人类基因组中加工假基因分布与重组率和基因密度的关系

分析了人类加工假基因在染色体上的分布,发现加工假基因密度与重组率负相关,而与基因密度正相关。加工假基因在低重组区的积累与插入有害模型和异位重组模型相吻合：在插入有害模型下,低重组区的选择强度由于Hill．Robertson干涉而变弱,所以加工假基因较多地插入到低重组区;在异位重组模型下,同源加工假基因家族（包括同源祖先基因）之内可能发生异位重组而对机体造成危害,所以加工假基因在高重组区的插入受到较强的负选择,导致加工假基因较多地分布在低重组区。除以上两种模型以外,加工假基因还可能通过降低重组率的方式对加工假基因密度与重组率的负相关有所贡献。加工假基因偏好分布在基因密区,这可能与异位重组在该区较少发生有关。     

人类基因组上的假基因

假基因是基因组上与编码基因序列非常相似的非功能性基因组DNA拷贝,一般情况都不被转录,且没有明确生理意义。假基因根据其来源可分为复制假基因和已加工假基因。迄今为止,明确鉴定的人类假基因多为已加工假基因,有8000个之多。在Swiss-Prot/TrEMBL收录的编码蛋白质的将近25500个基因序列中,约10％在基因组中有一个或多个近全长已加工假基因。其余的功能基因都没有已加工假基因。核糖体蛋白基因具有最多数量的已加工假基因,约有l700个(占已加工假基因数的22％),少数基因,如cyclophilinA、肌动蛋白(actin)、角蛋白(keratin)、GAPDH、细胞色素C(cytochromec)和nucleophosmin等则有很多份已加工假基因。总体上讲,假基因在人类染色体上的分布与染色体长度成比例,但已加工假基因在GC含量为41％～46％的染色体区域密度最高。已加工假基因的拷贝数和功能基因在生殖器官中的表达高度一致,说明许多假基因发生在胚胎阶段,另外也和基因中GC含量和基因大小密切相关。假基因的准确鉴定对基因组进化、分子医学研究和医学应用具有重要意义。     

脊椎动物假基因

假基因(pseudogene)是出现在特定种群基因组中无功能的基因拷贝,在不同的生命形式特别是脊椎动物中较为常见,未加工假基因和加工假基因的产生机制不同,在分子遗传学领域,识别假基因结构和功能有非常重要的作用。     

青蟹线粒体COI假基因的分离和特征分析

线粒体DNA标记在遗传结构和系统进化研究中得到广泛应用,然而核假基因的存在对此有很大威胁。本文以中国东南沿海的青蟹(Scylla paramamosain)为研究对象,利用线粒体COI基因的通用引物和特异性引物进行扩增,分别得到34个假基因（nuclear mitochondrial pseudogenes, Numts）和5个线粒体COI基因序列。在所获得的34个假基因中共定义了29种单倍型,根据序列的相似度,这些假基因可以分为2类,每类假基因都有各自保守的核苷酸序列。第Ⅰ类假基因存在2处插入序列和1处8 bp的缺失序列,这些位点导致了整个阅读框的移位;在第Ⅱ类假基因和5个线粒体COI序列中只有碱基替换,未发现插入和缺失序列。实验结果分析表明,这两类假基因分别代表了2次核整合事件,即核转移事件的最低值。研究结果提示了     

山茶属植物ITS的扩增及其序列特征分析

以山茶属(Camellia)植物为实验材料,通过在PCR反应液中加入二甲基亚砜(DMSO)来改进ITS区的扩增效果,并对获得的序列进行了相关的比较分析以及系统发育分析。分析结果表明:(1)PCR反应液中添加4%的DMSO能明显提高ITS的扩增效率,并能有效防止ITS假基因的扩增;(2)与功能拷贝相比,ITS假基因序列的ITS1和ITS2区存在多处碱基缺失,GC含量明显偏低,5.8S区二级结构稳定性降低;(3)应用ITS假基因进行种水平的系统发育分析会对其真实的系统关系造成影响,提示进行系统进化研究时应警惕假基因的干扰。     

假基因的功能及其在癌症疾病中的重要作用

假基因是一段具有与功能基因相似的DNA序列,但由于存在许多突变以致失去了原有的功能。过去的研究认为假基因是没有功能的DNA片段,是基因组进化过程中产生的噪音。然而,随着分子生物学技术的发展,越来越多的研究证明了假基因具有重要的生物学功能。假基因可与功能基因竞争性结合miRNA,从而调控功能基因的表达;假基因还可产生内源性小干扰RNA抑制功能基因的表达;甚至有的假基因还可以编码具有功能的蛋白质。文章通过假基因的分类、假基因的识别、假基因的功能和假基因与癌症疾病的关系等方面综述了假基因研究的最新进展。     

假基因的组成、分布及其分子进化

假基因(pseudogene)是指基因组中与正常基因序列相似, 但是缺乏功能的DNA 序列。通过序列同源性搜索, 可以收集基因组中假基因的群体特性、染色体分布和同源家族等特性。假基因很好地保留了数百万年前基因组中祖先基因的分子记录, 被视为“基因化石”, 因此假基因在进化和比较基因组学中是重要的资源。应用假基因和基因比较体系, 可以探究生物基因的进化史和基因组稳定性。如: 用Ka/Ks比值确定假基因的自然选择压、物种亲缘关系和进化距离, 分析假基因自身的进化趋势, 探讨DNA 突变的成因等。     

苏铁nrDNA ITS区的序列多态性:不完全致同进化的证据

本研究对苏铁(Cycas revoluta) nrDNA ITS进行克隆测序, 并以cDNA ITS为参照, 比较分析获得的序列的碱基变异、GC含量、5.8S二级结构的稳定性和5.8S保守基序的有无以及系统发育关系。结果发现苏铁nrDNA ITS存在较高的基因组内多样性, 同时, 这些分化的nrDNA ITS拷贝中包含有假基因的存在, 而且假基因与功能拷贝之间已经形成了较大的遗传分化, 这暗示假基因起源有较长历史。苏铁核仁组织区不仅多达16个, 而且分布在13条染色体上, 这可能是其nrDNA ITS致同进化不完全的主要原因。     

鲸类视觉退化分子机制的研究：基于GNAT2和CNGB3基因的进化分析

鲸类是一类次生性的水生哺乳动物,其陆生祖先大约53-56Ma从陆地返回海洋。为了适应水下的弱光环境,鲸类的光感受器以视杆细胞为主,视锥细胞功能大多退化,缺乏辨别颜色的能力,然而鲸类视觉退化的分子机制尚不清楚。本文选择在视锥细胞中表达且对光传导级联反应起重要作用的GNAT2和CNGB3基因,通过PCR扩增、测序以及在数据库中下载已有的基因序列,共获得8个鲸类代表性物种的同源序列。MEGA6.0软件比对发现侏儒抹香鲸和抹香鲸的GNAT2基因分别在148位和1012位插入了1个碱基 A,而抹香鲸的CNGB3分别在554位和1407位有1个碱基的缺失,导致提前终止密码子出现;另外,小须鲸CNGB3基因在1525-1527位出现了提前终止密码子,提示鲸类的这两个基因可能为假基因。通过I-TASSER在线预测GNAT2和CNGB3蛋白的三维结构,发现出现移码突变终止密码子的位置均位于这两个基因的重要功能域。另外,运用PAML软件的Branch model分析表明发生假基因的鲸类物种GNAT2和CNGB3基因出现选择压力放松,且假基因化可能是一个近期事件。侏儒抹香鲸、抹香鲸以及小须鲸的GNAT2和CNGB3基因出现假基因可能与其深潜习性以及完全的水生生境导致其色觉功能丢失相关。此外,Free-ratio model分析发现其他鲸类的ω值接近于1,说明GNAT2和CNGB3基因出现了选择压力放松,这可能是长期适应水生生境视觉退化的结果。     

鲸类视觉退化分子机制的研究：基于GNAT2和CNGB3基因的进化分析

鲸类是一类次生性的水生哺乳动物,其陆生祖先大约53-56Ma从陆地返回海洋。为了适应水下的弱光环境,鲸类的光感受器以视杆细胞为主,视锥细胞功能大多退化,缺乏辨别颜色的能力,然而鲸类视觉退化的分子机制尚不清楚。本文选择在视锥细胞中表达且对光传导级联反应起重要作用的GNAT2和CNGB3基因,通过PCR扩增、测序以及在数据库中下载已有的基因序列,共获得8个鲸类代表性物种的同源序列。MEGA6.0软件比对发现侏儒抹香鲸和抹香鲸的GNAT2基因分别在148位和1012位插入了1个碱基 A,而抹香鲸的CNGB3分别在554位和1407位有1个碱基的缺失,导致提前终止密码子出现;另外,小须鲸CNGB3基因在1525-1527位出现了提前终止密码子,提示鲸类的这两个基因可能为假基因。通过I-TASSER在线预测GNAT2和CNGB3蛋白的三维结构,发现出现移码突变终止密码子的位置均位于这两个基因的重要功能域。另外,运用PAML软件的Branch model分析表明发生假基因的鲸类物种GNAT2和CNGB3基因出现选择压力放松,且假基因化可能是一个近期事件。侏儒抹香鲸、抹香鲸以及小须鲸的GNAT2和CNGB3基因出现假基因可能与其深潜习性以及完全的水生生境导致其色觉功能丢失相关。此外,Free-ratio model分析发现其他鲸类的ω值接近于1,说明GNAT2和CNGB3基因出现了选择压力放松,这可能是长期适应水生生境视觉退化的结果。     

Evolution of human cytoplasmic actin gene sequences: chromosome mapping and structural characterizations of three cytoplasmic actin-like pseudogenes including one on the Y chromosome

Eight recombinant phage clones containing cytoplasmic actin-like gene sequences have been isolated from a human genomic library for structural characterization. Kpn I family repeat sequences flank six of these actin genes isolated, and Alu family repeats are scattered throughout the DNA inserts of all eight phage clones. Three of these genes are γ actin-like, and the other five are β actin-like. The complete nucleotide sequence analysis of one β and one γ actin-like genes and their flanking regions demonstrates that they both are processed pseudogenes. Using unique DNA sequences flanking these two pseudogenes as hybridization probes for human-mouse somatic cell hybrid DNAs, we have mapped the two actin pseudogenes on human chromosomes 8 and 3, respectively. We have also determined the DNA sequence of a human Y chromosome-linked, processed actin pseudogene. The different values of sequence divergence of these processed pseudogenes and their functional counterparts allow us to estimate the time of generation of the pseudogenes. The results suggest that the cDNA insertion events generating the human cytoplasmic actin-like pseudogenes have occurred at significantly different times during the evolution of primates, after their separation from other mammalian species.     

Isolation and characterization of rat and human glyceraldehyde-3-phosphate dehydrogenase cDNAs: genomic complexity and molecular evolution of the gene.

Full length cDNAs encoding the glycolytic enzyme glyceraldehyde-3-phosphate dehydrogenase (GAPDH) from rat and man have been isolated and sequenced. Many GAPDH gene-related sequences have been found in both genomes based on genomic blot hybridization analysis. Only one functional gene product is known. Results from genomic library screenings suggest that there are 300-400 copies of these sequences in the rat genome and approximately 100 in the human genome. Some of these related sequences have been shown to be processed pseudogenes. We have isolated several rat cDNA clones corresponding to these pseudogenes indicating that some pseudogenes are transcribed. Rat and human cDNAs are 89% homologous in the coding region, and 76% homologous in the first 100 base pairs of the 3'-noncoding region. Comparison of these two cDNA sequences with those of the chicken, Drosophila and yeast genes allows the analysis of the evolution of the GAPDH genes in detail.     

Similar integration but different stability of Alus and LINEs in the human genome

Alus and LINEs (LINE1) are widespread classes of repeats that are very unevenly distributed in the human genome. The majority of GC-poor LINEs reside in the GC-poor isochores whereas GC-rich Alus are mostly present in GC-rich isochores. The discovery that LINES and Alus share similar target site duplication and a common AT-rich insertion site specificity raised the question as to why these two families of repeats show such a different distribution in the genome. This problem was investigated here by studying the isochore distributions of subfamilies of LINES and Alus characterized by different degrees of divergence from the consensus sequences, and of Alus, LINEs and pseudogenes located on chromosomes 21 and 22. Young Alus are more frequent in the GC-poor part of the genome than old Alus. This suggests that the gradual accumulation of Alus in GC-rich isochores has occurred because of their higher stability in compositionally matching chromosomal regions. Densities of Alus and LINEs increase and decrease, respectively, with increasing GC levels, except for the telomeric regions of the analyzed chromosomes. In addition to LINEs, processed pseudogenes are also more frequent in GC-poor isochores. Finally, the present results on Alu and LINE stability/exclusion predict significant losses of Alu DNA from the GC-poor isochores during evolution, a phenomenon apparently due to negative selection against sequences that differ from the isochore composition.     

Identification of three human pseudogenes for subunit VIb of cytochrome c oxidase: a molecular record of gene evolution.

Three pseudogenes for the nuclear-encoded subunit VIb of cytochrome c oxidase (COX) were isolated by screening a human genomic library with cloned human cDNA coding for COX subunit VIb. The nucleotide sequences of the pseudogenes, designated psi COX6b-1, psi COX6b-2 and psi COX6b-3, were determined. Pseudogene psi COX6b-1 bears all the hallmarks of a processed pseudogene and diverged from the parental gene after the divergence of man and cow. Alu repetitive elements were integrated into the structural sequences of the other two pseudogenes. Comparison with the human and bovine cDNA sequences encoding COX subunit VIb suggests that psi COX6b-2 and psi COX6b-3 were formed earlier in evolution than psi COX6b-1. Genomic Southern analysis indicated that a few more pseudogenes for COX subunit VIb are likely to be present in the human genome. Identical nt differences with respect to the human cDNA sequence in the pseudogenes provide some clues on the evolution of the ancestral gene coding for COX subunit VIb.     

Structure and chromosomal distribution of human mitochondrial pseudogenes

Nuclear mitochondrial pseudogenes (Numts) have been found in the genome of many eukaryote species, including humans. Using a BLAST approach, we found 1105 DNA sequences homologous to mitochondrial DNA (mtDNA) in the August 2001 Goldenpath human genome database. We assembled these sequences manually into 286 pseudogenes on the basis of single insertion events and constructed a chromosomal map of these Numts. Some pseudogenes appeared highly modified, containing inversions, deletions, duplications, and displaced sequences. In the case of four randomly selected Numts, we used PCR tests on cells lacking mtDNA to ensure that our technique was free from genome-sequencing artifacts. Furthermore, phylogenetic investigation suggested that one Numt, apparently inserted into the nuclear genome 25-30 million years ago, had been duplicated at least 10 times in various chromosomes during the course of evolution. Thus, these pseudogenes should be very useful in the study of ancient mtDNA and nuclear genome evolution.