An antimicrobial peptide with antimicrobial activity against Helicobacter pylori

An antimicrobial peptide named odorranain-HP was identified from skin secretions of the diskless odorous frog, Odorrana grahami. It is composed of 23 amino acids with an amino acid sequence of GLLRASSVWGRKYYVDLAGCAKA. By BLAST search, odorranain-HP had si

Cloning of novel bombesin precursor cDNAs from skin of Bombina maxima

Amphibian skin is a rich resource of bioactive peptides like proline-rich bombesin from frog Bombina maxima. A novel cDNA clone encoding a precursor protein that comprises proline-rich bombesin and a novel peptide, designated as bombestatin, was isolated from a skin cDNA library of B. maxima. The predicted primary structure of the novel peptide is WEVLLNVALIRLELLSCRSSKDQDQKESCGMHSW, in which two cysteines form a disulfide bond. A BLAST search of databases did not detect sequences with significant similarity. Bombestatin possesses dose-dependent contractile activity on rat stomach strips. The differences between cDNAs encoding PR-bombesin plus bombestatin and PR-bombesin alone are due to fragment insertions located in 3'-coding region and 3'-untranslational region, respectively. (c) 2005 Elsevier B.V. All rights reserved.

A novel family of antimicrobial peptides from the skin of Amolops loloensis

While conducting experiments to investigate antimicrobial peptides of amphibians living in the Yunnan-Sichuan region of southwest China, a new family of antimicrobial peptides was identified from skin secretions of the rufous-spotted torrent frog, Amolops loloensis. Members of the new peptide family named amolopins are composed of 18 amino acids with a unique sequence, for example, NILSSIVNGINRALSFFG. By BLAST search, amolopins did no show similarity to any known peptides. Among the tested microorganisms, native and synthetic peptides only showed antimicrobial activities against Staphylococcus aureus ATCC2592 and Bacillus pumilus, no effects on other microorganisms. The CD spectroscopy showed that it adopted a structure of random combined with beta-sheet in water, Tris-HCl or Tris-HCl-SDS. Several cDNAs encoding amolopins were cloned from the skin cDNA library of A. loloensis. The precursors of amolopin are composed of 62 amino acid residues including predicted signal peptides, acidic propieces, and mature antimicrobial peptides. The preproregion of amolopin precursor comprises a hydrophobic signal peptide of 22 residues followed by an 18 residue acidic propiece which terminates by a typical prohormone processing signal Lys-Arg. The preproregions of precursors are very similar to other amphibian antimicrobial peptide precursors but the mature amolopins are different from other antimicrobial peptide families. The remarkable similarity of preproregions of precursors that give rise to very different antimicrobial peptides in distantly related frog species suggests that the corresponding genes form a multigene family originating from a common ancestor. (C) 2008 Elsevier Masson SAS. All rights reserved.

The first antimicrobial peptide from sea amphibian

The crab-eating frog, Rana cancrivora, is one of only a handful of amphibians worldwide that tolerates saline waters. It typically inhabits brackish water of mangrove forests of Southeast Asia. A large amount of antimicrobial peptides belonging to different families have been identified from skins of amphibians inhabiting freshwater. No antimicrobial peptide from sea amphibians has been reported. In this paper, we firstly reported the antimicrobial peptide and its cDNA cloning from skin secretions of the crab-eating frog R. cancrivora. The antimicrobial peptide was named cancrin with an amino acid sequence of GSAQPYKQLHKVVNWDPYG. By BLAST search, cancrin had no significant similarity to any known peptides. The cDNA encoding cancrin was cloned from the cDNA library of the skin of R. cancrivora. The cancrin precursor is composed of 68 amino acid residues including a signal peptide, acidic spacer peptide, which are similar to other antimicrobial peptide precursors from Ranid amphibians and mature cancrin. The overall structure is similar to other amphibian antimicrobial peptide precursors although mature cancrin is different from known peptides. The current results reported a new family of amphibian antimicrobial peptide and the first antimicrobial peptide from sea amphibian. (c) 2007 Elsevier Ltd. All rights reserved.

哺乳动物cd59基因的进化

采用PCR以及BLAST方法从5种哺乳动物中获得了cd59基因的编码区序列, 结合 GenBank中已有的序列, 计算cd59基因在哺乳动物中的核苷酸替换速率.对非同义替换速率和同义替换速率进行比较的结果显示, cd59在哺乳动物中总体上受到负选择作用; 用PAML软件"位点-特异"模型检测到4个受到正选择作用的位点, 4个位点分布于分子表面, 其中2个位于功能重要的区域; 此外, 用"支-位点-特异"模型在小鼠通过基因复制后形成的cd59a和cd59b上检测正选择引起的加速进化, 并检测到该支系特异的正选择位点1个.

Evolution of cd59 gene in mammals

The CD59-coding sequences were obtained from 5 mammals by PCR and BLAST, and combined with the available sequences in GenBank, the nucleotide substitution rates of mammalian cd59 were calculated. Results of synonymous and nonsynonymous substitution rates

山溪鲵皮肤β-microseminoprotein样蛋白的分离纯化及其cDNA克隆

从山溪鲵(Batrachuperus pachunii)皮肤匀浆液中经过Sephadex G-50凝胶过滤、AKTA(R)Resource Q阴离子柱和反向高压液相C4柱分离纯化得到相对分子质量为12 000的蛋白.利用其N端氨基酸序列设计引物,从山溪鲵皮肤的cDNA中克隆并筛选到该蛋白的cDNA序列.该cDNA序列的开放阅读框为339 bp,编码113个氨基酸残基组成的蛋白.在BLAST数据库搜寻比对分析表明,该蛋白的氨基酸序列与来自人类及其他哺乳动物β-microseminoprotein蛋白具有约40%的序列同源性.这也是首次在两栖类动物皮肤中确认β-microseminoprotein.初级结构分析表明,该蛋白属于亲水性蛋白;多重序列比较显示,其氨基酸序列中的10个半胱氨酸位点及15个其他氨基酸位点与高等脊椎动物β-microseminoprotein中同种氨基酸有相同的位点.由此推测该蛋白属于β-microseminoprotein家族.

XMolecular Cloning and Sequence Analyses of cDNAs Encoding Seven C2type Lectin2like Protein Subunits from Daboia russellii siamensis

按照Promega 公司的mRNA 提取试剂盒操作手册, 从圆斑蝰蛇( Daboia russellii siamensis ) 的毒腺中提 取mRNA ; 利用RT2PCR 的方法进行体外扩增, 获得C - 型凝集素蛋白的基因, 克隆到pMD182T 载体中。随机挑 选14 个阳性克隆进行核酸测序, 获得7 个编码不同蛇毒C - 型凝集素样蛋白亚基的cDNA , 分别命名为DRS2L1 、 DRS2L2 、DRS2L3 、DRS2L4 、DRS2L5 、DRS2L6 和DRS2L7 。由基因序列推导出的氨基酸序列表明, 克隆到的7 个蛇 毒C - 型凝集素样蛋白的亚基中均有糖识别结构域存在。BLAST 分析显示, 仅有DRS2L1 的蛋白序列与目前已知 的蛇毒C - 型凝集素样蛋白的α亚基相似。序列同源性比较并结合半胱氨酸位点分析, 推测DRS2L1 和DRS2L2 可能分别是圆斑蝰蛇毒Ⅹ因子激活剂的轻链LC2 和LC1 。DRS2L3 和DRS2L4 可能是高分子量的蛇毒C - 型凝集 素样蛋白的β亚基, 而DRS2L5 和DRS2L6 可能是低分子量的蛇毒C - 型凝集素样蛋白的β亚基。DRS2L7 可能是 类似于血小板膜糖蛋白Ib 结合蛋白的β亚基。

盐胁迫下杜氏盐藻(Dunaliella salina)cDNA文库的构建与新表达序列标签(EST)的获取、分析

在成功养殖杜氏盐藻(Dunaliella salina)的基础上,采用Takara cDNA文库构建试剂盒,构建盐胁迫下(1.5 mol/L NaCl)杜氏盐藻的cDNA文库.经鉴定,原始文库的滴度达1.2×106cfu/mL,重组率高达95%,且多数插入片段均在500 bp以上.对表达序列标签(expressed sequence tag,EST)分析发现,杜氏盐藻基因组中包含大量未鉴定的新基因.随机挑取60个单克隆进行序列测定,将所测序列经Blast比对等生物信息学方法分析后,发现其中三分之一,即20

彩鲫Ran基因全长cDNA的克隆及其组织表达特异性分析

根据Ran的保守区序列设计简并引物 ,结合SMARTcDNA合成及RACE PCR技术 ,克隆到彩鲫(Carassiusauratuscolorvariety)的Ran基因的cDNA全长 ,其编码区全长为 6 4 8bp ,编码 2 15个氨基酸。采用BLAST程序在NCBI数据库中对其进行同源基因搜索 ,结果表明 ,其推测的编码氨基酸序列与斑马鱼和鲑鱼的Ran基因编码的氨基酸序列的同源性分别高达 98%和 97%。还对其编码区全长序列进行了原核表达 ,以经纯化的表达蛋白免疫家兔 ,制备出了具有较高特异性

Upstream regulatory region of zebrafish lunatic fringe: Isolation and promoter analysis

Lunatic fringe (Lfng), one modulator of Notch signaling, plays an essential part in demarcation of tissues boundaries during animal early development, especially somitogenesis. To characterize the promoter of zebrafish 1fng and generate somite-specific transgenic zebrafish, we isolated the upstream regulatory region of zebrafish 1fng by blast search at the Ensembl genome database (http://www. ensembl.org) and analyzed the promoter activity using green fluorescent protein (GFP) as a reporter. Promoter activity assay in zebrafish shows that the 0.2-kb fragment containing GC-box, CAAT-box, and TATA-box can direct tissue-specific GFP expression, while the 0.4-kb and 1.2-kb fragments with further upstream sequence included drive GFP expression more efficiently. We produced 1fngEGFP-transgenic founders showing somite-specific expression of GFP and consequently generated a hemizygous 1fngEGFP-transgenic line. The eggs from 1fngEGFP-transgenic female zebrafish show strong GFP expression, which is consistent to the reverse-transcription polymerase chain reaction PCR (RT-PCR) detection of 1fng transcripts in the fertilized eggs. This reveals that zebrafish 1fng is a maternal factor existing in matured eggs, suggesting that fish somitogcnesis may be influenced by maternal factors.

Identification of three duplicated Spin genes in medaka (Oryzias latipes)

Gene and genomic duplications are very important and frequent events in fish evolution, and the divergence of duplicated genes in sequences and functions is a focus of research on gene evolution. Here, we report the identification and characterization of three duplicated Spindlin (Spin) genes from medaka (Oryzias latipes): OlSpinA, OlSpinB, and OlSpinC. Molecular cloning, genomic DNA Blast analysis and phylogenetic relationship analysis demonstrated that the three duplicated OlSpin genes should belong to gene duplication. Furthermore, Western blot analysis revealed significant expression differences of the three OlSpins among different tissues and during embryogenesis in medaka, and suggested that sequence and functional divergence might have occurred in evolution among them. © 2005 Elsevier B.V. All rights reserved.

Molecular cloning and characterisation of a fish PKR-like gene from cultured CAB cells induced by UV-inactivated virus

The double-stranded-RNA-dependent protein kinase (PKR) is an important component in an antiviral defence pathway that is mediated by interferon (IFN) in vertebrates. Previously, some important IFN system genes had been identified from an IFN-producing CAB (crucian carp Carassius auratus blastulae embryonic) cells after treatment with UV-inactivated GCHV (grass carp haemorrhage virus). Here, a fish PKR-like gene, named CaPKR-like, is cloned and sequenced from the same virally infected CAB cells. It has 2192 base pairs in length with a largest open reading frame (ORF) encoding a protein of 513 amino acid residues. BLAST search reveals that the putative CaPKR-like protein is most homologous to human PKR and also has a high-level homology with all members of a family of eIF2alpha kinases. Structurally, CaPKR-like possesses a conserved C-terminal catalytic domain of eIF2alpha kinase family and the most similarity to mammalian PKRs. Within its N-terminus, there are no dsRNA-binding domains conserved in mammalian PKRs instead of two putative Z-DNA binding domains (Zalpha). Like mammalian PKRs, CaPKR-like had a very low level of constitutive expression in normal CAB cells but was up-regulated in response to active GCHV, UV-inactivated GCHV and CAB IFN, implying that the transcriptional activation of CaPKR-like by viral infection is mediated possibly by newly produced CAB IFN, which was further supported by using cycloheximide, a potent inhibitor of protein synthesis. The results together suggested that CaPKR-like was the first identified fish gene most similar to mammalian PKRs. (C) 2004 Elsevier Ltd. All rights reserved.

INFLUENCE OF LINEAR ALKYLBENZENE SULFONATE (LAS) AS ORGANIC COSOLVENT ON LEACHING BEHAVIOR OF PCDD/FS FROM FLY-ASH AND SOIL

The leaching of polychlorinated dibenzo-p-dioxins and dibenzofurans (PCDD/Fs) was measured in soil and standard fly ash column eluted with pure water and linear alkylbenzene sulfonate (LAS)- water. The data obtained were used to evaluate the leachability of PCDD/Fs from waste dump like incineration residual slag and fly ash deposition. The leaching rate was shown to be increased significantly by using LAS water. The leachate contents of PCDD/Fs were above their known water solubility. Concentration of PCDD/Fs in the leachates as well as the relative leaching (calculated on the fly ash content) increased with increasing chlorinating degree and decreasing water solubility. LAS above the critical micelle concentration (CMC) probably enhances PCDD/Fs solubility.

Growth of aligned single-walled carbon nanotubes under ac electric fields through floating catalyst chemical vapour deposition

Through floating catalyst chemical vapour deposition(CVD) method, well-aligned isolated single-walled carbon nanotubes (SWCNTs) and their bundles were deposited on the metal electrodes patterned on the SiO2/Si surface under ac electric fields at relatively low temperature(280 degrees C). It was indicated that SWCNTs were effectively aligned under ac electric fields after they had just grown in the furnace. The time for a SWCNT to be aligned in the electric field and the effect of gas flow were estimated. Polarized Raman scattering was performed to characterize the aligned structure of SWCNTs. This method would be very useful for the controlled fabrication and preparation of SWCNTs in practical applications.