转座子在基因组和基因进化方面的研究进展

从转座子插入对基因组产生的重组、异位、倒位以及对基因产生的沉默、表达等方面阐述了转座子对基因组和基因进化的重要作用,从转座子在基因中插入位置的不同对基因表达和功能产生的影响进行了综述。     

piggyBac转座子在绒山羊基因组中的整合位点及其特征分析

【目的】构建piggyBac（PB）转座子表达载体系统,研究PB转座子在绒山羊基因组中的整合位点。【方法】构建pB-CMV-EGFP-Neo转座载体和pcDNA-Trans辅助载体,利用lipofectamine 2000介导共转染绒山羊胎儿成纤维细胞,经G418筛选获得稳定转染的转基因细胞系。提取转基因细胞基因组DNA,利用反向PCR检测piggyBac转座子整合位点。【结果】构建了转座系统并成功介导外源基因在绒山羊成纤维细胞染色体中整合,获得了转基因细胞系;反向PCR检测显示在绒山羊基因组中有21个PB转座子整合位点;整合位点TTAA毗邻一侧的5个碱基组成中,PB转座子3′倾向于插入到富含AT（58.35%）碱基的区域,PB转座子5′倾向于插入到富含GC（57.8%）。【结论】PB转座子可在绒山羊基因组中发生高效转座,获得的整合位点信息可为利用PB转座子开展绒山羊研究提供理论参考。     

Genetics. Transposons help sculpt a dynamic genome

New research is showing that the mobile genetic elements called transposons cause more extensive restructuring of the genome than previously thought. Researchers have known for about 20 years that transposons can expand the genome, resulting in the repetitive DNA sequences sometimes called "junk," but the new work indicates that transposons can also contribute to substantial DNA losses. What's more, these changes can be rapid--at least on an evolutionary scale--and may help organisms adapt to their environments.     

The genome sequence of Drosophila melanogaster

The fly Drosophila melanogaster is one of the most intensively studied organisms in biology and serves as a model system for the investigation of many developmental and cellular processes common to higher eukaryotes, including humans. We have determined the nucleotide sequence of nearly all of the approximately 120-megabase euchromatic portion of the Drosophila genome using a whole-genome shotgun sequencing strategy supported by extensive clone-based sequence and a high-quality bacterial artificial chromosome physical map. Efforts are under way to close the remaining gaps; however, the sequence is of sufficient accuracy and contiguity to be declared substantially complete and to support an initial analysis of genome structure and preliminary gene annotation and interpretation. The genome encodes approximately 13,600 genes, somewhat fewer than the smaller Caenorhabditis elegans genome, but with comparable functional diversity.     

Draft genome sequence of the sexually transmitted pathogen Trichomonas vaginalis

We describe the genome sequence of the protist Trichomonas vaginalis, a sexually transmitted human pathogen. Repeats and transposable elements comprise about two-thirds of the approximately 160-megabase genome, reflecting a recent massive expansion of genetic material. This expansion, in conjunction with the shaping of metabolic pathways that likely transpired through lateral gene transfer from bacteria, and amplification of specific gene families implicated in pathogenesis and phagocytosis of host proteins may exemplify adaptations of the parasite during its transition to a urogenital environment. The genome sequence predicts previously unknown functions for the hydrogenosome, which support a common evolutionary origin of this unusual organelle with mitochondria.     

Human immunodeficiency virus may encode a novel protein on the genomic DNA plus strand

The genome of the human immunodeficiency virus (HIV) is known to contain eight open reading frames (ORFs) on the minus strand of the double-stranded DNA replicative intermediate. Data presented here indicate that the DNA plus strand of HIV contains a previously unidentified ORF in a region complementary to the envelope gene sequence. This ORF could encode a protein of approximately 190 amino acid residues with a relative molecular mass of 20 kilodaltons if translation began from the first initiation codon. The predicted protein is highly hydrophobic and thus could be membrane associated. It is possible, therefore, that the HIV genome encodes a protein on antisense messenger RNA.     

A virophage at the origin of large DNA transposons

DNA transposons are mobile genetic elements that have shaped the genomes of eukaryotes for millions of years, yet their origins remain obscure. We discovered a virophage that, on the basis of genetic homology, likely represents an evolutionary link between double-stranded DNA viruses and Maverick/Polinton eukaryotic DNA transposons. The Mavirus virophage parasitizes the giant Cafeteria roenbergensis virus and encodes 20 predicted proteins, including a retroviral integrase and a protein-primed DNA polymerase B. On the basis of our data, we conclude that Maverick/Polinton transposons may have originated from ancient relatives of Mavirus, and thereby influenced the evolution of eukaryotic genomes, although we cannot rule out alternative evolutionary scenarios.     

PiggyBac转座元件的构建及其在团头鲂基因组中的转基因效率

鳞翅目昆虫粉纹夜蛾(Trichoplusia ni)的PiggyBac(PB)转座子已用于模式生物小鼠的转基因及插入诱变研究,目前,该转座子在养殖鱼类中的转基因效率如何还不清楚。构建了带PiggyBac转座子左臂、右臂、EF1α启动子和绿色荧光蛋白(eGFP)编码框的pPBs-EF1α-eGFP供体质粒。以50 ng/μL供体质粒和100 ng/μL体外转录的PB转座酶mRNA共同显微注射入团头鲂(Megalobrama amblycephala)1~2细胞期受精卵中,团头鲂eGFP的荧光表达率可达58.26%,PCR检测结果显示,该转座系统在团头鲂成鱼基因组中的整合效率为53.04%。表明PiggyBac转座子可高效介导基因在团头鲂基因组中的插入,为进一步开展团头鲂插入诱变研究奠定了基础。     

枯草芽胞杆菌Bs-916的全基因组分析

 【目的】对枯草芽胞杆菌Bs-916进行全基因组测序,为今后更好挖掘和利用该菌生防潜力提供较多的分子生物学信息。【方法】通过应用比较基因组学软件与另一测序菌株Bs168进行全基因组序列分析。【结果】Bs-916菌株全基因组大小为3 925 958 bp,GC含量为46.4%,与其它已测序全基因组芽孢杆菌相比,其GC含量最高;预测所得CDS数为4 056个,152个串联重复区域,转座子103个,IS（插入序列）序列数量37个, tRNA 46个,rRNA 39个;比较基因组学分析其含有8个NRPS/PKS基因簇,其中bacillomycin L,macrolactin,difficidin这3个基因簇在比较菌株Bs168中不存在;该菌还含有植酸酶基因、comAPQX及sfp等与生防因子相关的基因。【结论】Bs-916菌株全基因组含有多种抗菌物质编码基因簇,是一类具有极大生防潜力的典型根际革兰氏阳性菌株。该菌株全基因组序列的测定及其遗传操作方法的可行性,有利于其次生代谢产物的开发和利用。次生代谢产物具有潜在的应用价值,有望在未来开发成独特的农业生物工程制剂。     

Genome streamlining in a cosmopolitan oceanic bacterium

The SAR11 clade consists of very small, heterotrophic marine alpha-proteobacteria that are found throughout the oceans, where they account for about 25% of all microbial cells. Pelagibacter ubique, the first cultured member of this clade, has the smallest genome and encodes the smallest number of predicted open reading frames known for a free-living microorganism. In contrast to parasitic bacteria and archaea with small genomes, P. ubique has complete biosynthetic pathways for all 20 amino acids and all but a few cofactors. P. ubique has no pseudogenes, introns, transposons, extrachromosomal elements, or inteins; few paralogs; and the shortest intergenic spacers yet observed for any cell.     

花叶矢竹转录组中的转座子表达分析

以花叶矢竹Pseudosasa japonica f.akebonosuji 10种不同颜色和不同发育阶段的叶片转录组数据为基础,调查了花叶矢竹中转座子的种类、数量以及选择性表达特性。结果表明:花叶矢竹转录组中转座子类型丰富、数量繁多,其中RNA转座子明显多于DNA转座子,转座子LTR/Copia类型数量最多。绿叶的5个发育阶段中,转座子主要在第5发育阶段高表达,表明这些转座子可能参与了花叶矢竹叶片成熟过程。而在白叶5个发育阶段中,转座子主要在第1发育阶段高表达;对绿叶和白叶5个相对应的发育阶段分析表明,白叶中高表达的转座子多于绿叶,表明这些转座子可能参与了花叶矢竹叶色变异过程,也可能是逆境胁迫导致转座子转座激活。图3表3参25     

Common sequence polymorphisms shaping genetic diversity in Arabidopsis thaliana

The genomes of individuals from the same species vary in sequence as a result of different evolutionary processes. To examine the patterns of, and the forces shaping, sequence variation in Arabidopsis thaliana, we performed high-density array resequencing of 20 diverse strains (accessions). More than 1 million nonredundant single-nucleotide polymorphisms (SNPs) were identified at moderate false discovery rates (FDRs), and approximately 4% of the genome was identified as being highly dissimilar or deleted relative to the reference genome sequence. Patterns of polymorphism are highly nonrandom among gene families, with genes mediating interaction with the biotic environment having exceptional polymorphism levels. At the chromosomal scale, regional variation in polymorphism was readily apparent. A scan for recent selective sweeps revealed several candidate regions, including a notable example in which almost all variation was removed in a 500-kilobase window. Analyzing the polymorphisms we describe in larger sets of accessions will enable a detailed understanding of forces shaping population-wide sequence variation in A. thaliana.     

亚东鲑基因组中Tc1-like转座子的序列歧化特征分析

根据鱼类Tc1-like超家族转座子的末端反向重复序列设计单引物,对西藏亚东鲑基因组进行PCR扩增、回收、克隆和测序,鉴定出亚东鲑两条长度为1 607 bp和1 473 bp的Tc1-like超家族转座子序列,命名为Tbt1和Tbt2。序列分析表明,亚东鲑Tbt1转座子左右两端分别存在一个196 nt和225 nt的末端反向重复序列(Inverted terminal repeats,ITR),在左右ITR中分别包含2个12 nt的亚末端反向重复序列(Subterminal inverted repeats,SIR);亚东鲑Tbt2转座子分别存在一个32 nt和31 nt短的ITR,其左右ITR中各仅包含1个12 nt的SIR。亚东鲑Tbt1、Tbt2转座子的转座酶编码区在进化过程中各已积累了4个和9个终止突变,两者均不能表达完整的转座酶。亚东鲑Tbt2与其它鲑科鱼类Tc1-like转座子的相似度低于30%,而与金鱼Tca2转座子的序列相似度高达98%,显示该转座子的获得可能起源于基因水平转移(horizontal gene transfer,HGT)方式。     

Very low gene duplication rate in the yeast genome

The gene duplication rate in the yeast genome is estimated without assuming the molecular clock model to be approximately 0.01 to 0.06 per gene per billion years; this rate is two orders of magnitude lower than a previous estimate based on the molecular clock model. This difference is explained by extensive concerted evolution via gene conversion between duplicated genes, which violates the assumption of the molecular clock in the analyses of duplicated genes. The average length of the period of concerted evolution and the gene conversion rate are estimated to be approximately 25 million years and approximately 28 times the mutation rate, respectively.     

Xoryp_08180 of Xanthomonas oryzae pv. oryzicola,Encoding a Hypothetical Protein,is Regulated by HrpG and HrpX and Required for Full Virulence in Rice

Xanthomonas oryzae pv. oryzicola (Xoc) causes a destructive bacterial leaf streak disease in rice. Some of the gene products annotated as hypothetical proteins in the genome of Xoc may contribute to its virulence in rice. A mutant, Mxoc1679, screened from our previous Tn5-tagged mutant library for Xoc strain RS105, showed reduced virulence in rice. In this mutant, a gene named as Xoryp_08180 was disrupted by Tn5 insertion. Xoryp_08180 encodes a 1 306-aa hypothetical protein which is highly conserved in Xanthomonas spp. Non-polar mutation of Xoryp_08180 in RS105 strain led to a significant reduction in bacterial virulence and growth in rice, a delayed hypersensitive response (HR) in non-host tobacco, and a decrease in extracellular protease activity. The deficiencies above were restored to wild-type level in the complementary strain by expressing Xoryp_08180 in trans. In addition, the expression of Xoryp_08180 was repressed in hrpG and hrpX mutants in planta but not in a nutrient-rich condition. These results suggested that Xoryp_08180 is a virulence factor required for extracellular protease production, HR induction and full virulence of Xoc.     

薤白EPSP基因对亚麻的转化及检测

薤白EPSP基因编码5-烯醇丙酮莽草酸-3-磷酸合酶,对除草剂草甘瞵具有一定的抗性.利用该基因构建植物表达重组体.薤白EPSPcDNA重组到植物表达载体pWM101中,构建成35S启动子控制的植物组成型表达载体,利用农杆菌介导法将其转入亚麻,经过潮霉素筛选获得了抗性的愈伤组织,诱导分化后获得了亚麻苗.经特异引物的PCR扩增,证明目的基因已经整合到亚麻基因组中,对转基因亚麻愈伤组织及幼苗进行的草甘膦抗性检测表明,转基因亚麻对草甘膦的耐受性明显提高.