人血小板生成素重组载体的构建与体外表达

应用基因克隆技术,以人ＴＰＯｃＤＮＡ为目的基因,构建真核细胞表达重组体ＶＲＴＰＯ,藉脂质体将其转染于ＮＩＨ３Ｔ３细胞,应用ＰＣＲ、ＲＴ－ＰＣＲ及Ｗｅｓｔｅｒｎ印迹等技术对其转染及表达情况进行鉴定．结果表明：ＶＲＴＰＯ构建成功,在ＮＩＨ３Ｔ３细胞可表达人ＴＰＯ,为应用人ＴＰＯｃＤＮＡ进行质粒ＤＮＡ骨骼肌直接注射于动物体内的基因治疗研究奠定了基础     

Mapping of genes on the linear chromosome of the bacterium Borrelia burgdorferi: Possible locations for its origin of replication

Molecular clones of Borrelia burgdorferi, aetiologic agent of Lyme borreliosis, were isolated and analysed by DNA sequence determination. This procedure yielded B. burgdorferi homologues of gidA, gyrB, gyrA, ftsA and ftsZ. The genes were located on the physical map of the B. burgdorferi linear chromosome. Also mapped were the genes fla and p60 while dnaA was mapped using a heterologous probe. gyrA and gyrB were found to be in tandem and were mapped, along with dnaA at the centre of the chromosome. gidA was located close to the left hand extremity of the chromosome. Because gyrB, dnaA and gidA are normally located within 50 kb of the origin of replication (oriC), we propose two possible sites for oriC in the B. burgdorferi linear chromosome.     

pcDNA3.1(+)-VEGF165载体的构建及表达蛋白特性分析

构建含人VEGF165基因的重组真核表达质粒,并对其表达蛋白特性进行初步分析。将合成的VEGF165基因序列克隆至pCR2.1-TOPO载体,测序鉴定证实基因碱基序列无误后,再克隆至真核表达载体pcDNA3.1(+),构建成pcD-NA3.1(+)-VEGF165重组质粒。利用DNAstar软件分析基因序列并翻译成氨基酸序列,再用ExPASy protscale和Protean软件分析其疏水特性、蛋白二级结构。经基因测序、酶切鉴定和PCR鉴定证实pcDNA3.1(+)-VEGF165重组质粒构建成功。Ex-PASy protscale和Protean软件分析表明VEGF165蛋白具有较好的水溶性。本研究为VEGF165的基因治疗应用和VEGF165蛋白直接治疗勃起功能障碍奠定了基础。     

一种基于基因拓扑重要性的通路识别方法

癌症相关通路的识别是认识癌症发生发展过程机制的生物学基础。已有的通路识别方法很少考虑基因在通路中的拓扑重要性。重叠基因降权(PADOG)方法在基因集分析(GSA)方法的基础上融入了基因特异性的影响,提高了癌症相关通路的识别性能。为进一步提高癌症相关通路的识别性能,首先统计了KEGG通路数据集中基因出度的分布情况,根据基因出度的大小定义了基因的重要性。最后将基因的特异性和重要性融合在一起,提出了一种基于基因重要性和特异性的通路分析方法 PAGIS。在结肠癌、肺癌和胰腺癌3个数据集上的实验结果表明,PAGIS方法比PADOG能够提高很多癌症相关的排名,从而提高癌症相关通路的识别效果。     

大鼠前列腺甾体结合蛋白基因表达的种族专一性研究

以大鼠ＰＳＢＰ基因ｃＤＮＡ为探针,用Ｎｏｒｔｈｅｒｎ印迹法测定ｍＲＮＡ,Ｓｏｕｔｈｅｒｎ印迹法测定ＤＮＡ,明确了只在大鼠腹侧前列腺表达的ＰＳＢＰ基因,在其他种族,如小鼠、兔和人的前裂腺中不仅不表达,而且基因也不存在?澄清了文献中的混乱。     

Gene duplication and recombination in the evolution of mammalian Fc receptors

The immunoglobulin-related chains of cell-surface receptors for the Fc region of immunoglobulins (FCERIα, FcγRI, FcγRII, and FcγRIIIα) are encoded by members of a gene family. Phylogenetic analysis of representative members of this family from mammals revealed that FcγRIIIα genes of human, mouse, and rat are not orthologous to one another in the region of the gene encoding the Immunoglobulin C2-set domains. In phylogenetic trees of this region, FcγRIIIα and FcγRII clustered together. However, in trees based on both coding and noncoding regions 5′ and 3′ to the C2 domains, FcγRIIIα genes of human, mouse, and rat clustered together. This pattern of relationship is most easily explained as a result of two independent recombinational events occurring in the mouse and rat after these two species diverged, in each of which the exons encoding the C2 domains were donated to an FcγRIIIα gene by an FcγRII gene.     

Theory for the number of genes affecting quantitative characters

Summary A general expression for gene number estimation which encompasses the conventional formula was derived. It provides a basis for gene number estimation from the data of recurrent selection experiments that are not of sufficient duration to measure total response to selection.Gene number estimates are considerably more reliable when heritability is high. The effect of heritability on sampling variance is particularly important when gene number is large.Generally the most effective ways of decreasing the variance of a gene number estimate will be 1) to increase the number of generations in a primary selection program, 2) to increase the number of generations in the two way selection program and 3) to increase population size.From a thesis submitted by the author in partial fulfillment of the requirements for the Ph.D. degree. Received March, 1975. Work supported by Public Health Service Grant GM 16074, by the Minnesota Agricultural Experiment Station and by National Institutes of Environmental Health Sciences Grant No. 5T32 ES07011-02.Former Research Assistant, Genetics and Cell Biology, University of Minnesota; Currently Post-doctoral Fellow in Environmental Health Measurement and Statistics.     

牛β-乳球蛋白基因的克隆及其调控外源基因表达的研究

目的制备乳腺生物反应器所必需的乳腺特异性表达的调控序列,并验证其指导外源基因表达的能力.方法用PCR法从奶牛染色体上分5段扩增出了全长8.2Kb的牛β-乳球蛋白基因,包括1.8Kb的5′侧翼区、1.7Kb的3′侧翼区及4.7Kb的gDNA区.扩增出的各片段克隆到T-载体上,酶切鉴定及序列分析均证实了所扩增片段的正确性.将五个片段与荧光素酶cDNA拼接成荧光素酶瞬时表达载体并在小鼠乳腺中瞬时表达.结果注射荧光素酶瞬时表达载体的小鼠乳汁中明显测出了荧光素酶活性.结论所克隆的牛β-乳球蛋白基因表达调控序列能够指导外源基因在小鼠乳腺中表达.     

miR156靶基因SPL在植物中的系统分布和进化模式分析

Squamosa promoterbinding proteinlike genes （SPLs）在植物发育过程中具有重要作用。很多SPLs被miR156调节,然而,对于它们在植物中的系统分布和进化模式还知之甚少。本文对9个测序物种（藻类,苔藓,石松,单子叶和双子叶植物）的183个SPLs进行了生物信息学分析。结果表明miR156应答元件（MREs）仅在陆生植物SPLs中发现,藻类中不存在。系统进化分析显示陆生植物SPLs分为两大分支：group I和group II。 MiR156靶基因仅分布于group II,表明它们有着共同的祖先。Group II进一步分为7个亚支（IIaIIg）,miR156靶基因分布在除IId外的其余6个亚支的特定SPLs。系统分类与基因结构的相关性反映了SPL靶基因结构上的变化。在进化过程中,它们可能发生外显子的丢失且伴随MRE的丢失。另外,基因重复对SPL靶基因的丰度变化影响很大,尤其是被子植物与低等植物分歧后它们数量明显增加。以拟南芥为模式植物分析发现串联重复和片段重复是SPL靶基因扩张的主要机制。     

BIOREL: the benchmark resource to estimate the relevance of the gene networks

The progress of high-throughput methodologies in functional genomics has lead to the development of statistical procedures to infer gene networks from various types of high-throughput data. However, due to the lack of common standards, the biological significance of the results of the different studies is hard to compare. To overcome this problem we propose a benchmark procedure and have developed a web resource (BIOREL), which is useful for estimating the biological relevance of any genetic network by integrating different sources of biological information. The associations of each gene from the network are classified as biologically relevant or not. The proportion of genes in the network classified as "relevant" is used as the overall network relevance score. Employing synthetic data we demonstrated that such a score ranks the networks fairly in respect to the relevance level. Using BIOREL as the benchmark resource we compared the quality of experimental and theoretically predicted protein interaction data.     

GenOrigin: A comprehensive protein-coding gene origination database on the evolutionary timescale of life

The origination of new genes contributes to the biological diversity of life. New genes may quickly build their network, exert important functions, and generate novel phenotypes. Dating gene age and inferring the origination mechanisms of new genes, like primate-specific genes, is the basis for the functional study of the genes. However, no comprehensive resource of gene age estimates across species is available. Here, we systematically date the age of 9,102,113 protein-coding genes from 565 species in the Ensembl and Ensembl Genomes databases, including 82 bacteria, 57 protists, 134 fungi, 58 plants, 56 metazoa, and 178 vertebrates, using a protein-family-based pipeline with Wagner parsimony algorithm. We also collect gene age estimate data from other studies and uniformly distribute the gene age estimates to time ranges in a million years for comparison across studies. All the data are cataloged into GenOrigin (http://genorigin.chenzxlab.cn/), a user-friendly new database of gene age estimates, where users can browse gene age estimates by species, age, and gene ontology. In GenOrigin, the information such as gene age estimates, annotation, gene ontology, ortholog, and paralog, as well as detailed gene presence/absence views for gene age inference based on the species tree with evolutionary timescale, is provided to researchers for exploring gene functions.     

杨树和葡萄UBX蛋白质家族分析

UBX（泛素调控X因子）蛋白质家族在泛素化相关的过程中起着重要的作用,如细胞周期调控、转录调控、信号转导、发育、胁迫响应、细胞程序性死亡、内吞作用和DNA修复。然而,到目前为止。UBX家族在杨树和葡萄中还没有被研究过。为了更好的弄清这两个植物的UBX家族,我们对UBX的基因结构、染色体位置、基因重复、系统发育关系作了分析。该研究对葡萄和杨树的UBX蛋白质家族作了第一个系统的分析。基因的外显子／内含子结构和蛋白质基序组成在同一个组里相对比较保守。基因重复分析表明．串联重复和片段重复对于杨树和葡萄的UBX基因家族的扩张有一定贡献,基因缺失在UBX基因家族的扩张过程中也发生了作用。本研究为UBX蛋白质功能的研究奠定了基础。     

大肠杆菌亮氨酰－tRNA合成酶基因的克隆和鉴定

本文根据遗传互补原理,利用大肠杆菌亮氨酰－ｔＲＮＡ合成酶基因（ｌｅｎＳ）的温度敏感突变株ＫＬ２３１,从大肠杆菌基因文库一克隆（λ１５Ｄ７）中筛选出带完整ｌｅｕＳ基因的ＤＮＡ片段。该片段长度为３．２ｋｂ。对此片段做了１４种限制性内切酶图谱分析和部分ＤＮＡ序列鉴定,并与文献报道的ｌｅｎＳ基因序列进行了比较。发现在编码区和３’端非编码区各有一对碱基发生了转换。另外在３’端非编码区有一对碱基缺失。编码区的碱基对转换导致编码的氨基酸由组氨酸变成了精氨酸。带有ｌｅｎＳ基因的质粒（ｐＬｅｕＳ９１）转入大肠杆菌ＴＧＩ菌株中,测得转化子的亮氨酰－ｔＲＮＡ合成酶比活力是ＴＧ１菌株的１０倍以上。     

Cloning and expression of the Staphylococcus aureus glucosaminidase in Escherichia coli

Abstract A fragment of Staphylococcus aureus DNA encoding the glucosaminidase determinant was cloned in Escherichia coli by inserting the Sau 3A genomic fragments in the Bam HI site of the plasmid vector pBR322. One clone selected on the basis of its lytic activity was shown to contain a hybrid plasmid (pEU213) carrying a 4.7 kb insert of S. aureus DNA. Lytic activity was tested using different assays, and the enzyme production was confirmed by immunological reactions. An appreciable reduction of lytic activity was noted after few subcultures. The E. coli carrying pEU213 had a slower growth rate and increased autolytic activity compared to the parental strain. The possible reasons for this behavior are discussed.     

利用基因表达谱挖掘差异表达功能类的稳健性

Gene Ontology广泛地应用于基于基因芯片数据的差异表达功能类分析。基因芯片技术存在检测缺失与检测误差等问题。本文探讨上述这二个因素对利用基因表达谱挖掘Gene Ontology中差异表达功能类的影响。结果显示,差异表达功能类对于检测缺失与检测误差干扰等有一定的稳健性。     

逆转录病毒载体介导入GM-CSF基因转染肿瘤细胞

本文应用逆转录病毒载体pZIP Neo SV(X)介导人GM-CSF基因转染肿瘤细胞获得表达。经Lipofectin将含有人GM-GSF基因的重组逆转录病毒载体pZIP-GM转染兼性病毒包装细胞系PA317,继之用病毒收获液感染人肝癌细胞SMMC7721和人胃癌细胞BGC-823,经GM-CSF依赖细胞株TF1测活和双抗夹心法ELISA测定表明:人GM-CSF基因在人肿瘤细胞中获得稳定高效表达。为进一步建立GM-CSF的转基因治疗模型提供了基础。     

基因芯片的功能评价与辅助设计软件GeneHubD

为了比较特定的基因芯片的功能偏岐 ,我们研制开发了一个简单实用的基因芯片功能评价与辅助设计软件GeneHubD。通过结合GO、KEGG、BIND、MIPS、Unists数据库的基因功能注释信息 ,该软件有助于实验人员选择或设计合适的芯片开展芯片实验