Difference between revisions of "Part:BBa K2920001"

Latest revision as of 06:40, 20 October 2019

Peptide production process

-BBa-K2920727:We can prove our gene has the antimicrobial functions after we translate the constructed plasmid containing the peptide gene, so we can achieve the mass production of our specific peptide by genetic engineering approach and develop further applications.

-BBa:K1911005 Coding eGFP =eGFP = Enhanced green fluorescent protein which can be used as a reporter protein. eGFP was used in our project to show the effects of attaching the degradation tag to a target gene. In essence, the eGFP should be less prominent in E. coli cells which contain the tags compared to the E. coli cells that have been constructed, which do not have any degradation tags attached to the eGFP. eGFP can be used as a reporter protein and helps the observer easily see the coloration of E. coli cells which indicate a successful transformation.

-BBa:I719005 Regulatory (T7 Promoter): The constitutive promoter derived from the T7 bacteriophage allows a high expression of proteins only when the T7 polymerase is present.

-BBa: J18909 tag (His 6x tag) :This codes for six Histidine residues to allow for His column protein purification. It will work best if put at either the N or C terminus of your protein construct.

-BBa: K2323012 tag (SUMO tag): The SUMO tag increases the solubility of the tagged protein and can be cleaved with the SUMO protease.

-BBa: K731721 terminator(T7 terminator):a wild type terminator from T7 bacteriophage -BBa:K1638034 coding (Sce VMA Intein):This part consists of an intein from Saccharomyces cerevisiae VMA1 gene including a CBD-tag. This intein is a thiol-induced self-cleavable protein that enables the release of a C-terminal fused protein. The chitin-binding domain is an affinity tag that enables affinity purification on a chitin column. First, the target protein fused to intein is loaded and washed on the chitin column. Using a thiol reagent, like dithiothreitol (DTT), an on-column cleavage is induced and the target protein is released.

-BBa_I719005 (T7 Promoter): Constitutive promoter derived from the T7 bacteriophage. Allows high expression of proteins only when the T7 polymerase is present.

We used the different functions of the biobrick to increase the process efficiency. For example, the SUMO can help the peptide increases the solubility, the egfp protein can helps the observer easily see the coloration of E. coli , and the his tag will help us with the protein purification.

Antimicrobial peptide has been in the world for a while, but still couldn’t be common for people; there are several reasons, for instance, high prices, the production time is too long, the techniques couldn’t be accepted by people, etc. While there are various peptide company, but most of all are synthesized by chemical method, it takes lots of time, and the costs is high, e.g. 1 mg of peptide costs 700-1,000 NTD. To change disadvantages of the peptide company and improve it, for antimicrobial peptide, we are using the technique of genetic engineering to do the research. In this combination of biobrick, our final goal is to use an efficient, convenience way to produce it, in order to do that we add lots of proteins to help it produce. For example, his tag that could purify easily, sumo that could increase the solubility, and the t7 promoter that have highly specificity, etc.

Technically, so far, the manufacture of peptide could split to two categories: chemical method and genetic engineering method. For the peptide that has shorter sequence or the small-scale production, chemical method has the advantage, because of the foundation that built from synthetic chemical factor. However, as the peptide we need to synthesize is bigger, the accuracy would be decreased. Therefore, if consider to increase the scale of production, increase the production, or peptide sequence is longer, we are going to face some technique problems. Although, the technique of recombinants DNA is difficult, and require addition costs, once the scale is increased successfully, the number of productions would be increased, then apparently, the costs would be decreased; companies and factories could use this technique to prepare the procession, systematic the procession of the production, e.g. bacteria cultivation, disruption, purify, these all could be systematic in factories, thus, using the technique recombinant DNA to research on the peptide-making could be a great expectation of peptide development.

Sequence and Features