基于SLAF-seq的天竺桂群体遗传变异分析

【目的】探讨我国天竺桂资源的遗传多样性及群体的空间分布格局。【方法】分别对来自7个天然群体的30份天竺桂资源进行特异位点扩增片段测序(specific-locus amplified fragment sequencing, SLAF-seq)。基于检测到的SNP位点信息进行遗传变异分析。【结果】各样品的平均测序深度为15.11倍,开发获得 1 296 000个SLAF标签,其中377 250个SLAF标签在不同样品间具有多态性,共包含3 409 402个群体SNP,经过滤,最终获得268 821个高度一致性的群体SNP。基于这些SNP的系统进化分析发现,天竺桂是由中国东部向中国西部逐渐进化的。系统进化、主成分分析和群体结构分析均表明,来自5省(直辖市)7个天然群体的群体内变异小于群体间变异。30份天竺桂资源可分为2个大的亚群,其中,第1亚群位于中国第2阶梯上,第2亚群位于中国第3阶梯上,2个亚群间被大兴安岭—太行山脉—巫山—雪峰山山脉阻隔。第2亚群又可进一步分为3个小亚群,其中,来自浙江省和安徽桃岭的为第1个小亚群,来自安徽霍山的为第2个小亚群,而来自河南伏牛山的为第3个小亚群,第1个和第2个小亚群间被长江阻隔。【结论】山脉和湖泊的阻隔可能是造成天竺桂遗传分化的重要因素。本研究首次揭示天竺桂遗传结构及地理变化规律,为我国天竺桂资源的有效利用与科学保护提供理论依据。     

H2株甲肝减毒活疫苗K7的核苷酸全序列分析

应用ＲＴ－ＰＣＲ获取Ｈ２株甲肝减毒活疫苗（Ｋ７）的ｃＤＮＡ片段，经末端终止测序ＰＣＲ试剂盒和全自动测序仪（ＡＢＩ３７７）作序列测定，用Ｍａｘｔｒｉｇｅｎｅ软件进行计算机分析，Ｋ７疫苗病毒的核苷酸全长含７４４３个碱基．Ｋ７对比其亲代Ｈ２株，减毒的ＨＭ１７５株以及ＨＭ１７５野毒株，同源性分别为９９．７５％，９９．９５％和９９．６１％；彼此间在Ｐ１和Ｐ２连接区１６８个碱基的同源性为９９．４％～１００％，均为ⅠＢ基因亚型．前三者在５’非编码区均有１３１～１３４位和２０３位５个碱基缺失，且结构蛋白基因区序列完全一致．本文用分子病毒学方法证实了Ｋ７疫苗的优异纯毒水平，也为开展甲型肝炎分子流行病学工作提供了基础资料     

Thymidine kinase gene mutation leads to reduced virulence of pseudorabies virus

To explore correlation between the tk gene structure of pseudorabies virus (PRV) and its virulence, to study the effect of the gene mutation on PRV biological properties, and to investigate mechinism of reduced virulence, thymidine kinase (TK)-deficient mutant of pseudorabies virus strain Hubei (PRV HB) was isolated by selection for resistance to 5-bromodeoxyuridine. The tk genes of PRV HB and its TK^― mutant were cloned and sequenced. 1587 base pairs of the tk gene and flanking regions of wild-type (wt) virus were sequenced, which included an open reading frame (ORF) of 1098 bp encoding a protein of 366 amino acids. The ORF contained two 137-bp repeated sequences, which were connected by an adenosine. 1458 bp of the tk and flanking regions of TK^― mutant were sequenced. Analysis of the tk gene sequence of TK^― mutant indicated that one of 137 bp repeated sequence and the connecting adenosine in the tk gene of the wt virus was deleted and a repeated sequence of 8 nucleotides (GCGCGCC) was inserted. All other nucleotides of TK^―mutant were identical to that of wt virus. Deletion and insertion of the nucleotide sequence resulted in a frameshift and a premature chain termination, and the resultant TK protein was not active. Analysis of the amino acid sequence revealed that TK protein of PRV HB contained the conserved consensus sequence of herpesviral TKs and an additional conserved-DHR-motif. The results of this work also indicated that TK^― mutant was genetically stable. Compared to PRV HB, virulence of TK^― mutant was greatly decreased. Mice vaccinated with TK^― mutant were completely protected against a lethal challenge with virulent PRV (HB).     

Presence of NAD pyrophosphorylase in skeletal muscle in dystrophic mice

Summary NAD pyrophosphorylase (ATP:NMN adenylyltransferase) activity has been measured in the skeletal muscle of dystrophic mice. The amount of this enzyme in the dystrophic mice, as determined by three different methods, was about one half of that in the controls. In addition, the concentration of ATP was too low to be detected in crude extracts of dystrophic mouse skeletal muscle, which were prepared using Tris buffer alone or Tris buffer containing either 3 M KCl, or 1 mM PMSF.     

种群间遗传距离与进化趋异

遗传距离是衡量种群间遗传分化程度和差异大小的数量指标。近年来,不仅在生物进化与物种形成方面,而且在分类学、生态学、人类学等很多领域都有广泛的应用。同时,在人类遗传研究中也受到越来越普遍的注意。本文阐述遗传距离的概念、估算原理及其应用,并将其与蛋白质变异有关的等位基因差异联系起来。在此基础上讨论最小遗传距离D_(min)、标准距离D和最大距离D_(max)的概念与公式。并结合实例进行分析与释义。     

Nucleotide analogues as probes for DNA polymerases

Transmission of the genetic information from the parental DNA strand to the offspring is crucial for the survival of any living species. In nature, all DNA synthesis in DNA replication, recombination and repair is catalyzed by DNA polymerases and depends on their ability to select the canonical nucleobase pair from a pool of structurally similar building blocks. Recently, a wealth of valuable new insights into DNA polymerase mechanisms have been gained through application of carefully designed synthetic nucleotides and oligonucleotides in functional enzyme studies. The applied analogues exhibit features that differ in certain aspects from their natural counterparts and, thus, allow investigation of the involvement and efficacy of a chosen particular aspect on the entire complex enzyme mechanism. This review will focus on a depiction of the efforts that have been undertaken towards the development of nucleotide analogues with carefully altered properties. The different approaches will be discussed in the context of the motivation and the problem under investigation.Received 16 March 2005; received after revision 5 May 2005; accepted 8 June 2005     

Nucleotide binding domains of human CFTR: a structural classification of critical residues and disease-causing mutations.

Defective function of the cystic fibrosis (CF) transmembrane conductance regulator (CFTR) causes CF, the most frequent lethal inherited disease among the Caucasian population. The structure of this chloride ion channel includes two nucleotide-binding domains (NBDs), whose ATPase activity controls channel gating. Recently, the experimental structures of mouse and human CFTR NBD1 and our model of the human CFTR NBD1/NBD2 heterodimer have provided new insights into specific structural features of the CFTR NBD dimer. In the present work, we provide a structural classification of CF-causing mutations which may complement the existing functional classification. Our analysis also identified amino acid residues which may play a critical role in interdomain interaction and are located at the NBD1-NBD2 interface or on the surface of the dimer. In particular, a cluster of aromatic amino acids, which includes F508 and straddles the two NBDs, might be directly involved in the interaction of the NBD1/NBD2 heterodimer with the channel-forming membrane-spanning domains.Received 24 May 2005; received after revision 13 June 2005; accepted 18 June 2005     

Sr2+ can become incorporated into an agonist-sensitive,cytoplasmic Ca2+ store in a cell line derived from the equine sweat gland epithelium

We have explored the properties of a Ca²⁺-dependent cell-signalling pathway that becomes active when cultured equine sweat gland cells are stimulated with ATP. The ATP-regulated, Ca²⁺-influx pathway allowed Sr²⁺ to enter the cytoplasm but permitted only a minimal influx of Ba²⁺. Experiments in which cells were repeatedly stimulated with ATP suggested that Sr²⁺, but not Ba²⁺, could become incorporated into the agonist-sensitive, cytoplasmic Ca²⁺ store. Further evidence for this was provided by experiments using ionomycin, a Ca²⁺ ionophore which has no affinity for Sr²⁺.     

核酸水解酶及酶解法生产核苷酸研究进展

综述了目前主要核酸水解酶的种类、来源、蛋白质特性和生产方法;介绍了目前酶解法生产核苷酸的几种主要方法,比较了各种方法的优缺点及工业化应用前景;简单综述了水解液中四种核苷酸的分离提取方法;探讨了酶解法生产核苷酸的未来研究方向.结论为酶解法的关键是必须通过广泛采用现代高新生物技术来降低生产成本.     

强直性脊柱炎患者中IL-23 R基因rs7517847多态性分析

目的研究IL-23R基因rs7517847位点的单核苷酸多态性与强直性脊柱炎易感性的关系.方法采用PCRRFLP方法对100例强直性脊柱炎患者进行IL-23R基因多态性检测,并与35例健康者对照分析.结果 rs7517847位点各基因型频率和等位基因频率在AS组与对照组之间的分布差异均具有统计学意义(P0.05);并在假设遗传方式下,rs7517847位点的纯合突变GG基因型与(TG+TT)基因型比较,其频率分布差异在AS组与对照组之间也具有统计学意义(P0.05).结论 IL-23R基因rs7517847位点的多态性与吉林地区人群AS易感性有关;携带G等位基因且为GG基因型的个体患AS的危险性增大,这可能是患AS的易感因素之一.