SARS病毒NS-1株稳定性及培养条件的研究

用SARS病毒NS-1株接种Vero细胞,37℃培养,于培养1、2、3、4d收获病毒液,分别置-70℃冻融和4℃释放,于不同时期取样进行病毒滴定。结果显示,SARS病毒NS-1株各代次培养特性和形态学变化完全相同,有较好的抗原特异性,病毒滴度稳定,4℃保存125d,滴度下降2．25lgCCID50／ml,-70℃保存6个月,滴度未见明显下降。SARS病毒NS-1株未经冻融或释放,1d收获的病毒滴度比2、3、4d收获的病毒滴度低,经冻融或释放,1、2、3、4d收获的病毒滴度无明显区别,病毒收获时的病毒滴度与形态学变化成正相关。不同接种条件收获的病毒液,病毒滴度无明显区别。SARS NS-1株有较好的遗传稳定性和保存稳定性,培养2d,4℃释放收获病毒液,细胞悬液与病毒混合接种更简单,易操作,更适合疫苗规模生产。     

漆酶工程菌株的构建及其培养条件的研究

提取产漆酶白腐菌 (Fomelignosus)的总RNA ,利用RT_PCR的方法克隆到漆酶的cDNA ,并将其克隆到表达载体pPIC9k ,重组质粒经线性化、电激转化PichiapastorisGS115、通过底物显色反应筛选到漆酶生产工程菌株。其中 2 #工程菌株产酶活力较高。培养基、培养温度、诱导用甲醇浓度及培养基中铜离子浓度对该菌产酶活力均有一定的影响。在最适条件下 (用含 2 0 0 μmol LCu2 的BMMY培养基 ,于 2 0℃ ,2 5 0r min ;用 0 5 %甲醇诱导 )培养至第 6天时 ,该菌株产酶活力达到最高 (15 10U L)。     

无血清培养高密度乳猪肝细胞的形态和功能变化

本文研究了无血清培养高密度猪肝细胞的形态和功能变化。将分离的肝细胞以高密度(1×10~7/ml)培养在含激素、多种生长因子和营养成分的无血清培养基中,动态观察培养7天中肝细胞形态、活率、蛋白质合成功能、G-6-Pase活性、安定转化功能及LDH含量;同时以无血清培养低密度(5×10~5/ml)肝细胞作为对照组。研究结果表明:高密度培养的 肝细胞各项功能较低密度培养的肝细胞为低;高密度培养的肝细胞的形态、蛋白质合成功能在培养7天中保持稳定;活率随着培养时间的延长而下降,但均高于90％;安定转化功能在培养第2、3天最强;G-6-Pase活性在培养1天后明显下降,然后维持在较低水平;LDH含量在第1、2、3天较高。     

根癌农杆菌转化紫草的研究

紫草 (ＬｉｔｈｏｓｐｅｒｍｕｍｅｒｙｔｈｒｏｒｈｉｚｏｎＳｉｅｂ .ｅｔＺｕｃｃ)是传统中药。其根部含有萘醌类化合物—紫草素及其衍生物 ,具有显著的抗菌、抗炎、抗癌以及促进伤口愈合等生理活性。紫草素同时也是一种名贵化妆品染料。科学家对紫草的研究兴趣是基于其资源的缺乏及紫草植物本身所具有的一些特点 ;如 :紫草素及其衍生物的颜色特性可凭借肉眼观察 ,紫草素及其衍生物只在紫草的根部积累 ,紫草素合成的次生代谢途径受多种酶和外界条件 (光照 ,营养等 )的调节等。紫草细胞培养 (Ｆｕｊｉｔａ等 ,1983;叶和春等 ,1991)可以产…     

氧化葡萄糖酸杆菌SCB329染色体的测定

对氧化葡萄糖酸杆菌(\%Gluconobacter oxydans\%)SCB329的纯培养进行了研究,测定了其生长曲线,确定其对数期为4～27h。获得纯培养的对数期菌体后采用凝胶包埋法制备完整染色体,用脉冲场电泳方法对SCB329的染色体进行了分析,确定其有一条染色体和一个大质粒。染色体的长度在22Mb到35Mb之间。     

响应面法优化海洋镰刀腐皮菌产出HMG-CoA还原酶抑制剂的培养条件

【目的】采用响应面法对海洋微生物镰刀腐皮菌FG319发酵产生MFS(Metabolite of Fusarium solani FG319)培养条件进行优化。【方法】在单因素试验结果的基础上,依据Box-Behnken中心组合原则设计诱导物添加量、培养时间、培养温度的3因素3水平响应面实验,以MFS产出量为响应值优化镰刀腐皮菌FG319的培养条件。【结果】镰刀腐皮菌FG319最优培养条件为诱导物添加量0.6%、培养时间7 d、培养温度22°C,在此培养条件下,MFS最高产量达到20.11 mg/L,是优化前的4倍,与理论预测的相对误差为0.64%,实测值与响应面预测值拟合良好。镰刀腐皮菌FG319代谢产物MFS在HMG-CoA还原酶和NADPH构筑的分子评价反应体系,当MFS添加到100 mg/L时,具有类似洛伐他汀抑制HMG-CoA还原酶的最大抑制率。【结论】响应面试验设计对镰刀腐皮菌FG319培养条件的优化是有效的,其次生代谢产物MFS体外抑制HMG-CoA还原酶的效果也是明显的。     

Microbial Culture Collection (MCC) and International Depositary Authority (IDA) at National Centre for Cell Science,Pune

Culture collections are valuable resources for the sustainable use of microbial diversity and its conservation. Advances in biotechnology have further increased their importance and some of these have been recognized as International Depositary Authority (IDA) for the deposition of patent cultures. Microbial Culture Collection at National Centre for Cell Science was established by the Department of Biotechnology, Government of India is country’s newest culture collection with largest holdings. It is recognized as an IDA under the Budapest Treaty and Designated National Repository under the Biodiversity Act 2002. This article describes its various service related activities.     

纳豆枯草杆菌培养滤液对家兔离体肠平滑肌的作用

目的观察纳豆枯草杆菌培养滤液（culture filtrate,CF）对家兔离体肠平滑肌的作用,并初步探讨其作用机制。方法制备兔离体回肠标本,分别记录肠平滑肌的正常收缩张力和收缩频率作为给药前对照,然后按累积剂量分别加入CF小剂量（每次0．2mL）、CF大剂量（每次0．5mL）、肉汤（每次0．5mL）,给药间隔3min,共给药8次,并描记收缩曲线。观察不同剂量cF对肠平滑肌的作用。另取肠段按毛果芸香碱、CF或阿托品、再毛果芸香碱的顺序给药,观察CF对M胆碱受体的作用。结果CF小剂量组在累积给药达1．6mL时,CF大剂量组在累积给药达3．5mL和4．0mL时,兔离体肠平滑肌收缩张力下降,与给药前比较差异有统计学意义（P〈0．05）,其余各点差异均无统计学意义（P〉0．05）。CF小剂量组在累积给药达1．2、1．4、1．6mL时,CF大剂量组在累积给药达4．0mL时,兔离体肠平滑肌收缩频率明显降低,与给药前比较差异有统计学意义（P〈0．05或P〈0．01）,其余各点差异均无统计学意义（P〉0．05）。CF或阿托品可明显对抗毛果芸香碱引起的兔离体肠平滑肌收缩张力的增加（P〈0．05或P〈0．01）,同时CF还能使其收缩频率明显减少（P〈0．01）。结论纳豆枯草杆菌CF能明显抑制兔离体肠平滑肌蠕动,其作用机制可能与阻断M胆碱受体有关。     

Selection of DNA aptamers against epidermal growth factor receptor with high affinity and specificity

Epidermal growth factor receptor (EGFR/HER1/c-ErbB1), is overexpressed in many solid cancers, such as epidermoid carcinomas, malignant gliomas, etc. EGFR plays roles in proliferation, invasion, angiogenesis and metastasis of malignant cancer cells and is the ideal antigen for clinical applications in cancer detection, imaging and therapy. Aptamers, the output of the systematic evolution of ligands by exponential enrichment (SELEX), are DNA/RNA oligonucleotides which can bind protein and other substances with specificity. RNA aptamers are undesirable due to their instability and high cost of production. Conversely, DNA aptamers have aroused researcher’s attention because they are easily synthesized, stable, selective, have high binding affinity and are cost-effective to produce. In this study, we have successfully identified DNA aptamers with high binding affinity and selectivity to EGFR. The aptamer named TuTu22 with K_d 56 ± 7.3 nM was chosen from the identified DNA aptamers for further study. Flow cytometry analysis results indicated that the TuTu22 aptamer was able to specifically recognize a variety of cancer cells expressing EGFR but did not bind to the EGFR-negative cells. With all of the aforementioned advantages, the DNA aptamers reported here against cancer biomarker EGFR will facilitate the development of novel targeted cancer detection, imaging and therapy.