纯化

Methods The type Ⅳ pilin protein PILE was purified by HisTrapTM HP affinity columns ,then the purified protein was examined by SDS-PAGE and Western blot.

使用HisTrapTM HP亲和层析柱纯化Ⅳ型菌毛蛋白PILE，并对纯化蛋白进行SDS-PAGE和Westernblot鉴定。

ABSTRACT OBJECTIVE:To purify the extract from pleurotus ostreatus,and study on the physicochemical properties and antitumor activity in vitro of the purified component.

目的：对糙皮侧耳真菌提取液进行纯化，并对纯化组分的理化性质及体外抗肿瘤作用进行研究。

Primary culture of goat mammary gland cells could be derived by outgrowth of migrating cells from fragment of tissue. Fibroblast and epithelial cells could be pured according to their different sensibility to trypsin. Complex cultures treated by 0.25% trypsin in Hanks' for 5~7min at 37℃, the dispersed cells were mainly fibroblasts,then the rest treated by 0.15% trypsin-0.02% EDTA in Hanks'for 5~8min at 37℃, the majority cells harvested were mammary epithelial cells.

用组织块培养法可获得良好的山羊乳腺细胞原代培养物；培养的山羊乳腺成纤维细胞比上皮细胞对胰蛋白酶更敏感，据此可在传代过程中将二者分离纯化，获得纯细胞系；混合培养物先用0.25%胰蛋白酶在37℃消化5~7min 所收集的细胞主要为成纤维细胞；再加0.15%胰蛋白酶-0.02鞹A 消化液在37℃继续消化5~6min 所回收的细胞绝大多数为上皮细胞，经过2~3 代，即可得到纯化的乳腺上皮细胞系。

Primary culture of goat mammary gland cells could be derived by outgrowth of migrating cells from fragment of tissue. Fibroblast and epithelial cells could be pured according to their different sensibility to trypsin. Complex cultures treated by 0.25% trypsin in Hanks' for 5~7min at 37℃, the dispersed cells were mainly fibroblasts,then the rest treated by 0.15% trypsin-0.02% EDTA in Hanks'for 5~8min at 37℃, the majority cells harvested were mammary epithelial cells.

用组织块培养法可获得良好的山羊乳腺细胞原代培养物；培养的山羊乳腺成纤维细胞比上皮细胞对胰蛋白酶更敏感，据此可在传代过程中将二者分离纯化，获得纯细胞系；混合培养物先用0.25%胰蛋白酶在37℃消化5~7min 所收集的细胞主要为成纤维细胞；再加0.15%胰蛋白酶-0.02%EDTA 消化液在37℃继续消化5~6min 所回收的细胞绝大多数为上皮细胞，经过2~3 代，即可得到纯化的乳腺上皮细胞系。

The first passage cells were initially pured by enzyme digestion, then pured by enzyme digestion and repeated strapping alternatively in the second course of transferation and inoculation, the third period cultured cells showed typical clonali growth and were long fusiform fibroid cells with short growth cycle.

首次传代时应用胰酶消化法进行成纤维细胞的初次纯化，第2次转种时先后应用酶消化法和反复贴壁法再次进行细胞纯化，第3代以后基本可获得呈典型克隆性生长、长梭形纤维样细胞，生长周期短。

Object In this study we framed the refusion gene antibacterial peptide[ Cecropin A(l ~ 8)- Magainin (1 - 12),CA(1 ~ 8)-MA(l ~12),CAM] vetor and expressed it in the Escherichia.coli. Thenpurified the recombinded peptide in the Intein Mediated Purificationwith an Affinity Chitin-binding Tag-IMPACT system.

目的 构建重组基因抗菌肽[Cecropin A(1～8)-Magainin(1～12)，CA(1～8)-MA(1～12)，CAM]质粒载体并在大肠杆菌(Escherichia.coli)中表达，应用几丁质自切割分离纯化系统(InteinMediated Purification with an Affinity Chitin-binding Tag，IMPACT)分离纯化抗菌肽，并测定其抗菌活性。

Following attachment to the host immunized with purified antigens, midgut antigens, salivary gland antigens and reinfestation by I. sinensis, the midgut of tick revealed rather strikingly pathological changes, especially of the tick feeding on host immunized with 105KD purfied antigens the basal lamina became thinner, looser, and broken.

中华硬蜱叮咬纯化抗原、中肠抗原、唾液腺抗原免疫接种组和再次感染组宿主后，中肠可发生一系列明显的病理变化，尤以叮咬105KD纯化抗原组宿主后中肠病理变化最为严重。

With ultrasonic wave and lysozyme, the recombination N protein is split from bacteria. We dissolve it in guanidine hydrochloride, purify it with Ni〓 affinity chromatography column and renature it in vitro. By testing the antigen of the purified N protein, it shows that its activity is very high.

建立了用超声波和溶菌酶裂解菌体，用盐酸胍溶解，Ni〓亲合层析柱纯化及体外复性等方法，经检测证明纯化的VSV重组核蛋白抗原具有较高的活性。

Methods: A lysosome-targeted singal KFERQ was added to the C terminus of rRTA by DNA recombinant technology. A pKK223.3 expression system in E. coli was used to produce recombinant ricine A chain and rRTA-KFERQ. Recombinant proteins were purified by affinity chromatography using Blue-Sepharose 6B.

用DNA重组技术，将溶酶体的靶向信号肽KFERQ连接在RTA的羧基端；将构建好的重组质粒pKK223.3-RTA和pKK223.3-RTA-KFERQ转化感受态大肠杆菌JM109，经IPTG诱导表达RTA和RTA-KFERQ蛋白；重组蛋白质用Blue-Sepharose6B亲和柱纯化，以MTT法分别测定纯化后的RTA与RTA-KFERQ蛋白对体外培养的HEPG2、Hela、A5493种细胞的毒性作用。

Methods: A lysosome-targeted singal KFERQ was added to the C terminus of rRTA by DNA recombinant technology. A pKK223.3 expression system in E. coli was used to produce recombinant ricine A chain and rRTA-KFERQ. Recombinant proteins were purified by affinity chromatography using Blue-Sepharose 6B.

用DNA重组技术，將溶酶体的靶向信號肽KFERQ连接在RTA的羧基端；將构建好的重组质粒pKK223.3-RTA和pKK223.3-RTA-KFERQ转化感受態大肠桿菌JM109，经IPTG诱导表达RTA和RTA-KFERQ蛋白；重组蛋白质用Blue-Sepharose6B亲和柱纯化，以MTT法分别测定纯化后的RTA与RTA-KFERQ蛋白对体外培养的HEPG2、Hela、A5493种细胞的毒性作用。

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