Protein Expression Technology

Protein Expression refers to processes that describe how living cells or organisms synthesize, modify and regulate proteins. Custom protein expression can be synthesized through laboratory techniques, while recombinant protein production is done through cellular machinery. Protein expression has become an important laboratory technology in biochemistry, molecular biology, protein research and other scientific research fields that require functional proteins for research.

Commonly used protein expression systems are: bacterial protein expression system, yeast protein expression system, mammalian cell expression and insect protein expression system.

Take the Expression Protocol in E. coli as an example

• Codon optimized gene synthesis and vector construction

1. Optimize codon and mRNA structure.
2. Fuse gene to a protein tag and insert them into an expression vector.
3. Verify the correctness of construction by sequencing.

• Transform expression vector into E. coli competent cells

1. Add expression vector to thawed competent cells and heat-shocked cells to LB broth and shake.
2. Plate cell culture onto LB agar plates with appropriate antibiotic.

• Starter Culture

1. Pick single colony of expression strain into 5 ml of LB with appropriate antibiotic.
2. Shake at 37℃ for 3-5 h.

• Expansion of Starter Culture

1. Expand the culture by adding the starter culture to larger volume LB with antibiotic (room temp).
2. Incubate for 1-4 hours until culture density of OD600 reaches 0.5-0.6.

• Induction

1. Choose 37℃ induction or room temp (20℃) induction.

• Cells collection and lysis

1. Centrifuge the cells at 3,500 ×g for 20 min.
2. Resuspend cells in ice-cold PBS and re-centrifuge in an appropriate sized tube.
3. Remove the supernatant and freeze pellet for later processing.

Fig. 1 The general protocol of expression process from gene to protein.

• Mammalian expression systems
  Mammalian cells are ideal systems for expressing mammalian proteins, which require multiple post-translational modifications to perform proper protein functions. Most DNA constructs designed for mammalian expression utilize viral promoters to achieve robust expression after transfection. Mammalian systems can express proteins instantaneously or through stable cell lines, both of which can result in high protein production if transfection is successful.
• Insect expression systems
  Insect cells can also be utilized to produce complex eukaryotic proteins with the correct post-translational modifications. There are two common types of insect expression systems: baculovirus-infected and non-lytic insect cells.
• Yeast expression systems
  Yeast is a great expression system to generate large quantities of recombinant eukaryotic proteins. Saccharomyces cerevisiae is the most reliable and commonly used species for expression systems.
• Plant expression systems
  Plants provide a cheap and low-tech means of mass expression of recombinant proteins. Plant cells have been used to express a wide range of proteins including antibodies and pharmaceuticals, specifically interleukins.

• Production of purified proteins (structural, enzyme, drug discovery)
• Production of pharmaceutical protein
• Cell-based research

Laboratory Centrifuges

A centrifuge is any device that applies a sustained centrifugal force—that is, a force due to rotation. The widest use of centrifuges is for the concentration and purification of materials in suspension or dissolved in fluids. Suspended particles denser than the suspending liquid tend to migrate toward the periphery, while those less dense move toward the centre.

Pipette

Pipette is a small piece of apparatus which typically consists of a narrow tube into which fluid is drawn by suction (as for dispensing or measurement) and retained by closing the upper end. Pipettes are a common laboratory tool and come in a multitude of channels, displacements and construction materials. There are different pipettes for all kinds of uses within the lab.

Laboratory Rockers and Shakers

A rocker or shaker is a piece of laboratory equipment used to mix, blend, or agitate substances in a tube or flask by shaking them. These important tools are used for a variety of applications including cell culture, DNA extraction, mixing reagents, and staining gels and blots. A shaker contains an oscillating board that is used to place the flasks, beakers, or test tubes.

Laboratory Incubators

An incubator is a device used to grow and maintain microbiological cultures or cell cultures. It is made up of a chamber with a regulated temperature. An incubator maintains optimal temperature, humidity, and other conditions such as the CO2 and oxygen content of the atmosphere inside. Incubators are essential for much experimental work in cell biology, microbiology, and molecular biology.

Spectrophotometers

Spectrophotometer is an optical instrument that measures the intensity of light in relation to the wavelength. It can measure just about anything such as: plastics, paper, liquid, metal, fabric and so on. A spectrophotometer is used to check for the quality of product in a production facility and identify and quantify microscopic samples in analytical laboratories.

Laboratory Refrigeration

Laboratory refrigerators are designed with special features and options for use in the laboratory, including a range of refrigerator and freezer accessories. The primary function of a laboratory refrigerator is to maintain a defined, internal storage temperature in order to provide secure storage and protection of temperature-sensitive products, samples, specimens, chemicals, drugs, solutions, and other substances.

Electrophoresis Systems

Electrophoresis equipment applies an electric charge to molecules, causing them to migrate towards their oppositely charged electrode. The technique is found in all research and clinical laboratories utilizing DNA and protein applications, and is divided into gel and capillary techniques. Choice of electrophoresis equipment depends on the molecule of study, appropriate method of separation and downstream application.

Water Baths

A water bath is laboratory equipment made from a container filled with heated water. It is used to incubate samples in water at a constant temperature over a long period of time. Most water baths have a digital or an analogue interface to allow users to set a desired temperature. A laboratory water bath is a preferred heat source for heating flammable chemicals instead of an open flame to prevent ignition.

STEMart provides you with a variety of protein expression technology equipment or consumables to meet your various R&D and application needs. If you have any questions or requirements for protein expression technology, please feel free to contact us.