依赖RNA的RNA聚合酶介导RNA干扰的扩增效应

RNA干扰(RNA interference,RNAi)是一种非常高效的基因沉默效应,RNA依赖性RNA聚合酶(RNA—de—pendent RNA polymerase,RdRP)介导的扩增作用可能是RNAi具有高效性的一个主要原因。了解RdRP在生物体中存在的证据、RdRP及其复合体的结构、次级siRNA的产生及转移性RNAi的发生机制等问题,对深入理解RNAi的作用机制和促进RNAi的临床应用有重要意义。     

RNA诱导基因沉默

RNA沉默在动植物进化中起着重要作用,具有抵抗病毒入侵、抑制转座子活动等作用,并调控蛋白编码基因的表达。该文着重对RNA沉默现象的作用机制和模式、功能及其应用前景作一综述。     

RNA空间编码探析

RNA空间结构同线性结构一样包含着重要的生物信息。RNA空间编码蕴含了RNA功能信息。RNA空间编码具有简并性、通用性、动态性、重叠性、间隔性和方向性等性质。本文对RNA空间编码的概念和性质进行了初步探讨。     

反义RNA技术

反义RNA(antisence RNA)是一种与特异mRNA互补的RNA分子,它通过配对碱基间氢键作用与对应的RNA形成双链复合物,抑制RNA的翻译过程。利用人工合成或生物体合成特定互补RNA片段(或其化学修辞产物)抑制或封闭基因表达技术,称为反义RNA技术。本文综述了反义RNA作用机理,合成途径,并展示了该技术在研究基因功能,防治肿瘤、遗传病以及人工免疫等方面广阔的应用前景。一、反义RNA抑制基因表达机理许多实验证明,原核类生物如细菌、粘菌,     

环状RNA:潜在的新型miRNA活性调控分子

环状RNA(circular RNA,circRNA)是近年来RNA领域最新的研究热点.它是一类由特殊的选择性剪切产生且在真核细胞中广泛表达的环形内源性RNA分子.研究发现,circRNA富含microRNA(miRNA)结合位点,可以发挥竞争性内源RNA作用,作为miRNA"海绵"来解除对其靶基因的抑制效应.近年来,circRNA作为一种新型调控分子调控miRNA功能的发挥,受到众多研究者的青睐.本文综述circRNA的产生机制,及其调控miRNA的最新研究进展与研究方法等.     

tsRNAs的作用机制及其在相关疾病中的潜在应用*

近年来,转运RNA(transfer RNA,tRNA)衍生的小RNA(tRNA-derived small RNA,tsRNAs)被认为是一种新的、潜在的非编码RNAs(non-coding RNA,ncRNAs)。根据在前体或成熟tRNA上切割位置的不同,tsRNAs主要被分为两种类型,即tRNA halves(tRNA-derived stress-induced RNA,tiRNAs)和tRNA衍生片段(tRNA-derived fragment,tRFs)。越来越多的证据表明,tsRNAs参与翻译起始抑制、基因沉默和调节核糖体发生等多种细胞代谢过程,并在癌症、神经退行性疾病、代谢性疾病和病毒感染等相关疾病的发生、发展中都起着重要的作用。综述tsRNAs生物学功能和作用机制及其在相关疾病中的潜在应用,总结tsRNAs研究目前存在的问题和未来的研究方向。     

猪繁殖与呼吸综合征病毒ORF2～4特异siRNA的筛选及其抑制病毒复制效果的研究

RNA干扰(RNAi)技术是基因功能研究的有效工具,为了了解猪繁殖与呼吸综合征病毒(PRRSV)次要结构蛋白GP2、GP3、GP4在病毒复制中的作用,针对各自的编码基因ORF2、ORF3、ORF4分别选取4个小干扰RNA(siRNA)位点(共12个),构建相应的短发夹RNA(shRNA)表达载体,转染MARC-145细胞后,通过荧光定量PCR和病毒滴度检测干扰效果。筛选了可以减少GP2、GP3、GP4相应基因mRNA含量的ORF2、ORF3、ORF4特异shRNA表达载体,病毒效价滴定表明shRNA表达载体处理细胞可以减少GP2、GP3、GP4相应基因mRNA含量,细胞培养上清中的病毒滴度比对照低184~4.65倍。     

miRNA研究进展

近年来,在许多真核生物中发现了一类内源性的、长度约22个核苷酸的单链非编码RNA-微RNA,研究发现它与基因表达、细胞周期及个体发育过程都有关系,因此,微RNA的研究可能对基因功能研究、人类疾病防治及生物进化探索具有重要意义。就近几年来对微RNA的研究作简要概述。     

RNA调控与作物农艺性状改良

具有重要育种价值的基因资源挖掘与功能研究对于现代农业育种应用至关重要.已有大量研究显示,核糖核酸(ribonucleic acid, RNA)在多种植物的生长发育和环境信号应答中发挥重要作用,是调控多种农作物与经济作物复杂农艺性状的重要基因资源.本文综述了小RNA和长链非编码RNA的生成代谢和功能机制,着重介绍了小RNA、长链非编码RNA及其RNA修饰在调控作物产量与品质、抗病与抗逆等方面的研究进展.同时介绍了利用RNA技术改良遗传性状的研究,并展望了RNA技术的开发与利用对于未来农业生物技术应用的重要意义.     

猪札幌病毒的研究进展

猪札幌病毒(porcine sapovirus,PoSaV)是一种经粪-口途径传播引起猪急性胃肠炎的肠道病毒,对环境友好生态型养殖业构成一定威胁。研究表明,某些PoSaV与人SaV核苷酸序列具有很高的同源性,且越来越多来源于人和猪的SaV重组新毒株被发现,提示PoSaV具有跨种间感染及传播给人的潜在风险。迄今,PoSaV的入侵与感染、变异与迁移、免疫与致病、暴发与流行、跨种间感染与传播等机制尚不清楚。本文主要对PoSaV形态与抵抗力、基因组结构与功能、基因重组、传播方式、流行病学、受体等方面的研究进展进行综述。     

Discovery and genomic characterization of a novel bat sapovirus with unusual genomic features and phylogenetic position

Sapovirus is a genus of caliciviruses that are known to cause enteric disease in humans and animals. There is considerable genetic diversity among the sapoviruses, which are classified into different genogroups based on phylogenetic analysis of the full-length capsid protein sequence. While several mammalian species, including humans, pigs, minks, and dogs, have been identified as animal hosts for sapoviruses, there were no reports of sapoviruses in bats in spite of their biological diversity. In this report, we present the results of a targeted surveillance study in different bat species in Hong Kong. Five of the 321 specimens from the bat species, Hipposideros pomona, were found to be positive for sapoviruses by RT-PCR. Complete or nearly full-length genome sequences of approximately 7.7 kb in length were obtained for three strains, which showed similar organization of the genome compared to other sapoviruses. Interestingly, they possess many genomic features atypical of most sapoviruses, like high G+C content and minimal CpG suppression. Phylogenetic analysis of the viral proteins suggested that the bat sapovirus descended from an ancestral sapovirus lineage and is most closely related to the porcine sapoviruses. Codon usage analysis showed that the bat sapovirus genome has greater codon usage bias relative to other sapovirus genomes. In summary, we report the discovery and genomic characterization of the first bat calicivirus, which appears to have evolved under different conditions after early divergence from other sapovirus lineages.     

Quantitative detection of sapoviruses in wastewater and river water in Japan

Aims:  To detect sapoviruses at a wastewater treatment plant (WWTP) and in a river in Japan, quantitatively. Methods and Results:  Influent and effluent samples at a WWTP and river water samples were collected monthly for 1 year. The water samples were subjected to virus concentration using an HA electronegative filter, followed by quantification of sapoviruses using real‐time PCR. The concentration of sapoviruses in influent ranged from 2·8 × 10³ to 1·3 × 10⁵ copies per litre, showing a higher value in winter. Seven (58%) of 12 effluent samples were positive for sapoviruses, as were 23 (64%) of 36 river water samples collected from three sites along the Tamagawa River. Conclusions:  Sapoviruses were abundant in the influent even in the nonepidemic period, suggesting that sporadic and asymptomatic infections occur throughout the year. Increasing concentration of sapoviruses was discharged into the river during the epidemic period winter. Significance and Impact of the Study:  This is the first study demonstrating the quantitative detection of sapoviruses in aquatic environments.     

Genomic characterization and high prevalence of bocaviruses in swine

Using random PCR amplification followed by plasmid subcloning and DNA sequencing, we detected bocavirus related sequences in 9 out of 17 porcine stool samples. Using primer walking, we sequenced the nearly complete genomes of two highly divergent bocaviruses we provisionally named porcine bocavirus 1 isolate H18 (PBoV1-H18) and porcine bocavirus 2 isolate A6 (PBoV2-A6) which differed by 51.8% in their NS1 protein. Phylogenetic analysis indicated that PBoV1-H18 was very closely related to a ～2 Kb central region of a porcine bocavirus-like virus (PBo-LikeV) from Sweden described in 2009. PBoV2-A6 was very closely related to the porcine bocavirus genomes PBoV-1 and PBoV2 from China described in 2010. Among 340 fecal samples collected from different age, asymptomatic swine in five Chinese provinces, the prevalence of PBoV1-H18 and PBoV2-A6 related viruses were 45-75% and 55-70% respectively, with 30-47% of pigs co-infected. PBoV1-A6 related strains were highly conserved, while PBoV2-H18 related strains were more diverse, grouping into two genotypes corresponding to the previously described PBoV1 and PBoV2. Together with the recently described partial bocavirus genomes labeled V6 and V7, a total of three major porcine bocavirus clades have therefore been described to date. Further studies will be required to elucidate the possible pathogenic impact of these diverse bocaviruses either alone or in combination with other porcine viruses.     

Identification and Characterization of a Novel Recombinant Porcine Astrovirus from Pigs in Anhui,China

Porcine astroviruses (PAstVs) have wide distribution in swine herds worldwide. At present, five porcine astrovirus genotypes have been identified. In this study, using viral metagenomics, a novel PAstV strain (designated as Ahast) was identified in fecal samples from pigs in Anhui of China, and the complete genomic sequence of Ahast was obtained by assembling and PCR amplification. Genomic structural analysis indicated that Ahast had a typical ribosomal frameshifting signal, and some conserve amino acid motifs were also found in virally encoded proteins. Phylogenetic analysis and sequence comparison indicated that this virus belonged to porcine astrovirus genotype 4 (PAstV4), which formed a clade clustered with other PAstV4. Multiple recombinant events were confirmed by recombination analysis and indicated that Ahast was a potential recombinant. Epidemiological investigation indicated that PAstV4 has a 10.7% prevalence in this pig farm. The new recombinant identified in this study will be beneficial to comprehend the origin, genetic diversity, and evolution of porcine astroviruses in Anhui of China.Key words: porcine astroviruses, viral metagenomics, genome recombination     

Long-acting contraceptive agents: Analysis and purification of steroid esters

More than 200 samples of esters of norethisterone (17α-ethynyl-17β-hydroxyestr-4-en-3-one) and levonorgestrel (13β-ethyl-17α-ethynyl-17β-hydroxygon-4-en-3-one) have been analysed by a combination of techniques, including high performance liquid chromatography (HPLC). Compounds having a purity below the required limit (99.5%) were purified, mainly by preparative HPLC, prior to formulation and biological evaluation as long-acting progestogens.     

人源札幌病毒抗原多样性及遗传进化研究进展

人源札幌病毒(human sapovirus, HuSaV)是全球范围内引起散发性急性胃肠炎和相关疫情的重要病原,尤其对婴幼儿及免疫缺陷患者等高危人群存有致死的危险,人源札幌病毒具有丰富的抗原和遗传多样性,其抗原多样性及免疫原性主要位于P2亚结构域,并且衣壳蛋白免疫原性是人源札幌病毒疫苗研发的理论基础。由于人源札幌病毒可以耐受高衣壳突变而不失去病毒功能使它得以迅速进化,其在宿主体内进化过程中存在连续的氨基酸突变,且突变主要在VP1的P结构域内积累,少见于非结构蛋白和VP2中。序列和结构的改变使得人源札幌病毒逃脱先前存在的群体免疫,有必要进一步探索人源札幌病毒的免疫逃逸机制及其拮抗宿主的免疫应答。因此,本文针对人源札幌病毒在基因组特征、抗原多样性特点、遗传进化机制等领域的研究进展进行了系统综述,并对未来研究中亟待解决的科学问题进行了展望。     

Strain-rate dependent material properties of the porcine and human kidney capsule

This study was performed to characterize the mechanical properties of the kidney capsular membrane at strain-rates associated with blunt abdominal trauma. Uniaxial quasi-static and dynamic tensile experiments were performed on fresh, unfrozen porcine and human renal capsules at deformation rates ranging from 0.0001 to 7 m/s (strain-rates of 0.005-250 s(-1)). Single stroke, dynamic tests were performed on samples of porcine renal capsule at strain-rates of 0.005 s(-1) (n = 33), 0.05 s(-1) (n = 17), 0.5 s(-1) (n = 38), 2 s(-1) (n = 10), 4 s(-1) (n = 10), 50 s(-1) (n = 21), 100 s(-1) (n = 18), 150 s(-1) (n = 17), 200 s(-1) (n = 10), and 250 s(-1) (n = 17). Due to limited availability of human tissues, only quasi-static tests were performed (0.005 s(-1), n = 25). Porcine renal capsule properties were found to match the material properties of human capsular tissue sufficiently well such that porcine tissue material can be used as a human test surrogate. The apparent elastic modulus and breaking stress of the porcine renal capsule were observed to increase significantly with increasing strain-rate (p < 0.01). Breaking strain was inversely related to strain-rate (p < 0.01). The effect of increasing strain-rate on material properties diminished appreciably at rates exceeding 150 s(-1). Empirically derived mathematical models of constitutive behavior were developed using a hyperelastic/viscoelastic Ogden formulation, as well as a Cowper-Symonds law material curve multiplication.     

Recombinant porcine norovirus identified from piglet with diarrhea

ABSTRACT: BACKGROUND: Noroviruses (NoVs) are members of the family Caliciviridae and are emerging enteric pathogens of humans and animals. Some porcine NoVs are genetically similar to human strains and are classified into GII, like most epidemic human NoVs. So far, PoNoV have been exclusively detected in fecal samples of adult pig without clinical signs. METHODS: We collected 12 fecal samples from piglets with diarrhea and no accurate diagnosis of etiology from three commercial pig farms in Shanghai suburb. We tested for PoNoV, porcine circovirus type 2, porcine rotavirus, porcine transmissible gastroenteritis virus, porcine sapovirus, and porcine epidemic diarrhea virus using RT-PCR method. The full-genome sequence of the PoNoV was then determined and analyzed. Experimental infection of miniature pigs with fecal suspensions was performed to make sure if this strain can cause gastroenteritis in piglets. RESULTS: Result showed that 2 of the 12 evaluated fecal samples were positive for PoNoVs, one of which was positive for PoNoV alone, and the other was coinfected with porcine circovirus and PoNoV. Phylogenetic and recombination analysis showed that the PoNoV positive alone strain was a recombinant new genotype strain. Experimental infection of miniature pigs with fecal suspensions confirmed that this strain can cause gastroenteritis in piglets. CONCLUSION: This is the first report that recombinant new genotype PoNoV exised in pig herd of China, which cause diarrhea in pigs in nature condition. This find raised questions about the putative epidemiologic role of PoNoV.