巴斯德毕赤酵母表达系统在外源基因表达中的研究进展

巴斯德毕赤酵母是目前应用最广泛的外源蛋白表达系统。分别从的菌株、载体、外源基因整合、表达产物糖基化和外源基因高效表达等方面综述了毕赤酵母表达系统的研究进展。     

Hairpin DNA probe based electrochemical biosensor using methylene blue as hybridization indicator

In this paper, a label-free, rapid and simple method was proposed to study the hybridization specificity of hairpin DNA probe using methylene blue (MB) as a hybridization indicator. Thiolated hairpin DNA probe was immobilized on the gold electrode by self-assembly. The voltammetric signals of MB were investigated at these modified electrodes by means of cyclic voltammetry (CV) detection. Single-base mutation oligonucleotide and random oligonucleotide can be easily discriminated from complementary target DNA. The effect of mismatch position in target DNA was investigated. Experimental results showed that mutation in the center of target DNA had greatest effect on the hybridization with hairpin DNA probe. The relationship between electrochemical responses and DNA target concentration was also studied. The reduction current of MB intercalation decreased with increasing the concentration of target DNA. Taken together, these experiments demonstrate that the hybridization indicator MB provides great promise for rapid and specific measurement of target DNA.     

Regulatory elements in the immunoglobulin heavy chain gene 3'-enhancers induce c-myc deregulation and lymphomagenesis in murine B cells

Burkitt's lymphoma is invariably associated with chromosomal translocations that juxtapose the c-myc proto-oncogene with regulatory elements of the immunoglobulin heavy (IgH) or light chain loci resulting in the deregulation of c-myc expression. However, the enhancer elements mediating c-myc deregulation in vivo remain largely unidentified. To investigate the role of the IgH 3'-enhancers in c-myc deregulation, we used gene targeting to generate knock-in mice in which four DNase I hypersensitive regions from the murine IgH 3'-region were integrated into the 5'-region of the c-myc locus. The IgH 3'-enhancers induced the up-regulation of c-myc expression specifically in B cells of IgH-3'-E-myc mice. After approximately 10 months, the mice developed a Burkitt-like B cell lymphoma with the phenotype of B220+, IgM+, and IgD(low). Analysis of immunoglobulin gene rearrangements indicated that the lymphoma cells were of clonal origin. The presence of a rapidly expanding population of B cells in the spleen and bone marrow of young knock-in mice at 2-4 months of age was observed. Premalignant splenic B cells of knock-in mice showed higher spontaneous and induced apoptosis; however, malignant B cells were more resistant to apoptosis. The p53-ARF-Mdm2 pathway was disabled in half of the lymphomas examined, in most cases through Mdm2 overexpression. Although c-myc expression was increased in premalignant B cells, the promoter shift from P2 to P1 was observed only in malignant B cells. Our studies demonstrate that the IgH 3'-enhancers play an important role in c-myc deregulation and B cell lymphomagenesis in vivo.     

Bio‐preparation of (R)‐DMPM using whole cells of Pseudochrobactrum asaccharolyticum WZZ003 and its application on kilogram‐scale synthesis of fungicide (R)‐metalaxyl

Methyl (R)‐N‐(2,6‐dimethylphenyl)alaninate ((R)‐DMPM) is a key chiral intermediate for the production of (R)‐metalaxyl, which is one of the best‐selling fungicides. A new strain, Pseudochrobactrum asaccharolyticum WZZ003, was identified as a biocatalyst for the enantioselective hydrolysis of (R,S)‐DMPM. The key parameters including pH, temperature, rotation speed and substrate concentrations were optimized in the enantioselective hydrolysis of (R,S)‐DMPM. After the 48 h hydrolysis of 256 mM (R,S)‐DMPM under the optimized reaction conditions, the enantiomeric excess of product (e.e._p) was up to 99% and the conversion was nearly 50%. Subsequently, the unhydrolyzed (S)‐DMPM was converted to (R,S)‐DMPM through the n‐butanal‐catalyzed racemization. Furthermore, stereoselective hydrolysis of (R,S)‐DMPM catalyzed by whole cells of P. asaccharolyticum WZZ003 was scaled up to kilogram‐scale, offering (R)‐MAP‐acid with 98.6% e.e._p and 48.0% yield. Moreover, (R)‐metalaxyl was prepared at kilogram scale after subsequent esterification and coupling reactions. Therefore, a practical production process of (R)‐DMPM and (R)‐metalaxyl with the prospect of industrialization was developed in this study. © 2018 American Institute of Chemical Engineers Biotechnol. Prog., 34:921–928, 2018     

获得高抗虫转双基因烟草

利用DNA合成仪人工合成了豇豆胰蛋白酶抑制剂(cpTl)的cDNA编码全序列。合成的基因经过克隆和序列分析后．克隆到植物高效表达载体上,并转化农杆菌,通过共转化的方法,将cpTI基因和经人工改造的苏云金芽孢杆菌(B．T)δ－内毒素基因共转化烟草,得到经PCR扩增并Southern－blotting验证的分别含有CpTI和B．T基因的植株以及同时含有CpTI和B．T基因的植株。利用棉铃虫幼虫进行的杀虫测试表明．转基因烟草和对照烟草相比具有明显的杀虫活性-同时转双基因的烟草和转单一基因的烟草相比具有增强的杀虫活性。     

植物种子老化的生理学研究进展

种子品质的优劣对于农牧业生产、经济与遗传资源有效利用、生物多样性保护以及植物群落恢复与重建具有重要的作用.种子老化是其在贮藏过程中普遍存在的一种生理现象,是随着种子贮藏时间延长而发生和发展的自然不可逆过程,不仅关系到后续种、苗生长与产量、品质等问题,还对植物种质资源的保存、利用和开发等均具有重要影响.种子老化的生理机制...     

负压对机械转流管道中血流溶血过程的影响

目的：心脏转流辅助时为克服引流管道的阻力常需在血泵入口端引入负压区,但目前尚不了解负压对血细胞机械损伤过程的。本文了负压对机械转流管道中血流溶血过程的影响。方法：应用离心式血泵建立体外模拟的循环路线,循环液体对新鲜鲜血液２Ｌ,血球压积３０％,循环速度０．７Ｌ／ｍｉｎ。调节流入管道液柱高度以引入不同程度的负压区,其中Ａ组：－５ｍｍＨｇ,Ｂ组：－４５ｍｍＨｇ,Ｃ组：－９０ｍｍＨｇ,循环时间３０分钟。分     

杉木间伐材压缩密化利用的研究

以杉木间伐材为原料,通过对压缩密化处理材的物理力学性能指标测定、处理材的微观结构的变化和红外光谱分析,得到了本试验条件下的杉木间伐材压缩密化处理的优化工艺.结果表明,采用CH蒸煮添加剂软化效果好,具有环保性;最佳工艺条件为压缩率0%～60%、压缩后的厚度20 mm、压缩前的含水率0%、压缩时间20～30 min、热压温度180～200 ℃;处理材厚度吸湿回复率为2.68%,尺寸稳定性好,物理力学性能明显提高,可以与优良硬杂木(光皮桦)相媲美.处理材的微观结构只是细胞被挤压,细胞腔变小细胞壁未受到破坏.