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- 详细信息
- 文献和实验
- 技术资料
- 库存：
54
- 英文名：
Recombinant Human CHGA protein
- 保质期：
12个月
- 供应商：
上海帛科
- 保存条件：
-20 to -80 °C
- 规格：
50μg、100ug 、1mg
商品介绍：
内毒素: Please contact with the lab for this information.
蛋白构建: A DNA sequence encoding the human CHGA (Met158-Gly457) was fused with His tag
制剂: Supplied as solution form in PBS pH 7.5 or lyophilized from PBS pH 7.5.
运输方式: In general, proteins are provided as lyophilized powder/frozen liquid. They are shipped out with dry ice/blue ice unless customers require otherwise.
稳定性&储存: Use a manual defrost freezer and avoid repeated freeze thaw cycles.Store at 2 to 8 °C for one week .Store at -20 to -80 °C for twelve months from the date of receipt.
复溶: Reconstitute in sterile water for a stock solution.A copy of datasheet will be provided with the products, please refer to it for details.
分子别名: Chromogranin-A,CgA,Pituitary secretory protein I,SP-I,Vasostatin-1,Vasostatin I,Vasostatin-2,Vasostatin II,SL21.
背景介绍: The acidic glycoprotein chromogranin A (CHGA) is co-stored/co-secreted with catecholamines and crucial for secretory vesicle biogenesis in neuronal/neuroendocrine cells. CHGA is dysregulated in several cardiovascular diseases, but the underlying mechanisms are not well established. The CHGA promoter haplotype 2 (occurring in a large proportion of the world population) in enhancing CHGA expression in haplotype 2 carriers who may be at higher risk for cardiovascular/metabolic disorders. Chromogranin A (CHGA) plays a fundamental role in the biogenesis of catecholamine secretory granules. Changes in storage and release of CHGA in clinical and experimental hypertension prompted us to study whether genetic variation at the CHGA locus might contribute to alterations in autonomic function, and hence hypertension and its target organ consequences such as hypertensive renal disease (nephrosclerosis). Chromogranin A (CHGA) plays a catalytic role in formation of catecholamine storage vesicles and also serves as precursor to the peptide fragment catestatin, a catecholamine secretory inhibitor whose expression is diminished in the hypertensive individuals. The chromogranin/secretogranin proteins are costored and coreleased with catecholamines from secretory vesicles in chromaffin cells and noradrenergic neurons. Chromogranin A (CHGA) regulates catecholamine storage and release through intracellular (vesiculogenic) and extracellular (catecholamine release-inhibitory) mechanisms. CHGA is a candidate gene for autonomic dysfunction syndromes, including intermediate phenotypes that contribute to human hypertension.
表达载体在基因工程有以下几种元件：
（1）选择标志的编码序列；
（2）可控转录的启动子；
（3）转录调控序列(转录终止子，核糖体结合位点)；
（4）一个多限制酶切位点接头；
（5）宿主体内自主复制的序列。
重组蛋白质的诱导表达：
1. 挑取转化有质粒的单菌落， 接种于 3ml 选择性 LB 液体培养基中， 37 oC，250 rpm/min 振摇培养过夜。
2. 次日将培养过夜的菌液 500 μl 再接种于 10ml（1:20）选择性 LB 液体培养基 中， 37 oC，250 rpm/min 振摇培养至光密度（OD600=0.6）时，取 1 ml 样本作 为诱导前标本， 10000g 离心 1min 收集菌体沉淀，－20 oC 冻存备用。
3. 加入 1 mol/L IPTG 于菌液中，使 IPTG 终浓度为 1 mM ，37 oC ，250 rpm/min 振摇培养 4 ～ 5 小时。取 1 ml 样本作为诱导后标本，同上法收集菌体沉淀， -20 oC 冻存备用。
4. 将诱导前后菌体沉淀用 20 ~ 40 μl PBS（pH= 8.0）重悬， 加入等体积的 2×SDS 上样缓冲液， 煮沸加热 5min，SDS 聚凝胶（SDS-PAGE）电泳分离 ， 考马斯亮蓝染色 3 小时后，脱色观察结果。
5. 选取诱导成功的细菌克隆，扩大诱导规模，收集菌体沉淀，于－ 20 oC 保存，准备做下一步分析及纯化。
公司出售的相关产品：
重组蛋白质的可溶性鉴定：
1. 将按上法诱导培养后收集的菌体重悬于裂解液 1 (Lysis buffer under native conditions )中，然后在－80 oC 低温冰箱中放置 10 min。
2. 冰中解冻。
3. 在冰浴上用超声破碎仪破菌 6 次， 每次 10 sec，间歇 10 sec，电压 200-300 V。
4. 10000g，4oC，离心 20min，取上清（为溶液 A）， － 20 oC 保存； 另将沉淀用 同样裂解液 1 溶解（为溶液 B），同样 －20 oC 保存，供后继分析使用。
5. 将上述 A、B 溶液和诱导前后的细菌进行 SDS-PAGE 电泳， 考马斯亮蓝染色， 比较分析重组蛋白质的溶解性。如果诱导表达的蛋白质位于 A 溶液中， 则为可溶性蛋白；如果是在 B 溶液中，则为非可溶蛋白。
重组蛋白质的分离纯化：
1. 将菌体沉淀溶于适量裂解液 2（Lysis buffer under denaturing conditions ）中，室温下搅拌和吹打沉淀，避免泡沫生成。
2. 10000g ，4 oC，离心 30 min，收集上清液。
3. 将 Ni-NTAAgarose 充填柱子，并连接于 Pharmarcia 低压液相层析系统，用 5 倍柱体积的裂解液 2 平衡 Ni-NTAAgarose，调节 A280 值至零线。
4. 将适量上清液上样到 Ni-NTAAgarose 柱子中， 并用 lysis buffer 冲洗至 A280 值低于 0.01。
5. 分别用 5 ～ 10 倍柱体积的清洗液 1 和清洗液 2（Washbuffer 1 and 2）清洗柱 子，直至 A280 值低于 0.01。
6. 用洗脱液（Elution buffer）洗脱重组蛋白质， 在 A280 值监测下，收集出现峰 线后含有重组蛋白的所有洗脱液。
| 货号 | BK-DB2155 | 用途 | 仅供科研研究实验 |
| 纯度 | >90% as determined by SDS-PAGE | 预测分子量 | 35.5 kDa |
| 表达宿主 | E.coli | 种属 | Homo sapiens (Human) |
蛋白构建: A DNA sequence encoding the human CHGA (Met158-Gly457) was fused with His tag
制剂: Supplied as solution form in PBS pH 7.5 or lyophilized from PBS pH 7.5.
运输方式: In general, proteins are provided as lyophilized powder/frozen liquid. They are shipped out with dry ice/blue ice unless customers require otherwise.
稳定性&储存: Use a manual defrost freezer and avoid repeated freeze thaw cycles.Store at 2 to 8 °C for one week .Store at -20 to -80 °C for twelve months from the date of receipt.
复溶: Reconstitute in sterile water for a stock solution.A copy of datasheet will be provided with the products, please refer to it for details.
分子别名: Chromogranin-A,CgA,Pituitary secretory protein I,SP-I,Vasostatin-1,Vasostatin I,Vasostatin-2,Vasostatin II,SL21.
背景介绍: The acidic glycoprotein chromogranin A (CHGA) is co-stored/co-secreted with catecholamines and crucial for secretory vesicle biogenesis in neuronal/neuroendocrine cells. CHGA is dysregulated in several cardiovascular diseases, but the underlying mechanisms are not well established. The CHGA promoter haplotype 2 (occurring in a large proportion of the world population) in enhancing CHGA expression in haplotype 2 carriers who may be at higher risk for cardiovascular/metabolic disorders. Chromogranin A (CHGA) plays a fundamental role in the biogenesis of catecholamine secretory granules. Changes in storage and release of CHGA in clinical and experimental hypertension prompted us to study whether genetic variation at the CHGA locus might contribute to alterations in autonomic function, and hence hypertension and its target organ consequences such as hypertensive renal disease (nephrosclerosis). Chromogranin A (CHGA) plays a catalytic role in formation of catecholamine storage vesicles and also serves as precursor to the peptide fragment catestatin, a catecholamine secretory inhibitor whose expression is diminished in the hypertensive individuals. The chromogranin/secretogranin proteins are costored and coreleased with catecholamines from secretory vesicles in chromaffin cells and noradrenergic neurons. Chromogranin A (CHGA) regulates catecholamine storage and release through intracellular (vesiculogenic) and extracellular (catecholamine release-inhibitory) mechanisms. CHGA is a candidate gene for autonomic dysfunction syndromes, including intermediate phenotypes that contribute to human hypertension.
表达载体在基因工程有以下几种元件：
（1）选择标志的编码序列；
（2）可控转录的启动子；
（3）转录调控序列(转录终止子，核糖体结合位点)；
（4）一个多限制酶切位点接头；
（5）宿主体内自主复制的序列。
重组蛋白质的诱导表达：
1. 挑取转化有质粒的单菌落， 接种于 3ml 选择性 LB 液体培养基中， 37 oC，250 rpm/min 振摇培养过夜。
2. 次日将培养过夜的菌液 500 μl 再接种于 10ml（1:20）选择性 LB 液体培养基 中， 37 oC，250 rpm/min 振摇培养至光密度（OD600=0.6）时，取 1 ml 样本作 为诱导前标本， 10000g 离心 1min 收集菌体沉淀，－20 oC 冻存备用。
3. 加入 1 mol/L IPTG 于菌液中，使 IPTG 终浓度为 1 mM ，37 oC ，250 rpm/min 振摇培养 4 ～ 5 小时。取 1 ml 样本作为诱导后标本，同上法收集菌体沉淀， -20 oC 冻存备用。
4. 将诱导前后菌体沉淀用 20 ~ 40 μl PBS（pH= 8.0）重悬， 加入等体积的 2×SDS 上样缓冲液， 煮沸加热 5min，SDS 聚凝胶（SDS-PAGE）电泳分离 ， 考马斯亮蓝染色 3 小时后，脱色观察结果。
5. 选取诱导成功的细菌克隆，扩大诱导规模，收集菌体沉淀，于－ 20 oC 保存，准备做下一步分析及纯化。
公司出售的相关产品：
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| Phospho-MDM2 (Ser166) Antibody Blocking Peptide | Recombinant Human COL3A1 |
| BST1 Antibody Blocking Peptide | Recombinant Human LMNB2 |
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| AGTR1 Antibody Blocking Peptide | Recombinant Human JNK1 |
| Calpain 1 Antibody Blocking Peptide | Recombinant Human IMPA1 |
| connexin-30 Antibody Blocking Peptide | Recombinant Human KAP1 |
| Collagen II Antibody Blocking Peptide | Recombinant Human FAF2 |
| phospho-STMN1 (Ser24) Antibody Blocking Peptide | Recombinant Human CHGA proteinRecombinant Human Rubicon |
| CD300C Antibody Blocking Peptide | Recombinant Human SDCCAG8 |
| AK1 Antibody Blocking Peptide | Recombinant Human Macrophage Inflammatory3 alpha |
1. 将按上法诱导培养后收集的菌体重悬于裂解液 1 (Lysis buffer under native conditions )中，然后在－80 oC 低温冰箱中放置 10 min。
2. 冰中解冻。
3. 在冰浴上用超声破碎仪破菌 6 次， 每次 10 sec，间歇 10 sec，电压 200-300 V。
4. 10000g，4oC，离心 20min，取上清（为溶液 A）， － 20 oC 保存； 另将沉淀用 同样裂解液 1 溶解（为溶液 B），同样 －20 oC 保存，供后继分析使用。
5. 将上述 A、B 溶液和诱导前后的细菌进行 SDS-PAGE 电泳， 考马斯亮蓝染色， 比较分析重组蛋白质的溶解性。如果诱导表达的蛋白质位于 A 溶液中， 则为可溶性蛋白；如果是在 B 溶液中，则为非可溶蛋白。
重组蛋白质的分离纯化：
1. 将菌体沉淀溶于适量裂解液 2（Lysis buffer under denaturing conditions ）中，室温下搅拌和吹打沉淀，避免泡沫生成。
2. 10000g ，4 oC，离心 30 min，收集上清液。
3. 将 Ni-NTAAgarose 充填柱子，并连接于 Pharmarcia 低压液相层析系统，用 5 倍柱体积的裂解液 2 平衡 Ni-NTAAgarose，调节 A280 值至零线。
4. 将适量上清液上样到 Ni-NTAAgarose 柱子中， 并用 lysis buffer 冲洗至 A280 值低于 0.01。
5. 分别用 5 ～ 10 倍柱体积的清洗液 1 和清洗液 2（Washbuffer 1 and 2）清洗柱 子，直至 A280 值低于 0.01。
6. 用洗脱液（Elution buffer）洗脱重组蛋白质， 在 A280 值监测下，收集出现峰 线后含有重组蛋白的所有洗脱液。
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Recombinant Human CHGA protein
￥1290 - 3960
