红豆杉组织培养中褐变问题的研究

本研究针对红豆杉组织培养中经常遇到的褐变问题,采取多种方法进行试验,力求最大限度地避免或减轻褐变。结果表明,选择合适的外植体、培养基和培养条件是至关重要的,此外,抗氧化剂、金属离子螫合剂、热激处理和活性碳处理等方法可在一定程度上减轻褐变的程度。综合考虑各种因素,经过筛选可以获得生长良好的不发生褐变的组织。     

Scalable GMP compliant suspension culture system for human ES cells

Suspension bioreactors are an attractive alternative to static culture of human embryonic stem cells (hESCs) for the generation of clinically relevant cell numbers in a controlled system. In this study, we have developed a scalable suspension culture system using serum-free defined media with spinner flasks for hESC expansion as cell aggregates. With optimized cell seeding density and splitting interval, we demonstrate prolonged passaging and expansion of several hESC lines with overall expansion, yield, viability and maintenance of pluripotency equivalent to adherent culture. Human ESCs maintained in suspension as aggregates can be passaged at least 20 times to achieve over 1×10(13) fold calculated expansion with high undifferentiation rate and normal karyotype. Furthermore, the aggregates are able to differentiate to cardiomyocytes in a directed fashion. Finally, we show that the cells can be cryopreserved in serum-free medium and thawed into adherent or suspension cultures to continue passaging and expansion. We have successfully used this method under cGMP or cGMP-equivalent conditions to generate cell banks of several hESC lines. Taken together, our suspension culture system provides a powerful approach for scale-up expansion of hESCs under defined and serum-free conditions for clinical and research applications.     

叶绿体基因组在系统发育学及基因工程领域的应用

介绍了叶绿体基因组在系统发育学和基因工程这两个领域的应用研究进展：1)叶绿体基因组的DNA序列比较为植物系统发育学研究提供了可靠数据基础;2)叶绿体基因工程是高水平表达外源基因的重要途径之一,在生产医用蛋白、改良作物农艺性状和环境保护等方面有着广阔的应用前景。     

大数据时代杂合酶的设计及其新趋势

酶分子的高效性和稳定性是工业广泛应用的物质基础。利用分子生物学技术可以将不同酶分子通过串联、插入、翻译后融合等方式构建成符合工业需求的杂合酶,但应用中多结构域杂合酶在表达量与酶活等方面仍存在弊端,而基于特定蛋白质结构域的多功能设计成为新趋势。高通量测序技术的发展,使得生物学家正面临着爆炸式增长的大数据集。近年来"蛋白质功能区"概念的提出,拓宽了人们对蛋白质结构与功能组织层次的认知,功能区残基聚簇的协同演化可导致同一家族不同蛋白质功能的差异。基于海量大数据分析可以快速定位特定功能区以及协同进化的关键位点,再利用合成生物学技术就可实现多种功能残基在同一蛋白质中的精准嫁接,完成天然酶分子的再设计。这将是杂合酶技术发展的新阶段,也会成为生物大数据时代下蛋白质设计的新趋势。     

抑制O~6-甲基鸟嘌呤-DNA甲基转移酶活性与肿瘤细胞对亚硝脲药物敏感性关系的研究

80％以上的肿瘤细胞为O~6-甲基鸟嘌吟-DNA甲基转移酶(O~6-MT)活性较高的Mer~+型,能够修复亚硝脲药物(NU)造成的DNA烷化损伤,对NU不敏感。本实验证明,用0.75,0.50和0.25mmol/L甲基亚硝脲(MNU)分别处理Mer~+型的HeLaS3,SMMC-7721和表现Mer~-型特征的Cc801,均能明显降低细胞中O~6-MT活性,从而显著提高了三种细胞对嘧啶亚硝脲和双氯乙亚硝脲的敏感性,提示降低O~6-MT活性是使用NU对Mer~+型肿瘤进行有效治疗的前提。     

辛伐他汀对动脉粥样硬化大鼠凋亡相关基因Fas及FasL蛋白表达的影响

目的通过研究辛伐他汀对动脉粥样硬化大鼠血管壁中细胞凋亡相关基因Fas及FasL蛋白表达产物的影响,探讨其在预防动脉粥样硬化发生中的可能机制。方法复制动脉粥样硬化大鼠模型,以辛伐他汀干预,取胸主动脉,观察其斑块变化,采用免疫组化Elivision法测定动脉粥样硬化血管壁中Fas、FasL蛋白表达。结果Fas蛋白表达在实验组明显高于对照组及干预组(P<0.01,P<0.05),实验组FasL蛋白表达也明显高于对照组及干预组(P<0.05)。结论Fas及FasL基因通过促进细胞凋亡作用而诱发动脉粥样硬化过程,辛伐他汀可通过调节细胞凋亡过程发挥抗动脉粥样硬化作用。     

Refolding and purification of interferon-gamma in industry by hydrophobic interaction chromatography

A new technology for renaturation with simultaneous purification of the recombinant human interferon-gamma (rhIFN-gamma) in downstream of biotechnology is presented. The strategies to develop the new technology in industry scale were suggested. Based on chemical equilibrium and molecular interactions, the principle of rhIFN-gamma refolding by HPHIC was described. The kind of stationary and mobile phases were evaluated and found the former to contribute to the rhIFN-gamma refolding more than the latter. The extract containing the rhIFN-gamma in gram scale in 7.0 mol L(-1) guanidine hydrochloride solution of 700 mL was directly pumped into a unit of simultaneous renaturation and purification of proteins (USRPP, 10 x 300 mm i.d.) packed by small particle packings of hydrophobic interaction chromatography and a satisfactory recovery of bioactivity and mass of the rhIFN-gamma was obtained. With flow rate 100 mL min(-1) and a gradient elution by only one step in 4h, the purity and specific bioactivity approach to 95% and 8.7 x 10(7) IU(-1) mg, respectively. To evaluate the goodness of the presented new technology in this study, a usual method with the renaturation by dilution method firstly and then purification with a series of LC in literature was employed to compare with each other. The obtained result in terms of purity, recoveries of mass and bioactivity, cost time as well as expenses, the former is much better than the latter. Comparing the total bioactivity of rhIFN-gamma in the extract before to that after the renaturation by the USRPP, the total bioactivity of rhIFN-gamma increased 62-fold.     

低氧预处理对低氧／复氧心肌能量代谢的作用

目的：研究低氧预处理（HPC）对心肌的保护作用,方法：借助^31P－NMR图谱技术,在模拟Langendorff离体灌流大鼠心脏的正常生理条件下,跟踪心肌高能磷酸化合物含量的动态变化。结果：在30min低氧期,PCr、ATP相对含量及PCr/Pi值逐渐减小,但HPC组减小的速度比对照组慢;而在复氧期,HPC组能提高心肌高能磷酸化合物含量的恢复程度,特别是复氧初期,HPC组PCr 、ATP相对含量及PCr/Pi值立即有了恢复;在本实验中,HPC对pHi的改善不显著。结论：HPC能降低后续长时间低氧及复氧阶段的心肌能量代谢,对心肌的低氧／复氧损伤具有保护作用。     

水稻淀粉合成的分子生物学研究进展

本文综述了水稻淀粉合成的分子生物学研究的最新进展,主要内容是参与淀粉合成的酶鉴定及其基因表达调控,也介绍了对这些酶的遗传操作改良稻米淀粉品质等内容