毛竹微型颠倒重复序列的鉴定及分子标记开发

MITEs(miniature inverted-repeat transposable elements)又称颠倒重复序列,是缺少转座酶序列的非自主型转座子,在真核生物基因组含量丰富,是基因组多态性形成的重要驱动力之一。该研究利用MITE Tracker软件,在毛竹(Phyllostachys edulis)新版基因组中鉴定到1579个MITEs家族,共包括18373个全长MITEs,占毛竹基因组的0.34%,被归入到7个超家族。超家族的插入时间跨度为0.5~16.5 mya,其中有3个超家族在2~4 mya经历过一次扩增事件;2个超家族分别在1~2 mya和3~5 mya经历两次扩增事件;2个超家族在0.5~16.5 mya经历一次长期扩增。MITEs偏好插入基因或基因附近,且Micron-like超家族偏好插入ATT与ATA之间。该研究开发了3个分子标记,可从12份雷竹变种变型材料中鉴定出4份。综上所述,该文主要分析了毛竹基因组中MITEs的分布、进化及插入情况,并获得了3对可以区分雷竹变种变型的分子标记,为下一步验证MITEs功能奠定基础。     

水稻MITEs类转座子mPing研究进展

Miniature Ping(mPing)是小型反向重复转座子(Miniature Inverted-Repeat Transposable Elements,MITEs)类转座子Tourist-like超家族重要成员,是水稻基因组内检测到的第一个活跃的MITEs,是MITEs大家族中少数低拷贝且可以在自然状态下维持转座活性的成员之一,因此,mPing是转座子相关领域研究的良好素材。该文综合阐述了近年来国内外有关mPing的结构、转座酶供体、激活特性以及对基因组的影响等方面的研究进展,为进一步深入探究MITEs的转座机制以及mPing转座子的开发利用提供资料。     

高等植物转座子的超家族及其应用(综述)

转座子是广泛存在于高等植物基因组中的可移动的DNA分子。文中主要介绍高等植物的各种转座子超家族,包括LTR类反转录转座子、hAT、CACTA因子、Mutator和MULEs、Tc1/mariner、微小反向重复转座子MITEs等;另外还阐述了植物转座子标签体系和筛选方法,以及转座子在生物多样性与遗传连锁分析、植物基因组学研究与植物性状改良方面中的应用。     

Mutator超家族转座子研究进展

转座子是一类可以在基因组中不同遗传位点间移动的DNA序列,在其转移过程中有时会伴随自身拷贝数的增加。作为基因组的重要组成部分,转座子可以通过多种方式影响宿主基因及基因组的结构与功能,进而在宿主的演化过程中扮演重要角色。目前依据转座过程中间体类型的不同可以将其分为I类转座子和II类转座子。Mutator超家族转座子是20世纪70年代在玉米(Zea may L.)中发现的一类特殊的转座子,其属于II类转座子,广泛存在于真核生物基因组中,包含遗传特征明晰可分的众多转座子家族。此外,该超家族转座子转座频率高,倾向于插入基因富含区及低拷贝序列区,可快速产生大量新的突变体,目前已被广泛应用于正向及反向遗传学研究。本文结合近年来相关研究结果,围绕Mutator超家族转座子的分类组成、结构特征、转座机制、插入偏好、靶位点重复序列以及玉米自主性MULEs元件展开综述,并对转座子研究面临的问题及未来研究方向进行了探讨,旨在与研究领域内的同行探讨相关研究的可能突破点、未来发展方向及可能产生的重大影响。     

piggyBac转座子在牛基因组的整合位点及特征分析

piggyBac(PB)转座子作为一种遗传工具被广泛应用于多个物种的转基因及插入突变研究, 目前PB转座子在牛中的相关研究还较少。为了获得PB转座子在牛基因组中的整合位点, 总结其转座特征, 文章构建了PB[CMV-EGFP]和pcDNA-PBase二元转座系统, 利用细胞核电转技术共转染牛耳组织成纤维细胞, 经G-418筛选, 获得了稳定转染EGFP的转基因细胞系; 提取细胞基因组DNA, 利用基因组步移技术扩增PB转座子5′ Bac区插入位置的DNA序列; 通过与牛基因组序列进行BLAST比对, 得到PB转座子在牛基因组中的插入位点。文章共获得了8个有效的整合位点, 但仅有5个位点定位到染色体1、2、11和X染色体上。序列分析表明：在牛基因组中, PB转座子可特异性的插入到“TTAA”位置, 并整合到基因间的非调控区; 分析整合位点“TTAA”相邻一侧的5个碱基组成, 发现PB转座子5′端倾向于插入到GC(62.5%)碱基富集区。该研究表明, PB转座子可以在牛基因组中发生转座, 获得的整合位点信息为利用PB转座子在牛上开展遗传学研究提供了理论参考。     

真核生物转座子鉴定和分类计算方法

重复序列是真核生物基因组的重要组成成分,根据其序列特征及在基因组中的存在形式,可以进一步分为串联重复、片段重复和散在重复。其中,散在重复大多起源于转座子。根据转座介质的不同,转座子又可分为DNA和逆转录转座子。转座子的转座和扩增对基因的进化和基因组的稳定具有显著的影响;同时与其他类型的重复序列相比,转座子的结构和分类更为复杂多样,使得对转座子的鉴定和分类更为复杂和困难。鉴于此,文章简要概括了转座子的功能及分类,总结了真核生物转座子鉴定、分类和注释的3个步骤:(1)重复序列库的构建;(2)重复序列的校正和分类;(3)基因组注释。着重介绍了每一步骤所采用的不同计算方法,比较了不同方法的优缺点。只有把多种方法结合起来使用才能实现全基因组转座子的精确鉴定、分类和注释,这将为转座子的全基因组鉴定和分类提供借鉴意义。     

高等植物helitron转座子的研究进展

作为重复序列的一种主要类型,转座子在高等植物基因组中具有相当丰富的DNA含量,在改变基因结构、调节基因表达、影响基因组进化,以及创造新基因的过程中扮演着重要的角色。Helitron转座子是DNA转座子的一种,在转座过程中经常捕获基因或基因片段,以及插入到基因附近或基因内部,因此在改变基因组构成、影响基因组的进化过程以及改变基因型和表型等方面起着重要作用。该文对国内外近年来有关植物基因组中helitron转座子的结构特征、鉴定和分类方法、基因组中的含量和在染色体上的分布,以及转座扩增和基因片段的捕获等方面的研究进展进行了综述,并对helitron转座子研究过程中存在的问题进行了讨论,对今后helitron相关的研究进行了展望。     

家蚕LTR逆转录转座子的鉴定、分类及系统发育分析

转座子是真核生物基因组的重要组成成分。为了研究家蚕Bombyx mori长末端重复序列 (long terminal repeat, LTR)逆转录转座子的分类及进化, 本研究采用de novo预测和同源性搜索相结合的方法, 在家蚕基因组中共鉴定出了38个LTR逆转录转座子家族, 序列长度占整个基因组的0.64%, 远小于先前预测的11.8%, 其中有6个家族为本研究的新发现。38个家族中, 26个家族有表达序列标签 (expression sequence tag, EST)证据, 表明这些家族具有潜在的活性。对有EST证据的6个家族和没有EST证据的5个家族用RT-PCR进行了组织表达谱实验, 结果表明这11个家族在一些组织中有表达, 这进一步证实了这些家族具有转录活性, 基于此我们推测家蚕中大部分的LTR逆转录转座子家族很可能具有潜在活性。对转座子的插入时间进行估计, 结果表明绝大部分元件都是最近1百万年内插入到家蚕基因组中的。我们还比较了黑腹果蝇Drosophila melanogaster、 冈比亚按蚊Anopheles gambiae和家蚕B. mori中Ty3/Gypsy超家族分支的差异, 结果表明不同枝在不同昆虫中有着不同的扩张。家蚕中LTR逆转录转座子的鉴定和系统分析有助于我们理解逆转录转座子在昆虫进化中的作用。     

Tgf2转座子在团头鲂基因组中的插入效率

文章通过构建带金鱼Tgf2转座子左右臂、斑马鱼肌球蛋白轻链2(Mlyz2)启动子和红色荧光蛋白(RFP)的供体质粒Tgf2-Mlyz2-RFP, 与Tgf2转座酶mRNA共同显微注射入团头鲂1~2细胞期受精卵, 检测金鱼Tgf2转座子在团头鲂基因组中的整合效率。在团头鲂出膜仔鱼、30 d和180 d幼鱼阶段, 可在鱼体背部和侧面肌肉观察到荧光, 红色荧光蛋白的表达率为48.1%, PCR检测结果显示, 金鱼Tgf2转座系统在团头鲂成鱼基因组中的整合效率为31.5%; 对5尾阳性团头鲂进行了RT-PCR检测, 3尾团头鲂在12个组织均能检测到较高的RFP基因的表达, 2尾团头鲂仅在肌肉、皮和肾脏中存在较高的RFP基因的表达, 显示RFP基因在不同转基因团头鲂个体中的组织表达存在一定差异; 通过检测Tgf2转座子在团头鲂基因组插入位置5′端的侧翼序列, 检测出金鱼Tgf2转座系统在转基因团头鲂中的拷贝数至少为2个, 每尾鱼的平均拷贝数大约为5个, 50%以上插入位点的侧翼序列可找出其它脊椎动物的相关同源性序列。研究结果显示金鱼Tgf2转座子可高效介导基因在团头鲂基因组中插入, 为开展团头鲂转基因和基因捕获研究奠定了一定的基础。     

转座子插入宿主基因在稻属中的分布式样及其适应意义的初探

转座子在各类真核生物基因组中都占有很高的比例,它们对宿主基因组特别是关联的基因在结构、功能和进化上都起着重要的作用。基于生物信息学分析,本研究选择了水稻基因组中2个被转座子插入的宿主基因,通过PCR扩增和琼脂糖凝胶电泳分析,获得了转座子在稻属16个代表物种94份材料中的插入式样。结果表明,这2个转座子在稻属中的分布式样与插入时间不同,基因三DG-＆02926349中的转座子在AA-基因组的物种中全部存在,基因LOC-Os02945130中的转座子则插入稻属AA-基因组的部分物种中,与AA-基因组的物种的系统发育关系相吻合。转座子在宿主基因组中不同的分布与保留式样以及插入后已经固定在不同地理来源的群体中,暗示了它们在物种进化过程中对宿主基因可能存在适应性意义。     

Genome-wide characterization and evolution analysis of miniature inverted-repeat transposable elements (MITEs) in moso bamboo (<Emphasis Type="Italic">Phyllostachys heterocycla</Emphasis>)

Main conclusion

Moso bamboo MITEs were genome-wide identified first time, and data shows that MITEs contribute to the genomic diversity and differentiation of bamboo. Miniature inverted-repeat transposable elements (MITEs) are widespread in animals and plants. There are a large number of transposable elements in moso bamboo (Phyllostachys heterocycla var. pubescens) genome, but the genome-wide information of moso bamboo MITEs is not known yet. Here we identified 362 MITE families with a total of 489,592 MITE-related sequences, accounting for 4.74 % of the moso bamboo genome. The 362 MITE families are clustered into six known and one unknown super-families. Our analysis indicated that moso bamboo MITEs preferred to reside in or near the genes that might be involved in regulation of host gene expression. Of the seven super-families, three might undergo major expansion event twice, respectively, during 8–11 million years ago (mya) ago and 22–28 mya ago; two might experience a long expansion period from 6 to 13 mya. Almost 1/3 small RNAs might be derived from the MITE sequences. Some MITE families generate small RNAs mainly from the terminals, while others predominantly from the central region. Given the high copy number of MITEs, many siRNAs and miRNAs derived from MITE sequences and the preferential insertion of MITE into gene regions, MITEs may contribute to the genomic diversity and differentiation of bamboo.

     

The ancient mariner sails again: transposition of the human Hsmar1 element by a reconstructed transposase and activities of the SETMAR protein on transposon ends

Hsmar1, one of the two subfamilies of mariner transposons in humans, is an ancient element that entered the primate genome lineage approximately 50 million years ago. Although Hsmar1 elements are inactive due to mutational damage, one particular copy of the transposase gene has apparently been under selection. This transposase coding region is part of the SETMAR gene, in which a histone methylatransferase SET domain is fused to an Hsmar1 transposase domain. A phylogenetic approach was taken to reconstruct the ancestral Hsmar1 transposase gene, which we named Hsmar1-Ra. The Hsmar1-Ra transposase efficiently mobilizes Hsmar1 transposons by a cut-and-paste mechanism in human cells and zebra fish embryos. Hsmar1-Ra can also mobilize short inverted-repeat transposable elements (MITEs) related to Hsmar1 (MiHsmar1), thereby establishing a functional relationship between an Hsmar1 transposase source and these MITEs. MiHsmar1 excision is 2 orders of magnitude more efficient than that of long elements, thus providing an explanation for their high copy numbers. We show that the SETMAR protein binds and introduces single-strand nicks into Hsmar1 inverted-repeat sequences in vitro. Pathway choices for DNA break repair were found to be characteristically different in response to transposon cleavage mediated by Hsmar1-Ra and SETMAR in vivo. Whereas nonhomologous end joining plays a dominant role in repairing excision sites generated by the Hsmar1-Ra transposase, DNA repair following cleavage by SETMAR predominantly follows a homology-dependent pathway. The novel transposon system can be a useful tool for genome manipulations in vertebrates and for investigations into the transpositional dynamics and the contributions of these elements to primate genome evolution.     

Novel miniature inverted-repeat transposable elements derived from novel <Emphasis Type="Italic">CACTA</Emphasis> transposons were discovered in the genome of the ant <Emphasis Type="Italic">Camponotus floridanus</Emphasis>

Fourteen novel miniature inverted-repeat transposable element (MITE) families are found in the Florida carpenter ant genome, Camponotus floridanus. They constitute approximately 0.63 % of the entire genome. Analysis of their insertion time showed that most members of these MITEs were inserted into their host genome in less than 8 million years ago. In addition, the association between MITEs and the noncoding regions of genes in C. floridanus is random. Interestingly, an autonomous partner (named CfTEC) responsible for the amplification of these MITEs was also found in C. floridanus. Meanwhile, we present evidence, based on searches of publicly available databases, that this autonomous element was widespread in animals. Moreover, structure and phylogenetic analyses supported that TECs might represent a novel cade of transposons intermediate between the classic CACTA transposon and TRCs. Finally, their transposition mechanism and impact on host genome evolution were also discussed.     

Discovery of a large number of previously unrecognized mitochondrial pseudogenes in fish genomes

Nuclear inserted copies of mitochondrial origin (numts) vary widely among eukaryotes, with human and plant genomes harboring the largest repertoires. Numts were previously thought to be absent from fish species, but the recent release of three fish nuclear genome sequences provides the resource to obtain a more comprehensive insight into the extent of mtDNA transfer in fishes. From the sequence analyses of the genomes of Fugu rubripes, Tetraodon nigroviridis, and Danio rerio, we have identified 2, 5, and 10 recent numt integrations, respectively, which integrated into those genomes less than 0.6 million years (Myr) ago. Such results contradict the hypothesis of absence or rarity of numts in fishes, as (i) the ratio of numts to the total size of the nuclear genome in T. nigroviridis was superior to the ratio observed in several higher vertebrate species (e.g., chicken, mouse, and rat), and only surpassed by humans, and (ii) the mtDNA coverage transferred to the nuclear genome of D. rerio is exceeded only by human and mouse, within the whole range of eukaryotic genomes surveyed for numts. Additionally, 335, 336, and 471 old numts (>12.5 Myr) were detected in F. rubripes, T. nigroviridis, and D. rerio, respectively. Surprisingly, old numts are inserted preferentially into known or predicted genes, as inferred for recent numts in human. However, because in fish genomes such integrations are old, they are likely to represent evolutionary successes and they may be considered a potential important evolutionary mechanism for the enhancement of genomic coding regions.     

A RAD-Tag Genetic Map for the Platyfish (Xiphophorus maculatus) Reveals Mechanisms of Karyotype Evolution Among Teleost Fish

Mammalian genomes can vary substantially in haploid chromosome number even within a small taxon (e.g., 3–40 among deer alone); in contrast, teleost fish genomes are stable (24–25 in 58% of teleosts), but we do not yet understand the mechanisms that account for differences in karyotype stability. Among perciform teleosts, platyfish (Xiphophorus maculatus) and medaka (Oryzias latipes) both have 24 chromosome pairs, but threespine stickleback (Gasterosteus aculeatus) and green pufferfish (Tetraodon nigroviridis) have just 21 pairs. To understand the evolution of teleost genomes, we made a platyfish meiotic map containing 16,114 mapped markers scored on 267 backcross fish. We tiled genomic contigs along the map to create chromosome-length genome assemblies. Genome-wide comparisons of conserved synteny showed that platyfish and medaka karyotypes remained remarkably similar with few interchromosomal translocations but with numerous intrachromosomal rearrangements (transpositions and inversions) since their lineages diverged ∼120 million years ago. Comparative genomics with platyfish shows how reduced chromosome numbers in stickleback and green pufferfish arose by fusion of pairs of ancestral chromosomes after their lineages diverged from platyfish ∼195 million years ago. Zebrafish and human genomes provide outgroups to root observed changes. These studies identify likely genome assembly errors, characterize chromosome fusion events, distinguish lineage-independent chromosome fusions, show that the teleost genome duplication does not appear to have accelerated the rate of translocations, and reveal the stability of syntenies and gene orders in teleost chromosomes over hundreds of millions of years.     

A family of Tc1-like transposons from the genomes of fishes and frogs: evidence for horizontal transmission.

Tc1-like transposons are very widely distributed within the genomes of animal species. They consist of an inverted repeat sequence flanking a transposase gene with homology to the mobile DNA element, Tc1 of the nematode Caenorhabditis elegans. These elements seem particularly to infest the genomes of fish and amphibian species where they can account for 1% of the total genome. However, all vertebrate Tc1-like elements isolated so far are non-functional in that they contain multiple frameshifts within their transposase coding regions. Here I describe a Tc1-like transposon (PPTN) from the genome of a marine flatfish species (Pleuronectes platessa) which bears conserved inverted repeats flanking an apparently intact transposase gene. Closely related, although degenerate, Tc1-like transposons were also isolated from the genomes of Atlantic salmon (SSTN, Salmo salar) and frog (RTTN, Rana temporaria). Consensual nucleic acid sequences were derived by comparing several individual isolates from each species and conceptual amino acid sequences were thence derived for their transposases. Phylogenetic analysis of these sequences with previously isolated Tc1-like transposases shows that the elements from plaice, salmon and frog comprise a new subfamily of Tc1-like transposons. Each member is distinct in that it is not found in the genomes of the other species tested. Plaice genomes contain about 300 copies of PPTN, salmon 1200 copies of SSTN and frog genomes about 500 copies of RTTN. The presence of these closely related elements in the genomes of fish and frog species, representing evolutionary lines, which diverged more than 400 million years ago, is not consistent with a vertical transmission model for their distributions.     

Plant MITEs: useful tools for plant genetics and genomics

MTTEs (Miniature inverted-repeat transposabie elements) are reminiscence ot non-autonomous DNA (class Ⅱ) elements, which are distinguished from other transposable elements by their small size, short terminal inverted repeats (TIRs), high copy numbers, genie preference, and DNA sequence identity among family members. Although MITEs were first discovered in plants and still actively reshaping genomes, they have been isolated from a wide range of eukaryotic organisms. MITEs can be divided into Tourist-like, Stowaway-like, and pogo-like groups, according to similarities of their TIRs and TSDs (target site duplications). In despite of several models to explain the origin and amplification of MITEs, their mechanisms of transposition and accumulation in eukaryotic genomes remain poorly understood owing to insufficient experimental data. The unique properties of MITEs have been exploited as useful genetic tools for plant genome analysis. Utilization of MITEs as effective and informative genomic markers and pot     

Origin and evolution of avian microchromosomes

The origin of avian microchromosomes has long been the subject of much speculation and debate. Microchromosomes are a universal characteristic of all avian species and many reptilian karyotypes. The typical avian karyotype contains about 40 pairs of chromosomes and usually 30 pairs of small to tiny microchromosomes. This characteristic karyotype probably evolved 100-250 million years ago. Once the microchromosomes were thought to be a non-essential component of the avian genome. Recent work has shown that even though these chromosomes represent only 25% of the genome; they encode 50% of the genes. Contrary to popular belief, microchromosomes are present in a wide range of vertebrate classes, spanning 400-450 million years of evolutionary history. In this paper, comparative gene mapping between the genomes of chicken, human, mouse and zebrafish, has been used to investigate the origin and evolution of avian microchromosomes during this period. This analysis reveals evidence for four ancient syntenies conserved in fish, birds and mammals for over 400 million years. More than half, if not all, microchromosomes may represent ancestral syntenies and at least ten avian microchromosomes are the product of chromosome fission. Birds have one of the smallest genomes of any terrestrial vertebrate. This is likely to be the product of an evolutionary process that minimizes the DNA content (mostly through the number of repeats) and maximizes the recombination rate of microchromosomes. Through this process the properties (GC content, DNA and repeat content, gene density and recombination rate) of microchromosomes and macrochromosomes have diverged to create distinct chromosome types. An ancestral genome for birds likely had a small genome, low in repeats and a karyotype with microchromosomes. A "Fission-Fusion Model" of microchromosome evolution based on chromosome rearrangement and minimization of repeat content is discussed.     

鱼类特异的基因组复制

辐鳍鱼类是脊椎动物中种类最多、分布最广的类群,其基因组大小不等。过去的观点认为,在脊椎动物进化历程中曾发生了两次基因组复制。近期的系统基因组学研究资料进一步提出,在大约350百万年,辐鳍鱼还发生了第三次基因组复制,即鱼类特异的基因组复制(fish-specificgenomeduplication,FSGD),且发生的时间正处在“物种极度丰富”的硬骨鱼谱系(真骨总目)和“物种贫乏”的谱系(辐鳍鱼纲基部的类群)出现分歧的时间点,表明FSGD与硬骨鱼物种和生物多样性的增加有关。进一步开展鱼类比较基因组学和功能基因组学研究将进一步验证FSGD这一假说。