Difference between revisions of "Part:BBa K4073000:Design"
 
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BBa_K407300 regulates salt tolerance in plants
 
  
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This composite part consists of a CrGDPH3 salt-inducible promoter, a medium-strength RBS, the AtHKT1 gene, and a terminator. The regulation of the expression of the AtHKT1 gene is the function of this part. The CrGPDH3 salt-inducible promoter was used for the overexpression of proteins in a transgene in microalga Chlamydia reinhardtii.  We then decided to utilize this promoter because of its high NaCl response elements. The RBS initiates translation of the HKT1 gene, which is essential for its protein synthesis. Finally, we used a composite terminator which signals the end of transcription in our part. This is essential, as if we did not utilize a terminator, the AtHKT1 gene could potentially be over-expressed, leading to decreased salt concentrations which could inhibit plant growth (salt is an essential nutrient used in plant growth).
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__NOTOC__
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<partinfo>BBa_K4073000 short</partinfo>
  
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The source of the medium strength RBS, HKT1 gene, and terminator all came from the iGEM registry. The RBS is the part labeled Bba_B0032, the HKT1 gene is the protein coding sequence of the part labeled "BBa_K2665007”, while the terminator is the part "BBa_B0015." The promoter was derived from the NCBI sequence for the CrGPDH3 salt-inducible promoter.
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<partinfo>BBa_K4073000 SequenceAndFeatures</partinfo>
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===Design Notes===
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Some design considerations that we had to deal with were the regions of the promoter that we wanted to focus on. For example, both RIA1 and RIA3 were regions of interest, and so we had to decide which one of them would yield the most enhanced salinity mechanisms in the form of NaCl response elements. We ended up choosing the restriction enzymes SpeI to KpnI, encompassing both the RIA1 and the RIA3 regions, for the maximum effect of the mechanisms under theoretical NaCl treatments. Additionally, a medium strength RBS was used so that a decent strength of expression of the HKT1 gene was achieved (our other composite parts touch upon the other RBS strengths). Another important aspect to consider is that our design could have had the capability to be altered based on how experimental lab procedures followed through. However, due to the lack of access to a lab, we were unable to test the effects of this composite part on the salinity mechanisms of Arabidopsis Thaliana, and therefore most of our design considerations were centered around research gathered from original papers (https://link.springer.com/content/pdf/10.1007/s00253-019-09733-y.pdf).
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===Source===
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The source of the medium strength RBS, HKT1 gene, and terminator all came from the iGEM registry. The RBS is the part labeled Bba_B0032, the HKT1 gene is the protein coding sequence of the part labeled "BBa_K2665007”, while the terminator is the part "BBa_B0015." The promoter was derived from the NCBI sequence (​​https://www.ncbi.nlm.nih.gov/nuccore/1601834299) for the CrGPDH3 salt-inducible promoter.  
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===References===

Latest revision as of 06:43, 17 October 2021


BBa_K4073000 regulates salt tolerance in plants.


Assembly Compatibility:
  • 10
    COMPATIBLE WITH RFC[10]
  • 12
    COMPATIBLE WITH RFC[12]
  • 21
    COMPATIBLE WITH RFC[21]
  • 23
    COMPATIBLE WITH RFC[23]
  • 25
    INCOMPATIBLE WITH RFC[25]
    Illegal NgoMIV site found at 755
  • 1000
    INCOMPATIBLE WITH RFC[1000]
    Illegal BsaI site found at 1912
    Illegal BsaI.rc site found at 225


Design Notes

Some design considerations that we had to deal with were the regions of the promoter that we wanted to focus on. For example, both RIA1 and RIA3 were regions of interest, and so we had to decide which one of them would yield the most enhanced salinity mechanisms in the form of NaCl response elements. We ended up choosing the restriction enzymes SpeI to KpnI, encompassing both the RIA1 and the RIA3 regions, for the maximum effect of the mechanisms under theoretical NaCl treatments. Additionally, a medium strength RBS was used so that a decent strength of expression of the HKT1 gene was achieved (our other composite parts touch upon the other RBS strengths). Another important aspect to consider is that our design could have had the capability to be altered based on how experimental lab procedures followed through. However, due to the lack of access to a lab, we were unable to test the effects of this composite part on the salinity mechanisms of Arabidopsis Thaliana, and therefore most of our design considerations were centered around research gathered from original papers (https://link.springer.com/content/pdf/10.1007/s00253-019-09733-y.pdf).


Source

The source of the medium strength RBS, HKT1 gene, and terminator all came from the iGEM registry. The RBS is the part labeled Bba_B0032, the HKT1 gene is the protein coding sequence of the part labeled "BBa_K2665007”, while the terminator is the part "BBa_B0015." The promoter was derived from the NCBI sequence (​​https://www.ncbi.nlm.nih.gov/nuccore/1601834299) for the CrGPDH3 salt-inducible promoter.

References