纯化

Methods:(1) Silica column chromatography and LH-20 were used in phytochemistry study.(2) Based on the extraction ratio of arctiin, the best extraction procedure was acheived under the direction of L9(34) orthogonal experimental design. From 10 types of resins, the one with the maximal binding ability for Arctii was selected and silica-column choramatograpy was used in its seperation and purification procedure.(3) The animal diabetic experimental model in Wistar rats was established with streptozotocin and many biochemical factors were used to indicate the pharmacological effects of arctiin, including glycosylated hemoglobin, serum creatinine, seralbumin, monoxide nitrogen, glycerin trilaurate, cholesterol, HDL-ch, LDL-ch, Blood urea nitrogen, creatinine, SOD, ET, MDA, total Urine protein and albuminuria.

(1)采用硅胶柱层析、聚酰胺层析、LH-20等方法对牛蒡子的化学成分进行研究；(2)采用正交实验设计方法，以牛蒡子苷为指标，确定了牛蒡子乙醇回流提取的最佳工艺；对10种类型大孔吸附树脂进行筛选，筛选出对牛蒡子苷吸附效果最佳的树脂；采用硅胶柱层析法，对牛蒡子苷进一步分离纯化；(3)建立实验性糖尿病大鼠模型，以GLU、TC、TG、HDL-ch、LDL-ch、TB、ALB、BUN、CREA、GHbAlc、NO、SOD、ET、MDA和尿液中TB、ALB为指标，研究牛蒡子苷药理活性；(4)以差速消化法纯化Wistar大鼠的主动脉内皮细胞并进行传代培养，测定牛蒡子苷对高糖条件下RAECs活性、RAECs释放LDH、MDA和NO的变化以及RAECs表达内皮型一氧化氮合酶表达的影响。

Chicken IL-15(ChIL-15), Which was discovered in 2001, plays a main role in activating NK cells and CD8+ memory T cells, and may therefore have potential as a vaccine adjuvant.According to published ChIL-15 gene sequence, a pair of primers were designed and synthesized to clone ChIL-15 cDNA from ConA-activated chicken splenocytes by RT-PCR. Recombinant plasmid pUC19ChIL-15 carrying the ChIL-15 gene was constructed. The gene encoding mature ChIL-15 (mChIL-15) was amplified from pUC19ChIL-15 by PCR and cloned into pMD 18-T vector. After digested with Kpn I /Pst I , the mChIL-15 gene was subcloned into prokaryotic expression vector pPRoEX橦Ta and transformed into E.coli DH5a .The transformant was induced by IPTG, and the recombinant ChIL-15(rChIL-15) was expressed as a fusion protein with a histidine hexamer tag at the N-terminal end of the protein. Following expression, the protein was purified by ProBond?Nickel-Chelating Resin. The uninduced and induced protein lysates and the purified protein were separated by SDS-PAGE and transferred onto nitrocellulose membrane to identify rChIL-15 by Western blot. The rChIL-15 was used to immune the guinea pig to obtain rChIL-15 polyclonal antibody.

参考已发表的ChIL-15基因序列，设计合成引物，应用RT-PCR技术从刀豆球蛋白A活化的白来航鸡脾淋巴细胞中克隆ChIL-15 cDNA，将其与SmalⅠ酶切处理的pUC19载体连接，构建重组质粒pUC19ChIL-15；用PCR技术从pUC19ChIL-15中扩增出去信号肽的成熟ChIL-15(mChIL-15)基因，与pMD 18-T载体相连构建重组质粒pMDChIL-15；然后用KpnⅠ／PstⅠ双酶切下mChIL-15基因片段，定向亚克隆到经同样酶切处理的带有6个组氨酸标签的表达载体pP_RoEX~HTa中，构建重组质粒pP_RoChIL-15，对其测序确认读框正确后，转入大肠杆菌DH5α中诱导表达并进行纯化，用重组ChIL-15(rChIL-15)免疫豚鼠，制备多克隆抗体；再用HindⅢ／PstⅠ从质粒pP_RoChIL-15上切下mChIL-15基因，定向亚克隆到经同样酶切处理的杆状病毒转移载体pMelBacB中，经酶切、PCR和序列测定鉴定后，与杆状病毒线形化DNABac-N-Blue~(TM DNA共转染Sf9昆虫细胞，经三轮蚀斑纯化，构建重组杆状病毒rBac-ChIL-15，用该重组病毒感染处于对数生长期的Sf9细胞，按不同的时间收取样品，离心后对其上清和沉淀进行SDS-PAGE和Western blot分析。

Methods The method of sucrose gradient centrifugation was used to purify Human herpesvirus-6B(HHV-6B); and in order to make sure the range of positive samples collection, the viron was detected by the methods of PCR, Dot blotting, Western blotting.

采用蔗糖密度梯度离心法进行病毒纯化；通过PCR法、斑点杂交法、蛋白免疫印迹法对纯化样品中HHV-6B进行鉴定，以确定收集阳性样品的范围。

The iso-electric point of was 4.8-5.2. The amino acid analysis indicated that the recombinant protein is non- N-glycosylated.The ability of 0.5 ug/m rhsTRAIL to induce apoptosis of liver cancer cells SMMC7721 and BEL 7402 could be visioned morphologically. This was further verified by Electrical microscope observision of induced SMMC7721 cells and agarose gel electrophoresis of apoptosis DNA. MTT test results also showed that 0.01 u g/ml purified rhsTRAIL has strong effect to induce apoptosis of SMMC7721 cell.

形态学上观察了rhsTRAIL对肝癌细胞SMMC7721和BEL7402的诱导凋亡活性，发现rhsTRAIL在0.5μg/ml时显示了很好的诱导凋亡活性，并通过电镜观察rhsTRAIL诱导后的SMMC7721细胞，证实了凋亡；同时提取凋亡细胞DNA，进行琼脂糖电泳分析，在分子水平证明凋亡；最后利用SMMC7721细胞分组加入纯化后的rhsTRAIL蛋白，结果显示纯化蛋白含量在0.01μg/ml时细胞就呈现了诱导凋亡活性。

Two molecular forms of Xenopus hatching enzyme, 60kD and 40kD molecules, was obtained during preparation and purification. Both of them were verified as the hatching enzyme molecules, using anti GST UVS.2 antibody as the probe. 60kD molecule was digested or autodigested easily into 40kD molecule during purification. It was indicated that 40kD molecule was probably derived from 60kD molecule with its two CUB repeats lost, and the two CUB repeats may play an important role in recognizing and/or modifying of the molecular structure of vitelline envelope.

在分离纯化非洲爪蟾孵化酶时，得到了60kD和40kD两种分子，用孵化酶的特异性抗GST-UVS.2抗体进行Western杂交的结果证明二者均为孵化酶分子。60kD分子很不稳定，在纯化过程中极易降解，40kD分子可能是由60kD分子经过降解或自身降解丢失了两个CUB重复区而形成的，而CUB重复区很可能在对受精膜分子结构的修饰或改造中具有重要作用。

Coli BL21 (DE3) plysS for expression under induction of IPTG. The expressed product in a form of inclusion body was denaturalized with urea, purified by nickel ion chelate affinity chromatography, renaturalized by gradient dialysis and identified by Western blot.

coli BL21(DE3) plysS进行诱导表达，采用Ni(上标 2+)螯合亲和层析法纯化经尿素变性的包涵体，梯度透析复性纯化的目的蛋白，并进行Western blot鉴定。

The recombinant adenovirus was then large amplified and purified with Viraprep purification column and desalted.

大规模扩增重组腺病毒并用Vira-prep纯化柱纯化。

The crude DNA pellets were recovered by dialyzing and then could be used for molecular operation such as amplification of 16S rRNA gene and digestion of restriction enzymes.The plasmid pUC19 was prepared and digested with BamHI.

粗提DNA用半透膜透析回收纯化，纯化后的DNA可以进行16sRNA基因PCR扩增，酶切等分子生物学操作，具有较高的质量。

The gradual process of allele homologizing in a closed inbreeding population is imitated quantitatively by using linear algebra model.

线性代数模型定量地模拟了封闭近交群体内等位基因逐步纯化的过程，同时能客观分析不同自交代之间的生物数学特征及内在联系，根据模型还能由初始条件预报纯化基因型的频率。

The target protein was purified under the native condition and the polyclonal antibody was prepared by immunizing rabbits.

因此，可以在自然条件下进行目的蛋白的纯化，并将纯化的蛋白对家兔进行免疫而制备多克隆抗体。

As she looked at Warrington's manly face, and dark, melancholy eyes, she had settled in her mind that he must have been the victim of an unhappy attachment.

每逢看到沃林顿那刚毅的脸，那乌黑、忧郁的眼睛，她便会相信，他一定作过不幸的爱情的受害者。

Maybe they'll disappear into a pothole.

也许他们将在壶穴里消失