Difference between revisions of "Part:BBa K4605007"
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==Description==
 
==Description==
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BpsA stands for the Blue-pigment indigoidine synthetase gene.Itself is derived from Streptomyces lavendulae and is used in the synthesis of indigo. It can only be activated from inative apo-form to the active holo-bpsA by the addition of CoA to its PCP, catalyzed by PPTase, which synthesizes two molecules of glutamine into one molecule of indigo. Corynebacterium glutamicum is usually used to express bpsA for high indigo production.
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To explore whether more colored fibers could be expressed in K.xylinus. We wanted to express EGFP in K.xylinus using a strong promoter (J23100) as a way to get fluorescent cellulose membranes.
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==Experiments==
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In this experiment we will modify Komagataeibacter xylinus to express bpsA for one-step synthesis of colored fibers, and also codon optimize the bpsA coding sequence.
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We chose to use the Anderson family promoter, first, because it is compatible with K.xylinus, and there is enough references to report that these promoters have been tested in several of bacteria such as G. xylinus 700178, G. hansenii 53582, and K. rhaeticus iGEM.Then we selected three of the strong promoters:J23100,J23104 and J23119.We linked it to bpsA in the hope that fluorescent cellulose could be expressed.
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==Experiment==
 
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===<strong>Expression of bpsA in K. xylinus</strong>===
 
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With previous basic explorations, we will use a wood vinegar compatible PSB1A2 plasmid backbone, ligated with promoters such as strong promoters (J23104, J23102, etc.), and bpsA sequences to try to express bpsA in K. xylinus while binding to bacterial cellulose membranes.
 
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===<strong>References</strong>===
 
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[1] Mohammad Rifqi Ghiffary, Cindy Pricilia Surya Prabowo, Komal Sharma, Yuchun Yan, Sang Yup Lee, and Hyun Uk Kim.High-Level Production of the Natural Blue Pigment Indigoidine from Metabolically Engineered Corynebacterium glutamicum for Sustainable Fabric Dyes ACS Sustainable Chemistry & Engineering 2021 9 (19), 6613-6622
 
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[2]Teh MY, Ooi KH, Danny Teo SX, Bin Mansoor ME, Shaun Lim WZ, Tan MH. An Expanded Synthetic Biology Toolkit for Gene Expression Control in Acetobacteraceae. ACS Synth Biol. 2019 Apr 19;8(4):708-723.
 
  
 
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Latest revision as of 06:30, 8 October 2023


Strong promoters to improve the bpsA expression

Description

To explore whether more colored fibers could be expressed in K.xylinus. We wanted to express EGFP in K.xylinus using a strong promoter (J23100) as a way to get fluorescent cellulose membranes.

Experiments

We chose to use the Anderson family promoter, first, because it is compatible with K.xylinus, and there is enough references to report that these promoters have been tested in several of bacteria such as G. xylinus 700178, G. hansenii 53582, and K. rhaeticus iGEM.Then we selected three of the strong promoters:J23100,J23104 and J23119.We linked it to bpsA in the hope that fluorescent cellulose could be expressed.

Sequence and Features


Assembly Compatibility:
  • 10
    COMPATIBLE WITH RFC[10]
  • 12
    INCOMPATIBLE WITH RFC[12]
    Illegal NheI site found at 7
    Illegal NheI site found at 30
  • 21
    COMPATIBLE WITH RFC[21]
  • 23
    COMPATIBLE WITH RFC[23]
  • 25
    COMPATIBLE WITH RFC[25]
  • 1000
    COMPATIBLE WITH RFC[1000]