HBV基因型分析及B/C型共感染的血清病毒种群分析

 为了解HBV大三阳青年妇女的HBV基因型及体内病毒种群特征,从19~35岁青年妇女HBV感染者血清样本中筛选出24例HBV血清学标志物检测结果为大三阳的样本进行HBV基因分型检测,14例为B型,9例为C型,1例为B/C基因型共感染。对14例大S蛋白基因通过特异性扩增、序列克隆和测序确认的分型结果与分型检测一致。发现的1例B/C共感染样本来自阿德福韦酯抗病毒治疗35周患者,大S蛋白基因序列分析显示10条序列中4条为B型,准种分散值为2.8;其6条为C型,准种分散值为0.6。α决定簇分析该患者体内病毒种群包含3种血清型。病毒种群分析结果提示本地区青年妇女HBV感染的基因型主要为B型和C型,B/C基因型共感染患者体内HBV病毒种群呈复杂的多基因型分布,不同基因型内部以准种形式存在。     

A draft sequence of the rice (Oryza sativa ssp. indicd) genome

The sequence of the rice genome holds fundamental information for its biology, including physiology, genetics, development, and evolution, as well as information on many beneficial phenotypes of economic significance. Using a “whole genome shotgun” approach, we have produced a draft rice genome sequence ofOryza sativa ssp.indica, the major crop rice subspecies in China and many other regions of Asia. The draft genome sequence is constructed from over 4.3 million successful sequencing traces with an accumulative total length of 2214.9 Mb. The initial assembly of the non-redundant sequences reached 409.76 Mb in length, based on 3.30 million successful sequencing traces with a total length of 1797.4 Mb from anindica variant cultivar93-11, giving an estimated coverage of 95.29% of the rice genome with an average base accuracy of higher than 99%. The coverage of the draft sequence, the randomness of the sequence distribution, and the consistency of BIG-ASSEMBLER, a custom-designed software package used for the initial assembly, were verified rigorously by comparisons against finished BAC clone sequences from bothindica andjapanica strains, available from the public databases. Over all, 96.3% of full-length cDNAs, 96.4% of STS, STR, RFLP markers, 94.0% of ESTs and 94.9% unigene clusters were identified from the draft sequence. Our preliminary analysis on the data set shows that our rice draft sequence is consistent with the comman standard accepted by the genome sequencing community. The unconditional release of the draft to the public also undoubtedly provides a fundamental resource to the international scientific communities to facilitate genomic and genetic studies on rice biology. These authors contributed equally to this work.     

Rapid genome evolution in Pms 1 region of rice revealed by comparative sequence analysis

Pms1, a locus for photoperiod sensitive genic male sterility in rice, was identified and mapped to chromosome 7 in previous studies. Here we report an effort to identify the candidate genes for Pms1 by comparative sequencing of BAC clones from two cultivars Minghui 63 and Nongken 58, the parents for the initial mapping population. Annotation and comparison of the sequences of the two clones resulted in a total of five potential candidates which should be functionally tested. We also conducted com-parative analysis of sequences of these two cultivars with two other cultivars, Nipponbare and 93-11, for which sequence data were available in public databases. The analysis revealed large differences in sequence composition among the four genotypes in the Pms1 region primarily due to retroelement activity leading to rapid recent growth and divergence of the genomes. High levels of polymorphism in the forms of indels and SNPs were found both in intra- and inter-subspecific comparisons. Dating analysis using LTRs of the retroelements in this region showed that the substitution rate of LTRs was much higher than reported in the literature. The results provided strong evidence for rapid genomic evolution of this region as a consequence of natural and artificial selection.     

HBV基因型与干扰素及核苷类似物抗病毒治疗疗效的关系

目的:介绍乙型肝炎病毒(HBV)基因型与抗病毒治疗疗效关系的最新进展.方法:综合分析近年国内外有关HBV基因型与抗病毒治疗疗效关系的文献资料.结果:感染HBV基因型A的慢性乙型肝炎患者对干扰素治疗的应答率高于基因型D,基因型B高于基因型C;HBV基因型与核苷类似物治疗疗效的关系目前尚存争议;基因型E、F、G、H是否影响抗病毒疗效报道较少.结论:HBV基因型与抗病毒治疗疗效存在一定的相关性;HBV基因型的研究将有助于临床抗病毒治疗方案的优化.     

Hepatitis B virus DNA integration in a sequence homologous to v-erb-A and steroid receptor genes in a hepatocellular carcinoma

Hepatitis B virus (HBV) is clearly involved in the aetiology of human hepatocellular carcinoma (HCC) and the finding of HBV DNA integration into human liver DNA in almost all HCCs studied suggested that these integrated viral sequences may be involved in liver oncogenesis. Several HBV integrations in different HCCs and HCC-derived cell lines have been analysed after molecular cloning without revealing any obvious role for HBV. From a comparison of a HBV integration site present in a particular HCC with the corresponding unoccupied site in the non-tumorous tissue of the same liver, we now report that HBV integration places the viral sequence next to a liver cell sequence which bears a striking resemblance to both an oncogene (v-erb-A) and the supposed DNA-binding domain of the human glucocorticoid receptor and human oestrogen receptor genes. We suggest that this gene, usually silent or transcribed at a very low level in normal hepatocytes, becomes inappropriately expressed as a consequence of HBV integration, thus contributing to the cell transformation.     

Presence of integrated hepatitis B virus DNA sequences in cellular DNA of human hepatocellular carcinoma

Hepatitis B virus (HBV) may be one of the agents involved in the aetiology of human primary liver cancer. This hypothesis is supported by (1) the similarity between the geographical distribution of chronic carriers of the viral surface antigen (HBsAg) and that of hepatocellular carcinoma (HCC); (2) the increase in the prevalence of HBV markers in serum of patients with primary liver cancer when compared with the general population; (3) the observation that HBV infection precedes the development of the tumour. Moreover, these epidemiological indications of an association between HBV infecton and hepatocellular carcinoma are supported by the detection of HBV markers such as HBsAg or viral DNA sequences, although in a non-integrated form in tumour tissue. To study the relationship between HBV and primary liver cancer further, we looked for the presence of free or integrated viral DNA in tumour tissue of human hepatocellular carcinomas and in a HBsAg-producing human hepatoma cell line. Using the blot-transfer hybridization technique and cloned HBV DNA as a probe, we have now demonstrated that the viral DNA is integrated in the cellular genome both in tumour tissue and in a hepatoma cell line.     

Comparative analyses of multi-species sequences from targeted genomic regions

The systematic comparison of genomic sequences from different organisms represents a central focus of contemporary genome analysis. Comparative analyses of vertebrate sequences can identify coding and conserved non-coding regions, including regulatory elements, and provide insight into the forces that have rendered modern-day genomes. As a complement to whole-genome sequencing efforts, we are sequencing and comparing targeted genomic regions in multiple, evolutionarily diverse vertebrates. Here we report the generation and analysis of over 12 megabases (Mb) of sequence from 12 species, all derived from the genomic region orthologous to a segment of about 1.8 Mb on human chromosome 7 containing ten genes, including the gene mutated in cystic fibrosis. These sequences show conservation reflecting both functional constraints and the neutral mutational events that shaped this genomic region. In particular, we identify substantial numbers of conserved non-coding segments beyond those previously identified experimentally, most of which are not detectable by pair-wise sequence comparisons alone. Analysis of transposable element insertions highlights the variation in genome dynamics among these species and confirms the placement of rodents as a sister group to the primates.     

肝癌染色体高频缺失区肿瘤相关基因cSNP在HBV患者和正常人群中的多态分布研究

从定位于肝癌高频缺失区的肿瘤相关基因入手,查询单核苷酸多态性(SNP)数据库信息,获得编码区SNP(cSNP)序列,设计引物,根据SNP位点设计寡核苷酸探针,构建SNP芯片．分别从正常人和HBV患者血样中提取基因组DNA,PCR扩增标记含SNP位点的序列,将地高辛标记的PCR产物与SNP芯片杂交．结果表明,正常人基因组与HBV患者基因组肿瘤相关基因SNP之间存在差异,检测到EGFL3(rs947 345),Gas- pase9(rs2 308 950),E2F2(rs3 218 171)三个cSNP位点的基因频率在两组人群中差异显著．HBV患者中存在的高频多态位点可能与其肝癌易感性相关．     

Numerous potentially functional but non-genic conserved sequences on human chromosome 21

The use of comparative genomics to infer genome function relies on the understanding of how different components of the genome change over evolutionary time. The aim of such comparative analysis is to identify conserved, functionally transcribed sequences such as protein-coding genes and non-coding RNA genes, and other functional sequences such as regulatory regions, as well as other genomic features. Here, we have compared the entire human chromosome 21 with syntenic regions of the mouse genome, and have identified a large number of conserved blocks of unknown function. Although previous studies have made similar observations, it is unknown whether these conserved sequences are genes or not. Here we present an extensive experimental and computational analysis of human chromosome 21 in an effort to assign function to sequences conserved between human chromosome 21 (ref. 8) and the syntenic mouse regions. Our data support the presence of a large number of potentially functional non-genic sequences, probably regulatory and structural. The integration of the properties of the conserved components of human chromosome 21 to the rapidly accumulating functional data for this chromosome will improve considerably our understanding of the role of sequence conservation in mammalian genomes.     

HBs和HCVc基因双顺反子质粒在BALB/c小鼠体内的分布

 采用注射含双顺反子质粒后,PCR法扩增不同组织中HBsAg和HCVc基因,同时检测HBV和HCV抗体应答水平.研究注射基因免疫用双顺反子质粒在小鼠组织中的分布.pcDNA3.0BApc154S₂S 1次注射BALB/c小鼠后,24 h内主要分布于血液、肝、脾、骨髓、淋巴结、注射部位肌肉、肺和肾等含血液丰富的组织中.注射后2 d主要在骨髓、血液和注射部位肌肉中存在.7 d后仅在注射部位肌肉中能检测出来,并可持续到第9周.组织切片镜检质粒注射初期肌肉细胞轻度浊肿,随后肌膜细胞轻度增生,未见其它明显的组织病理学变化.质粒多次免疫BALB/c小鼠未见明显的临床症状和病理组织学变化.质粒在小鼠体内不同组织存在时间不一致.     

RNA interference-mediated inhibition of Hepatitis B Virus replication

Persistent and recurrent infection of hepatitis B virus (HBV) represents one of the most common and severe viral infections of humans, and has caused a formidable health problem in the affected countries. Currently used antiviral drugs have a very limited success on controlling HBV replication and infection. RNA interference (RNAi), a process by which double-stranded RNA (dsRNA) directs sequence-specific degradation of target mRNA in mammalian and plant cells, has recently been used to knockdown gene expression in various species. In this study, we sought to determine whether RNAi-mediated silencing of HBV viral gene expression could lead to the effective inhibition of HBV replication. We first developed RNAi vectors that expressed small interfering RNA (siRNA) and targeted the HBV core or surface gene sequence. Our results demonstrated that these specific siRNAs efficiently reduced the levels of corresponding viral RNAs and proteins, and thus suppressed viral replication. Treatment with siRNA gave the greatest reduction in the levels of HBsAg (92%) and in HBeAg (85%) respectively in the cultured cell medium. Our findings further demonstrated that the RNAi-mediated antiviral effect was sequence-specific and dose-dependent. Therefore, our findings strongly suggest that RNAi-mediated silencing of HBV viral genes could effectively inhibit the replication of HBV, hence RNAi-based strategy should be further explored as a more efficacious antiviral therapy of HBV infection.     

Biodiversity and biogeography of phages in modern stromatolites and thrombolites

Viruses, and more particularly phages (viruses that infect bacteria), represent one of the most abundant living entities in aquatic and terrestrial environments. The biogeography of phages has only recently been investigated and so far reveals a cosmopolitan distribution of phage genetic material (or genotypes). Here we address this cosmopolitan distribution through the analysis of phage communities in modern microbialites, the living representatives of one of the most ancient life forms on Earth. On the basis of a comparative metagenomic analysis of viral communities associated with marine (Highborne Cay, Bahamas) and freshwater (Pozas Azules II and Rio Mesquites, Mexico) microbialites, we show that some phage genotypes are geographically restricted. The high percentage of unknown sequences recovered from the three metagenomes (>97%), the low percentage similarities with sequences from other environmental viral (n = 42) and microbial (n = 36) metagenomes, and the absence of viral genotypes shared among microbialites indicate that viruses are genetically unique in these environments. Identifiable sequences in the Highborne Cay metagenome were dominated by single-stranded DNA microphages that were not detected in any other samples examined, including sea water, fresh water, sediment, terrestrial, extreme, metazoan-associated and marine microbial mats. Finally, a marine signature was present in the phage community of the Pozas Azules II microbialites, even though this environment has not been in contact with the ocean for tens of millions of years. Taken together, these results prove that viruses in modern microbialites display biogeographical variability and suggest that they may be derived from an ancient community.     

大理吸毒人群HBV感染的调查及基因型研究

目的：调查大理吸毒人群乙型肝炎病毒（HBV）的基因型和亚型。方法：采集大理市某戒毒所吸毒人群血清共192份，采用酶联吸附免疫法（ELISA）测定HBV血清标志物（HBVM），再采用基因型特异性引物PCR和聚合酶链反应-限制性片段长度多态性（PCR—RFLP）方法测定HBV感染血清基因型和亚型。结果：192份血清中，HBsAg阳性率8．85％（17／192）。17份HBsAg阳性标本中基因型分别为：B型3例，均为Ba型；C型1例，为Cs型；B＋C混合型3例（Ba＋Cs）；未扩增10例。结论：大理吸毒人群HBV感染率较高；HBV感染的基因型为B型、C型和B＋C型，基因亚型为Ba和Cs。     

Ancient DNA sequences of rice from the low Yangtze reveal significant genotypic divergence

Rice (Oryza sativa) was first domesticated in the lower and middle Yangtze regions of China, and rice remains have been found in many Chinese archaeological sites. Until now, only phenotypic archeobotanical evidence, such as the spikelet bases of ancient grains, has been used to speculate on the domestication process and domestication rate of rice. In this study, we sequenced 4 genomic segments from rice remains in Tianluoshan, a site of the local Hemudu Neolithic culture in the low Yangtze and two other archaeological sites (??2400 and 1200 BC, respectively). We compared our sequences with those of the current domesticated and wild rice (O. rufipogon) populations. At least two genotypes were found in the remains from each site, suggesting a heterozygotic state of the rice seeds. One ancient genotype was not found in the current domesticated population and might have been lost. The rice remains belonged to the japonica group, and most if not all were japonica-type, suggesting that the remains might be at an early stage of indica-japonica divergence or an indica-japonica mixture. We also identified sequences with significant similarity to those from species of Sapindales, Zygophyllales, and Brassicales, which is consistent with the identification of other plant remains in the Tianluoshan site and the common rice field weeds such as mustards in southern China.     

HIV-1和HBV复合型DNA免疫的初步研究

近年的研究表明,在啮齿类和非人灵长类免疫带有编码病毒和细菌抗原基因的质粒ＤＮＡ可以激发体液和细胞免疫应答．在本实验中,将ＨＢＶ的Ｓ基因和ＨＩＶ－１的ｇｐ１６０基因以融合形式插入到载体ｐｃＤＮＡ３中,其能表达ＨＢｓＡｇ和ｇｐ１６０的融合蛋白,并将此质粒ＤＮＡ分别直接注射到Ｂａｌｂ／ｃ小鼠和Ｓｗｉｓ小鼠．三次免疫后,用ＥＬＩＳＡ的方法初步检测ＨＢｓＡｇ和ｇｐ１６０抗原特异的抗体免疫应答均为阳性．结果说明,带有ＨＢＶ和ＨＩＶ－１融合基因的质粒ＤＮＡ直接免疫小鼠后,均激发了小鼠产生相应的免疫应答反应,这个结果为研究和生产多价疫苗提供了新的思路     

DNA序列作为信息隐藏载体的研究

研究了DNA序列能否成为信息隐藏的载体.通过对DNA序列进行分析证实了:由于核苷酸序列中有强的随机噪声,DNA序列可以作为信息隐藏的载体;进而通过对DNA序列特征进行分析,提出了一个嵌入对策:秘密消息嵌入非编码区的高复杂度区域有很好的安全性.该文的研究对提出以DNA序列为载体的信息隐藏算法具有重要的指导作用.     

50例中国人IX因子基因的限制性片段长度多态性

用IX因子基因内探针F9(VⅢ)对TaqI,BamHI和EcoRI酶切的50例中国人基因组DNA进行杂交分析。结果表明,所有个体经TaqI酶切的杂交片段为4.5kb和1.8kb,BamHI和EcoRI酶切的杂交片段分别为23kb和5.0 kb。基因组DNA样本中未发现限制性片段长度多态性(RFLP),这与欧美国家的民族群体中存在着IX因子基因内TaqI和BamHI的RFLP的结论不同。造成不同种族间DNA水平差异的原因,很可能与长期在不同地理环境中的进化适应有关。     

乙肝病毒感染数量的一个数学模型

乙型肝炎是由乙型肝炎病毒(HBV)引起的肝病,该病毒干扰肝功能并造成病理损害.一小部分受感染者无法消灭该病毒而成为慢性感染,进而面临极高的死于肝硬化和肝癌的危险.乙型肝炎病毒通过与受感染者的血液或体液接触传播,这与人类免疫缺陷病毒(艾滋病毒)的方式相同.但是,乙型肝炎病毒的感染性比艾滋病毒高50至100倍.接种乙型肝炎疫苗是预防乙型肝炎的主要方法.如何有效防控乙肝的传染,不只是政府的事,也是每个国民应关注的问题.文章建立一个乙肝病毒传染的数学模型,并对模型进行实证分析;同时,对乙肝病毒的传染也做了一个预测.