Abstract

Background: The primary interest for most molecular biologists has remained genes. The isolation and amplification of genes are the foremost criteria for gene-related studies.

Main Body: A simple strategy for the same is gene cloning by incorporating a gene into a vector or vehicle carrying a DNA molecule. This is further taken through the replication process with the help of living cells. A recombinant DNA molecule comes into the picture as a result of combining two DNAs possessing disparate sources. The required gene isolated from any source can be taken to the cloning process by inserting it into a suitable vector with desired properties to create recombinant DNA. Cloning involves the use of restriction enzymes termed restriction endonucleases or molecular scissors isolated from various sources. Various other enzymes like DNA ligases, used for joining the DNA fragments, DNA polymerases, used in producing multiple replicas of a source DNA are involved in cloning procedures. Furthermore, the produced recombinant DNA can be introduced into the host system in order to synthesize the product, the gene codes for. The obtained protein product is finally concentrated and purified by affinity chromatography, followed by analysis and confirmation by western blotting.

Conclusion: Cloning of genes and expressing the recombinant protein in bacterial system followed by purification by chromatography involves important tools and techniques in molecular biology.

Keywords: DNA Insertion; Bacterial Expression System; Molecular Cloning; Recombinant DNA Technology; Affinity Chromatography; Western Blotting

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Citation

Citation: Ansab Akhtar. “Cloning, Expression, and Purification of Recombinant Protein Utilizing Bacterial Expression System”. Acta Scientific Microbiology 4.11 (2021): 82-94.

Copyright

Copyright: © 2021 Ansab Akhtar. This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.