技术文章您现在的位置:首页 > 技术文章 > 自动诱导超级肉汤培养基 Super Broth

自动诱导超级肉汤培养基 Super Broth

发布时间:2022-05-08   点击次数:285次

自动诱导超级肉汤培养基Super Broth产品描述:

自动诱导超级肉汤培养基(Autoinductive Super Broth)也称自动诱导SB,用于大肠杆菌Lac启动子驱动的的重组蛋白表达,无需添加IPTG。自诱导培养基能够获得高细胞密度,蛋白表达率高出IPTG诱导过程好几倍。

SB自动诱导培养基工作原理

SB培养基由于其高密度的营养成分,非常适合重组大肠杆菌的快速生长。超级肉汤培养基是一种加富培养基,配方基于TB培养基(Terrific Broth),后者由LB培养基改进而来,目的是达到高细胞密度。E. coli在该培养基中生长速度快于其在LB培养基或 2x YT肉汤中的生长速度。SB培养基被认为是很佳的质粒扩增培养基。一般来说,在液体培养基中大肠杆菌细胞生长的质量如下:LB Broth < 2x YT Broth < Terrific Broth < Super Broth。

该培养基中含有理想比例的葡萄糖和α-乳糖作为碳源和能量来源。胰蛋白胨提供氮源、维生素、矿物质和氨基酸,这些营养成分是细胞生长所必需的。酵母提取物是丰富的B族维生素来源。Studier salts为细菌细胞的生长提供很佳的生理环境。

通常,外源蛋白在细菌中表达时常使用IPTG诱导的启动子,如Lac启动子。当细胞密度达到理想值时,通过向培养基中加入IPTG的方法诱导蛋白表达。使用这个自动诱导培养基,不再需要监测细胞密度和添加IPTG。葡萄糖作为半乳糖操纵子的阻遏因子阻止细菌利用α-乳糖,而被优先代谢,促进高密度生长。一旦培养基中的葡萄糖被耗尽(通常发生在对数期的后期),乳糖被?-半乳糖苷酶转换成异乳糖(葡萄糖-1,6-半乳糖),而后者作为IPTG诱导型启动子的诱导剂,引起乳糖阻遏子从与DNA结合的位点上释放,启动重组蛋白的表达。对于DE3基因型的E.coli中,异乳糖使T7lac启动子去阻遏,并诱导lacUV5启动子表达T7 RNA聚合酶。通过这种方式,在细菌培养物生长到某一特定点时蛋白表达自发开始,去掉了监测细胞密度(OD600)和添加IPTG这两个步骤。与传统IPTG诱导的蛋白表达过程相比,使用自动诱导培养基极大地方便和简化了试验流程。


使用步骤:

1. 称取45.85g 脱水培养基粉末,用1L去离子水加热溶解。

2. 煮沸1min。

3. 115°C 灭菌20min。

不要过度加热,避免乳糖分解和培养基颜色加深。冷却后保存在2-8oC,如果不染菌,可以保存2个月左右。


Quantity: 500g

Appearance: Beige powder. Autoclaved medium should be amber.

Storage: 2oC – 25oC. When not in use, keep container closed to avoid hydration.


配方Formulation (g/L)

Tryptone: 16,00

Yeast Extract: 10,00

MgSO4: 0,15

(NH4) 2SO4: 3,30

KH2PO4: 6,80

Na2HPO4: 7,10

Glucose 0,50

Alpha Lactose 2,00

Final pH (25oC): 7,0 ± 0,2


配置Preparation:

Add 45,85g of the dehydrated medium to one liter of distilled water. Mix well and dissolve by heating with regular agitation. Boil for 1 minute in order to dissolve completely. Dispense in appropriate containers and sterilize by autoclaving at 121oC for 15 to 20 minutes. Store at 2oC to 8oC.


使用Usage

Commonly, heterologous protein expression is carried out in bacterial systems where the expression is under the control of an IPTG-inducible promoter, such as the Lac promoter. Cells are grown until a desired density and protein expression is subsequently induced by adding IPTG to the medium. With this Auto Induction Medium (AIM), it is no longer required to monitor cell density and to add IPTG at the proper stage, as the medium contains an optimized ratio of glucose and alpha lactose as carbon sources. Glucose, who serves as a repressor of the Lac operon, by preventing uptake of alpha lactose (hence and IPTG) is metabolized preferentially during growth, promoting high cell density. Once glucose is depleted, usually in mid to late log phase, lactose enters the cell where it is converted by ?galactosidase into allolactose, which in turn serves as the inducer of the IPTG-inducible promoter, resulting in protein expression. This is a great convenience and simplifies manual or automatic induction and analysis of multiple clones compared to conventional IPTG  induction.

靶点科技(北京)有限公司

靶点科技(北京)有限公司

地址:中关村生命科学园北清创意园2-4楼2层

© 2022 版权所有:靶点科技(北京)有限公司  备案号:京ICP备18027329号-2  总访问量:99679  站点地图  技术支持:化工仪器网  管理登陆

QQ在线客服
联系方式

15210359867

4000043669