the sorghum chromosomes. Most SnRK2 genes have 8 introns, while the CIPK genes have a few （no intron or less than 3 introns） or more than I0 introns. Phylogenetic analysis revealed that SnRK2 genes belong to one cluster and CIPK genes form the other independent cluster. The sorghum SnRK2s are subgrouped into three parts, and CIPK into five parts. More than half SnRK2 and CIPK genes present in homologous pairs, suggesting gene duplication may be due to the amplification of SnRK family genes. The kinase domains of SnRK2 family are highly conserved with 88.40% identity, but those of the CIPK family are less conserved with 63.72% identity. And the identity of sorghum CBLinteracting NAF domains of CIPKs is 61.66%. What＇s more, regarding to the sorghum SnRK2 and CIPK kinases, they are characterized with distinct motifs and their subcellular localization is not necessarily the same, which suggests they may be divergent in functions. Due to less conserved sequences, complex subcellular localization, and more family members, sorghum CIPK genes may play more flexible and multiple biological functions. According to the phylogenetic analysis of SnRK genes and SnRK functional studies in other plants, it is speculated that sorghum SnRK2 and CIPK genes may play important roles in stress response, growth and development.     

The dog genome: survey sequencing and comparative analysis

A survey of the dog genome sequence (6.22 million sequence reads; 1.5x coverage) demonstrates the power of sample sequencing for comparative analysis of mammalian genomes and the generation of species-specific resources. More than 650 million base pairs (>25%) of dog sequence align uniquely to the human genome, including fragments of putative orthologs for 18,473 of 24,567 annotated human genes. Mutation rates, conserved synteny, repeat content, and phylogeny can be compared among human, mouse, and dog. A variety of polymorphic elements are identified that will be valuable for mapping the genetic basis of diseases and traits in the dog.     

Whole-genome shotgun assembly and analysis of the genome of Fugu rubripes

The compact genome of Fugu rubripes has been sequenced to over 95% coverage, and more than 80% of the assembly is in multigene-sized scaffolds. In this 365-megabase vertebrate genome, repetitive DNA accounts for less than one-sixth of the sequence, and gene loci occupy about one-third of the genome. As with the human genome, gene loci are not evenly distributed, but are clustered into sparse and dense regions. Some "giant" genes were observed that had average coding sequence sizes but were spread over genomic lengths significantly larger than those of their human orthologs. Although three-quarters of predicted human proteins have a strong match to Fugu, approximately a quarter of the human proteins had highly diverged from or had no pufferfish homologs, highlighting the extent of protein evolution in the 450 million years since teleosts and mammals diverged. Conserved linkages between Fugu and human genes indicate the preservation of chromosomal segments from the common vertebrate ancestor, but with considerable scrambling of gene order.     

猪围脂滴蛋白基因cDNA部分片段的克隆及序列分析

采用比较基因组学和简并PCR方法,根据人、小鼠和大鼠等物种PLIN基因的序列,设计简并引物,克隆了猪PLIN基因包括起始密码子的部分cDNA序列,大小为978 bp(GenBank登录号为EU416277)。该序列与人和小鼠相应序列的相似性分别为87%和83%,编码325个氨基酸,具有1个PAT-1结构域、2个蛋白激酶A位点和1个富含谷氨酸的酸性基序,预测的氨基酸序列与人、小鼠、牛和绵羊相应区域序列的相似性分别为87%,82%,86%,86%。该基因的克隆为进一步研究其功能奠定了基础。     

伪狂犬病毒Fa株gB、gC、gD基因的克隆与序列分析

对猪伪狂犬病病毒闽A株(Fa株)gB、gC、gD基因进行了克隆和序列测定,结果表明:gB、gC、gD基因序列全长分别为2 745 bp、1464 bp、1 215 bp,编码914、487、404个氨基酸残基组成的多肽。序列分析结果显示:Fa株与其他毒株gB、gC、gD基因核苷酸序列的同源性为94.9%~99.9%,氨基酸序列的同源性为91.4%~99.9%。不同PRV毒株gB、gC、gD基因在核苷酸和氨基酸水平上高度保守。基于gB、gC、gD基因的进化树分析表明,中国分离毒株与韩国分离毒株可归为一个进化分支,而日本分离株与欧美分离株归为另一个分支。在前一个分支里,闽A株与鄂A株的亲缘关系特别近,3个基因的同源率均在99.5%以上。在gB蛋白氨基酸序列全长上中国分离株(Ea株、Fa株)比欧美分离株多了1个氨基酸,氨基酸残基的突变主要集中在第50~100氨基酸。在gC基因序列第186~211 bp,Ea株、Fa株比欧美分离株多插入了21个碱基。在gD蛋白氨基酸序列全长上,Ea株、Fa株比其他分离株多了2~6个氨基酸。在gD基因序列第803~837 bp,Fa株核苷酸AGGCCC串联重复最多,达到7个。