Flow cytometric analysis of antibody producing cells using double immunofluorescent staining

Summary Double fluorescent labeling, with fluorescein isothiocyanate (FITC)-labeled F(ab)₂ specific for the heavy chain and R-phycoerythrin (R-PE)-labeled F(ab)₂ specific for the light chain, was demonstrated as a convenient means for the accurate evaluation of a heterogeneous non-antibody-producing population. Furthermore, it could be used for monitoring the changes in each immunoglobulin (Ig) chain content of the cells during the batch culture, which will facilitate the study on antibody synthesis, assembly and secretion.     

Further progress towards a catalogue of all Arabidopsis genes: analysis of a set of 5000 non-redundant ESTs

Nearly 7000 Arabidopsis thaliana -expressed sequence tags (ESTs) from 10 cDNA libraries have been sequenced, of which almost 5000 non-redundant tags have been submitted to the EMBL data bank. The quality of the cDNA libraries used is analysed. Similarity searches in international protein data banks have allowed the detection of significant similarities to a wide range of proteins from many organisms. Alignment with ESTs from the rice systematic sequencing project has allowed the detection of amino acid motifs which are conserved between the two organisms, thus identifying tags to genes encoding highly conserved proteins. These genes are candidates for a common framework in genome mapping projects in different plants.     

Poplar (Populus nigra L.) plants transformed with aBacillus thuringiensis toxin gene: insecticidal activity and genomic analysis

Insect-resistant poplar (Populus nigra L.) plants have been produced by infecting leaves withAgrobacterium tumefaciens strains carrying a binary vector containing different truncated forms of aBacillus thuringiensis (B.t.) toxin gene under a duplicated CaMV 35S promoter. Putative transgenic plants were propagated by cuttings at two experimental farms (in Beijing and Xinjiang, China). At 2–3 years after transformation, 17 of them were selected on the bases of insect-tolerance and good silvicultural traits, and evaluated for insect resistance, for the presence of theB.t. toxin DNA fragment (Southern blots and PCR) and for the expression of the transgene (western and northern blots). Somaclonal variation, as suggested by the appearance of permanent changes in the shape of the leaves, was also investigated with molecular tools (RFLP (restriction fragment length polymorphism), RAPD (random amplified polymorphic DNA) and microsatellite DNA).Bioassays withApochemia cineraius andLymantria dispar on the leaves of the selected clones showed different and, in some cases, high levels of insecticidal activity. The molecular analysis demonstrated integration and expression of the foreign gene. Somatic changes were correlated to extensive genomic changes and were quantified in dendrograms, in terms of genomic similarity. The analysis of control plants suggested that genomic changes were correlated to thein vitro culture step necessary forA. tumefaciens-mediated gene transfer, rather than to the integration of the foreign genes.Three transgenic clones (12, 153 and 192), selected for insect resistance, reduced morphological changes and promising silvicultural traits, are now under large-scale field evaluation in six different provinces in China.     

Physicochemical basis for the rapid time-action of LysB28ProB29-insulin: dissociation of a protein-ligand complex.

The rate-limiting step for the absorption of insulin solutions after subcutaneous injection is considered to be the dissociation of self-associated hexamers to monomers. To accelerate this absorption process, insulin analogues have been designed that possess full biological activity and yet have greatly diminished tendencies to self-associate. Sedimentation velocity and static light scattering results show that the presence of zinc and phenolic ligands (m-cresol and/or phenol) cause one such insulin analogue, LysB28ProB29-human insulin (LysPro), to associate into a hexameric complex. Most importantly, this ligand-bound hexamer retains its rapid-acting pharmacokinetics and pharmacodynamics. The dissociation of the stabilized hexameric analogue has been studied in vitro using static light scattering as well as in vivo using a female pig pharmacodynamic model. Retention of rapid time-action is hypothesized to be due to altered subunit packing within the hexamer. Evidence for modified monomer-monomer interactions has been observed in the X-ray crystal structure of a zinc LysPro hexamer (Ciszak E et al., 1995, Structure 3:615-622). The solution state behavior of LysPro, reported here, has been interpreted with respect to the crystal structure results. In addition, the phenolic ligand binding differences between LysPro and insulin have been compared using isothermal titrating calorimetry and visible absorption spectroscopy of cobalt-containing hexamers. These studies establish that rapid-acting insulin analogues of this type can be stabilized in solution via the formation of hexamer complexes with altered dissociation properties.     

静电作用与修饰铜锌超氧化物歧化酶的稳定性

铜锌超氧化物歧化酶(Cu, Zn-SOD)表面的赖氨酸经化学修饰后, 酶的稳定性显著提高. 赖氨酸被修饰后, 酶的电荷结构遂发生变化, 从而影响到酶分子电场. 使用FDPB方法(有限差分法求解Poission-Boltzman方程)计算了酶修饰前后的静电场变化, 以及对维持酶的结构稳定起重要作用的Cu, Zn配位结构的影响.结果表明, Cu, Zn配位体的两级离解常数在酶修饰后分别约下降10³, 10⁶.     

鼠肝抽提液中磷酸酶活性终止方法的改进

改进了一种分析磷酸酶活性的终止酶反应方法.该方法通过在酶反应进行到一定程度时,在反应混合物中加入酶反应终止液(1mol/L NaOH-0.2mol/L EDTA),从而使测定更简捷、精确.     

DNA水平上的植物系统学研究方法

本文简要总结了近年来在ＤＮＡ水平上的植物系统学研究方法,着重介绍了限制性长度多态性分析,ＰＣＲ技术在植物系统学上的应用等这一领域最新的进展,并对分子数据的分析方法及系统树的构建进行了详细讨论。     

中药秦艽根内中柱裂分的发育解剖学研究

通过发育解剖学研究表明,秦艽根的初生结构正常,初生木质部四原型。次生生长早期阶段也是正常的,但天以后的次生生长过程中,由于木质部内部分薄壁细胞的分裂,且迅速 化成异常形成层细胞,并与原维管形成层相连,从而形成多个新的形成层环,将木质部柱分为几个子木质部。     

印产毛喉鞘蕊花的引种栽培研究

本文报道印产毛喉鞘蕊花肉质根稳定性状的分离及栽培技术路线．肉质根株提纯到９９．５％;求得了露地最佳效益种植法,最优水平搭配是：３０ｄ、１５株／ｍ２、高垄、覆膜、底肥、整枝、去花穗,产鲜根８７００ｋｇ／ｈｍ２．