一种定量检测人血清高敏C反应蛋白的化学发光免疫方法

旨在建立一种可定量检测人血清高敏CRP的化学发光检测方法 (High-sensitivity C-reactive protein quantifiable chemiluminescent immunoassay,hs-CRP CLIA)。首先利用亲和层析和离子交换层析技术从肝硬化病人腹水中纯化出高纯度的天然CRP作为免疫原制备了22株CRP单克隆抗体 (单抗),其中13株单抗在磷酸胆碱配体捕获ELISA中呈阳性,然后利用方正滴定法筛选出单抗10C5和10C11建立了hs-CRP CLIA。试剂盒评估结果显示：该方法对血清中干扰物质IgG、血红蛋白、甘油三酯等无非特异性反应;该方法检测灵敏度高,在0.04~20.38 mg/L范围内定量检测人血清CRP标准品呈良好线性关系 (R2＞0.993);该方法准确性高、可重复性好,平均回收率为99％,批内差为4.2％~5.8％,批间差为9.0％~11.5％;该方法与进口商品化高敏CRP ELISA试剂盒平行比较检测90份血清标本,结果显示两者有良好的可比性 (r=0.968)。综上,建立的hs-CRP CLIA是一种准确、可靠、可定量的高灵敏C反应蛋白检测方法,该方法的临床应用,有利于改善我国心脏病风险评估及肠炎性疾病预后判断。     

云南药用野生稻BIBAC文库混合克隆池制备及筛选

在已构建完成的云南药用野生稻BIBAC( binary bacterial artificial chomosome)文库的基础上,将文库制备成一、二、三级混合克隆池,各级混合池的数量分别为3 360、140和14个.根据Xa21抗病基因序列设计1对特异引物,利用4步PCR法对文库混合克隆池进行逐级筛选,初步确定了3个抗病基因阳性克隆.为今后以PCR法高效利用云南药用野生稻BIBAC文库挖掘其优异基因奠定了良好的基础.     

Construction of a fusion enzyme system by gene splicing as a new molecular recognition element for a sequence biosensor.

A bifunctional fusion enzyme system constructed by gene splicing is proposed as a new model to develop sequence biosensors, taking maltose biosensor as an example. The cDNA fragment of Aspergillus niger glucoamylase (E.C 3.2.1.3, GA) was fused to the 3' end of Aspergillus niger glucose oxidase (E.C 1.1.3.4, GOD) gene with the insertion of a flexible linker peptide [-(Ser-Gly)5-] coding sequence. The fusion gene was cloned into the vector pPIC9 and expressed in Pichia pastoris GS115 under the control of the AOX1 promoter. It was found that a bifunctional hybrid protein with a molecular weight of 430 kDa was secreted after induction with methanol. The fusion enzyme GOD-(Ser-Gly)5-GA (GLG) was purified using Q Sepharose Fast Flow ion-exchange chromatography. Kinetic analysis demonstrated that GLG retained the typical kinetic properties of both GA and GOD. After being immobilized on an aminosilanized glass slide through covalent bonding by glutaraldehyde, GLG showed much higher sequential catalytic efficiency than the mixture of separately expressed GA and GOD (GA/GOD). Maltose biosensors were fabricated with GLG and GA/GOD, respectively. The performance characteristics of the maltose biosensor with respect to reproducibility, signal level, and linearity were effectively improved by using the fusion enzyme. Our findings offer a basis for the development of other sequence biosensors.     

Analysis of dysregulation of immune system in pancreatic cancer based on gene expression profile

The aim of this study was to explore the dysregulated expression of the immune system in pancreatic cancer and clarify the pathogenesis of pancreatic cancer. The Dataset GSE15471 was downloaded from GEO database, Student’s t test was used to screen differentially expressed genes (DEGs) between the pancreatic cancer group and the normal control group. Kyoto Encyclopedia of Genes and Genomes (KEGG) provides functional annotation was employed to explore the significant DEGs involved in biological functions. We got 988 significantly DEGs, including 832 up-regulated genes and 156 down-regulated genes. The ratio of up-regulated genes and down-regulated genes was 5.3. Total 13 biological pathways which were significant enriched with DEGs by KEGG pathway enrichment analysis. Finally, we constructed a overall network of the immune system in pancreatic cancer with these biological pathways information. Our study reveals that dysregulated pathways in pancreatic cancer associated with the immune system. Besides, we also identify some important molecular biomarkers of the pancreatic cancer, including CXCR4 and CD4. Dysfunctional pathways and important molecular biomarkers of pancreatic cancer will provide useful information for potential treatment of pancreatic cancer.     

基因工程雄性不育烟草及其温度敏感性

将含有抗溴苯腈基因bxn和雄性不育基因的重组载体pTA29-Barnase/bxn导入农杆菌(Agrobacterium tumefaciens)后转化烟草(Nicotiana tabacum L.),得到33个转基因植株。在27℃/23℃培养的16株中,有7株表现部分不育,另9株全部可育。而在20℃/15℃培养的17株中,12株表现不育,5株表现部分不育。部分不育的植株上同时开放可育花朵和不育花朵,不过其中的不育花朵中的花粉萌发活力很低。将在20℃/15℃条件下表现不育的12个不育株从20℃/15℃温室移至27℃/23℃温室后30d左右,其中9株表现程度不同的育性恢复现象:5株表现部分可育,另4株表现完全可育;但仍有3株表现雄性不育。雄性不育花朵的花丝变短,花药皱瘪,不散粉。细胞学观察证明,转基因植株的花药绒毡层降解提早于四分体时期,至单核小孢子时期降解殆尽。     

不同碳源对须糖多孢菌生长发育及丁烯基多杀菌素生物合成的影响

本文研究了不同碳源对须糖多孢菌生长以及丁烯基多杀菌素生物合成的影响,通过寻找优势碳源优化发酵培养基配方,促进须糖多孢菌丁烯基多杀菌素的生物合成。试验共设11个处理,1个对照,通过单因素试验比较不同处理组菌体OD600值和丁烯基多杀菌素产量,筛选获得最优碳源及其发酵培养基配方。结果表明,除可溶性淀粉和木糖外,须糖多孢菌在9种碳源中都能进行生长,对不同构型碳源显示较好的利用率。在以半乳糖、葡萄糖、果糖和甘露糖作为碳源时具有较好的生长速率,而以甘露糖为碳源时能显著促进丁烯基多杀菌素的合成。选择甘露糖最佳添加浓度为5 g/L,须糖多孢菌最高菌体浓度和丁烯基多杀菌素产量分别是初始配方条件的1. 32倍和1. 78倍,显著提高了丁烯基多杀菌素的产量。上述结果为培养基碳源对丁烯基多杀菌素生物合成影响机制的研究及丁烯基多杀菌素大规模工业化发酵生产提供了科学依据和新的技术途径。     

歌德与生物学

歌德（Goethe）是德国古典文学和民族文学的杰出代表,也是世界文学史上最杰出的作家之一。青年时代,他的书信体小说《少年维特之烦恼》使他获得了巨大的成功;而他晚年的作品《浮士德》则是他一生丰富的思想总结和艺术探索的结晶,因而与荷马史诗、但丁的《神曲》和莎士比亚的《哈姆雷特》并列为欧洲文学的四大古典名著。歌德在世界文学史上的重要地位是毋庸置疑的,但是他在生物学上的重要贡献就鲜为人知了。