Bacterial Protein Expression

Published on May 15, 2013 at 10:21 PM

AMSBIO provides bacterial protein expression services. This includes several aspects including gene synthesis, mutations, cleavage site construction, refolding, Tag cleavage and test expression.

The services provide a full spectrum of protein purification methods to achieve the best suitable level of protein purity for downstream applications.

Phases of bacterial protein expression

The whole process involves two phases:-

Phase 1

This phase involves cloning the gene of interest into a bacterial expression vector along with a His-tag or other selected tag (common ones in use include tags with GST, His, or Thioredoxin).

Once cloned this gene is amplified and then isolated out of a customer-supplied vector and is subcloned onto a bacterial expression vector. The subcloned gene is verified and authenticated using restriction enzyme digest and sequencing.

The recombinant constructs are then transformed into high efficiency expression bacterial strain. Mini-induction to over-express the target protein when required.

The tests for expression of the target protein for verification include SDS-PAGE and/or western blot, where customer provides the antibody or AMSBIO provide it from their vast library of over 65,000.

The turnaround time is generally around 2 to 4 weeks.

Phase 2

This stage involves large scale culture and purification of the over- expressing cell line.

In phase 2 500ml of the bacterial culture will be induced with IPTG and this is harvested for protein purification. For protein purification Ni-NTA (for 6xHIS-tagged proteins) may be used or glutathione (for GST-tagged proteins) may be chosen.

Either method uses beads at either native or denatured conditions. The solubility of the purified protein is not guaranteed and the turnaround time is about 2 weeks.

Final deliverable

The final deliverable is a 5 μg construct, purified protein along with verification documentation on sequencing data, design form, Western blot and SDS-PAGE result. If both phases are performed the product includes purified protein.

Advantages of AMSBIO bacterial protein expression services

  • Comprehensive expression systems from bacterial cell lines.
  • Faster turnaround times with as little as 4 weeks time for product development.
  • Verification and authentication of purity of protein as well as verification of target protein expression using SDA-PAGE and/or Western Blot.
  • One-stop services that provides gene synthesis, stable cell line generation and bacterial protein expression

Source

http://www.amsbio.com/custombacterialproteinexpression.aspx


AMSBIO is a premier provider of quality life science research reagents and services helping customers develop innovative methods, processes, products and medicines. This is achieved by offering small and medium size manufacturers, academic groups and revenue generating biotechs a unique partnership for the global market and by providing state of the art and cost effective solutions to end users and partners.

Last updated: May 15, 2013 at 10:56 PM

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