人源天然ScFv噬菌体抗体库的构建及鉴定

噬菌体抗体库技术是获得治疗性抗体的一条重要途径。以20份健康人外周血为样本,通过提取淋巴细胞、逆转录－PCR（RT PCR）、抗体可变区基因的扩增、重叠PCR获得单链抗体（ScFv）基因,将ScFv克隆入噬粒载体,通过近300次的电转化获得了库容量为1.3×109的全人源天然ScFv噬菌体抗体库。通过随机挑克隆测序和用5种不同抗原筛选对抗体库进行了初步验证。随机测序表明抗体库具有较好的多样性,用5种不同抗原对其进行筛选,均获得了特异性噬菌体抗体的不同富集,表明成功构建了一个多样性良好的人源天然ScFv噬菌体抗体库。     

克氏原螯虾嗜水气单胞菌噬菌体的分离鉴定和应用

为分离获得致病性嗜水气单胞菌烈性噬菌体,鉴定和研究其生物学特性,为克氏原螯虾细菌性疾病的防治和治疗提供依据,研究以患病克氏原螯虾分离到的致病性嗜水气单胞菌为宿主菌,从湖泊水体中分离到多株噬菌体,对其中一株烈性噬菌体Ph-0进行了生物学特性的分析,包括噬菌斑形态的观察,最佳感染复数的测定,一步生长曲线的绘制,热稳定性的测...     

表面活性剂对林可霉素发酵生产的影响

在林可霉素发酵过程中,当向培养基中加入表面活性剂十二烷基磺酸钠(SDS)、吐温80(Tween 80)和曲拉通(Triton X-100)时,林可霉素的产量受到较大影响。本研究应用响应面设计法(Response surface design)对表面活性剂的配比进行了优化,得到的优化配比为:十二烷基磺酸钠为31.13 mg/100 mL,吐温80为51.97 mg/100 mL,曲拉通为16.9 mg/100 mL。将该优化配比应用于林可霉素发酵,产量提高了36.67%。     

一株南海红树林底泥抗真菌放线菌(No.H74-18)的化学成分研究

对我国南海红树林底泥中分离的一株放线菌(No.H74-18)的发酵菌丝体采用95%乙醇提取,并对具有抗真菌活性的乙酸乙酯部位进行研究,通过硅胶开放柱色谱分离得到了一个结晶样品。通过制备型反相高效液相色谱分离,从此结晶样品中分离纯化出3个化合物,经NMR、MS等光谱学方法分别鉴定为抗霉素A1(1)、抗霉素A2a(2)、抗霉素A3(3),这些化合物都是抗霉素类化合物。采用LC-MS联用技术分析了此结晶样品中存在的抗霉素类的可能组份。     

人源抗狂犬病毒单克隆抗体Fab段基因的获得和表达

运用噬菌体表面呈现（phage display）技术获得了人源抗狂犬病毒糖蛋白基因工程单克隆抗体Fab段基因及其表达。从狂犬病毒PM株Vero细胞疫苗免疫的人抗凝血中分离获得外周淋巴细胞,提取细胞总RNA,通过RTPCR方法,用一组人IgG Fab基因4特异性引物,从合成的cDNA中扩增了一组轻链和重链Fab段基因,将轻链和重链Fab段基因,将轻链和重链先后克隆入噬菌体载体pComb3,成功地建立了抗狂犬病毒抗原的方法,对此抗体库进行富积筛选表达,成功地获得了抗狂犬病毒的人源单抗Fab段基因及其在大肠杆菌中的有效表达,对其中一株单抗G10进行了较为系统的分析,发现它与一株鼠源中和性狂犬病毒糖蛋白特异性单抗存在竞争,证实该单抗能识别狂犬病毒糖蛋白,其序列资料分析表明,该单抗为一株新的抗狂犬病毒人源基因工程抗体。     

国家人勇担国家责，积极推动中国生物经济与生物产业发展

作为国家级研究机构,中国科学院各级研究所一直以来的重要使命就是心系国家事、肩扛国家责、永做国家人,站在国际科学发展前沿的高度,紧抓国家社会发展中重大需求的科学与技术问题,布局研究所及其项目和吸引人才。本文以个人视角,回顾了2004年回国接任微生物研究所所长后,开始思考工业生物技术的发展,布局天津研发中心,经过多年努力,在院党组的领导与支持下,中国科学院天津工业生物技术研究所建成,已经取得了重要成果;还回顾了建所初衷和早期成长发展历程以及与微生物研究所的关系。通过思考科学、技术、工程(医学)之间的关系,提出从事基础科学研究的科研人员进行转化应用的路径,即“想法-假说-实验-概念-论文-技术-样品-产品-商品”的9层逻辑关系,希望科学家能够做出解决实际问题的实用工作,并展望了未来工业生物技术的远景。     

Transmembrane diffusion of hydrophobic antimicrobial agents and cell surface hydrophobicity in Bacteroides fragilis

The transmembrane diffusion of hydrophobic antimicrobial agents, e.g. lincomycin and clindamycin, was examined in Bacteroides fragilis which is sensitive to these agents. The results showed that these agents penetrate efficiently through the outer membrane. Cell surface hydrophobicity measured by the partition assay between water and p-xylene revealed that the cell surface of B. fragilis is more hydrophobic than that of Salmonella typhimurium or Pseudomonas aeruginosa. Furthermore, treatment with low concentrations of surfactant caused cell lysis. These results suggest that the cell surface hydrophobicity in B. fragilis plays an important role in the efficient transmembrane penetration of hydrophobic compounds. This efficiency explains the susceptibility of B. fragilis to hydrophobic antimicrobial agents.     

Fertilization and wounding of the style induce the expression of a highly conserved plant gene homologous to a Plasmodium falciparum surface antigen in the wild potato Solanum chacoense Bitt.

Pistil tissues are actively involved in pollen tube growth and respond to the presence of the growing pollen tubes by modulating the expression of specific genes. Once fertilization has occurred, complex developmental programs lead to embryogenesis, ovary maturation, and seed set. In order to understand the early events that follow pollination and fertilization we have used a subtractive hybridization approach to characterize genes which are related to pollination and fertilization events. One cDNA clone isolated and named SPP30 (Solanum pollinated pistil) was found to share significant sequence identities with a Plasmodium falciparum (malaria parasite) surface antigen and a yeast gene of unknown function. Searches in recent EST databases also revealed that SPP30 homologues are found in both monocot and dicot species. The presence of this conserved gene in evolutionarily distant organisms such as yeast, Plasmodium, and plants suggests that it codes for an essential cellular function. This is also strengthened by its extremely high sequence conservation in both monocots and dicots where virtually all substitutions tolerated are conservative.     

pH-dependent conformational changes of ferricytochrome c induced by electrode surface microstructure

pH-dependent processes of bovine heart ferricytochrome c have been investigated by electronic absorption and circular dichroism (CD) spectra at functionalized single-wall carbon nanotubes (SWNTs) modified glass carbon electrode (SWNTs/GCE) using a long optical path thin layer cell. These methods enabled the pH-dependent conformational changes arising from the heme structure change to be monitored. The spectra obtained at functionalized SWNTs/GCE reflect electrode surface microstructure-dependent changes for pH-induced protein conformation, pK_a of alkaline transition and structural microenvironment of the ferricytochrome c heme. pH-dependent conformational distribution curves of ferricytochrome c obtained by analysis of in situ CD spectra using singular value decomposition least square (SVDLS) method show that the functionalized SWNTs can retain native conformational stability of ferricytochrome c during alkaline transition.     

Molecular cloning of protamine-2 and expression with aging in Japanese monkey (<Emphasis Type="Italic">Macaca fuscata</Emphasis>)

In order to isolate genes relating spermatogenesis with postnatal development and aging, we have attempted to obtain genes showing differences in expression in testis of Japanese monkeys (Macaca fuscata) by means of differential display PCR, and we have cloned, sequenced and characterized protamine-2 (PRM2) of Japanese monkey. The predicted open reading frame encoded a protein of 103 amino acid residues, most of which are common to those of other mammals. Northern analysis revealed that the PRM2 gene is expressed at adult and aged stages, but not at the juvenile stage. In situ hybridization revealed that the PRM2 gene is expressed mainly in late spermatids and its expressional potential is decreased from adult to aged stages. It suggests that PRM2 in spermiogenesis is mediated by the age of the animal.