GoldBio’s BL21 Chemically Competent E. coli cells are suitable for transformation and routine protein expression from non-T7 vectors. BL21 chemically competent cells feature a widely used host background, and are deficient in both lon (1) and ompT proteases. In addition, BL21 Chemically Competent E. coli cells are resistant to phage T1 (fhuA2).
Product SpecificationsCompetent cell type: Chemically CompetentDerivative of: BL21Species: E. coliFormat: TubesTransformation efficiency: ≥1 x 106 cfu/µg pUC19 DNABlue/white screening: Yes
Storage/Handling: This product may be shipped on dry ice. BL21 Chemically Competent E. coli cells should be stored at -80°C, pUC19 Control DNA should be stored at -20°C and recovery medium should be stored at 4°C immediately upon arrival. When stored under the recommended conditions and handled correctly, these products should be stable for at least 1 year from the date of receipt.
Genomic Features
- >1 x 106 cfu/µg efficiency
- Widely used host background
- Routine non-T7 vector expression.
- Deficient in both lon (1) and ompT proteases.
- Resistant to phage T1 (fhuA2).
GenotypeF- dcm ompT hsdS(rB- mB-) gal [malB+]K-12(λS)
Reagents Needed for One Reaction
- BL21 chemically competent cells: 50 µl
- DNA (or pUC19 Control, 10 pg/µl): 1 µl
- Recovery medium: 1 ml
Quality ControlTransformation efficiency is tested by using the pUC19 control DNA supplied with the kit and using the protocol given below. Transformation efficiency should be ≥1 x 106 CFU/µg pUC19 DNA. Untransformed cells are tested for appropriate antibiotic sensitivity.
General Guidelines
- Handle competent cells gently as they are highly sensitive to changes in temperature or mechanical lysis caused by pipetting.
- Thaw competent cells on ice, and transform cells immediately following thawing. After adding DNA, mix by tapping the tube gently. Do not mix cells by pipetting or vortexing.
Calculation of Transformation EfficiencyTransformation Efficiency (TE) is defined as the number of colony forming units (cfu) produced by transforming 1 µg of plasmid into a given volume of competent cells.
- TE = Colonies/µg/Dilution
- Colonies = the number of colonies counted
- µg = amount of DNA transformed in µg
- Dilution = total dilution of the DNA before plating
Example: Transform 1 µl of (10 pg/µl) control plasmid into 25 µl of cells, add 975 µl of Recovery Medium. Dilute 10 µl of this in 990 µl of Recovery Medium and plate 50 µl. Count the colonies on the plate the next day. If you count 250 colonies, the TE is calculated as follows:Colonies = 250µg of DNA = 0.00001 Dilution = 10/1000 x 50/1000 = 0.0005 TE = 250/0.00001/0.0005 = 5.0 × 1010
GoldBio活体成像技术：早在1999年由美国哈佛大学Weissleder博士率先提出了分子影像学(molecularimaging，MI)的概念，即应用影像学的方法对活体状态下的生物过程进行细胞和分子水平的定性和定量研究。活体成像便是基于分子影像学孕育而生的，通过这个成像系统，可以观测活体动物体内肿瘤的生长及转移，感染性疾病的发展进程，特定基因的表达等生物学过程。活体成像技术主要采用生物发光（bioluminescence）与荧光（fluorescence）两种技术。★生物发光是用荧光素酶基因标记细胞或DNA。★荧光技术则采用荧光报告基团（GFP、RFP,Cyt及dyes等）进行标记。★这一技术对肿瘤微小转移灶的检测灵敏度极高，不涉及放射性物质和方法,非常安全。操作极其简单、所得结果直观、灵敏度高。
活体成像两种检测技术介绍活体成像特点优点缺点生物发光检测bioluminescence★荧光素酶（Luciferase）对基因、细胞和活体动物进行标记；★荧光素酶催化底物（例如荧光素钾盐）反应后，会产生化学发光。这种光是由化学反应而来，不需要激发光；★标记方法是通过克隆技术，将荧光素酶的基因插入到预期观察的细胞染色体内，通过对克隆细胞进行筛选，培养出能稳定表达荧光素酶的细胞株。再将细胞株转移至特定的小鼠体内形成模型。★特异性强，无自发荧光；★高灵敏度，在体内可检测到几百个细胞，检测的深度在3-100px；★定量精确 ★信号较弱，检测时间较长；★仪器精密度要求较高；★细胞或基因需要转基因标记；★不可用于人体，不适用于抗体、多肽等标记荧光检测fluorescence★采用荧光报告基因(GFP、RFP等)或荧光染料进行标记；★需要外接激发光源，利用报告基因、荧光蛋白质或染料产生的荧光，就可以形成体内的生物光源。★荧光染料、蛋白标记能力强；★信号强，成像速度快，操作简便，实验成本较低；★未来可用于人；★适用范围广，可以是动物、细胞、微生物，也可以是抗体、药物、纳米材料等。★存在自发荧光，影响灵敏度；★光容易被动物组织吸收；★检测深度受限；★背景光干扰，定量准确度低
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