Difference between revisions of "Part:BBa K1720000:Design"

 
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===Design Notes===
 
===Design Notes===
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This subunit coding part with a GFP reporter was then transfected into HEK293 cells and in vivo green fluorescence signal was observed under fluorescence microscope. After that we used lentiviral vector to transfected alpha 3 unit and beta 3 unit to HEK293 cells together. Since soluble guanylate cyclases (sGC) are heterodimeric proteins that catalyze the conversion of GTP to 3',5'-cyclic GMP(cGMP) and pyrophosphate. The level of cGMP will be up regulated.The positive control was HEK293 cells that treat with Sodium Nitroprusside ,a NO donator that activate sGC and up regulate the level of cGMP. A negative control was made by transfecting an empty vector that does not contain sGC subunit. We used Elisa to detect cGMP level.  
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The vector we used in our experiment was created using scarless golden gate assembly.We cloned this basic part into a psb1c3 vector without promoter , so that user can choose their own promoter for different purpose. After standardizing our part, a promoter can be easily inserted in front of our part. This part is only a subunit of human guanylate cyclase,.To form holoenzyme this part needs to work with BBa_K1720001 together.
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===Source===
 
===Source===
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We get the information of Human guanylate cyclase 1,soluble, alpha 3 unit here
 
http://www.ncbi.nlm.nih.gov/gene/2982
 
http://www.ncbi.nlm.nih.gov/gene/2982
  
 
===References===
 
===References===
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1. Gheorghiade M, Marti CN, Sabbah HN, Roessig L, Greene SJ, Bohm M, Burnett JC, Campia U, Cleland JG, Collins SP et al: Soluble  guanylate cyclase: a potential therapeutic target for heart failure. Heart failure reviews 2013, 18(2):123-134.
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2. Sharina IG, Cote GJ, Martin E, Doursout MF, Murad F: RNA splicing in regulation of nitric oxide receptor soluble guanylyl cyclase. Nitric oxide : biology and chemistry / official journal of the Nitric Oxide Society 2011, 25(3):265-274.
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3. Marinko M, Novakovic A, Nenezic D, Stojanovic I, Milojevic P, Jovic M, Ugresic N, Kanjuh V, Yang Q, He GW: Nicorandil directly and cyclic GMP-dependently opens K channels in human bypass grafts. Journal of pharmacological sciences 2015.

Latest revision as of 07:30, 4 September 2015

Human guanylate cyclase1,soluble, alpha 3 unit


Assembly Compatibility:
  • 10
    INCOMPATIBLE WITH RFC[10]
    Illegal PstI site found at 1498
    Illegal PstI site found at 1827
  • 12
    INCOMPATIBLE WITH RFC[12]
    Illegal PstI site found at 1498
    Illegal PstI site found at 1827
  • 21
    COMPATIBLE WITH RFC[21]
  • 23
    INCOMPATIBLE WITH RFC[23]
    Illegal PstI site found at 1498
    Illegal PstI site found at 1827
  • 25
    INCOMPATIBLE WITH RFC[25]
    Illegal PstI site found at 1498
    Illegal PstI site found at 1827
  • 1000
    INCOMPATIBLE WITH RFC[1000]
    Illegal BsaI.rc site found at 1576
    Illegal SapI site found at 1446


Design Notes

The vector we used in our experiment was created using scarless golden gate assembly.We cloned this basic part into a psb1c3 vector without promoter , so that user can choose their own promoter for different purpose. After standardizing our part, a promoter can be easily inserted in front of our part. This part is only a subunit of human guanylate cyclase,.To form holoenzyme this part needs to work with BBa_K1720001 together.

Source

We get the information of Human guanylate cyclase 1,soluble, alpha 3 unit here http://www.ncbi.nlm.nih.gov/gene/2982

References

1. Gheorghiade M, Marti CN, Sabbah HN, Roessig L, Greene SJ, Bohm M, Burnett JC, Campia U, Cleland JG, Collins SP et al: Soluble guanylate cyclase: a potential therapeutic target for heart failure. Heart failure reviews 2013, 18(2):123-134.

2. Sharina IG, Cote GJ, Martin E, Doursout MF, Murad F: RNA splicing in regulation of nitric oxide receptor soluble guanylyl cyclase. Nitric oxide : biology and chemistry / official journal of the Nitric Oxide Society 2011, 25(3):265-274.

3. Marinko M, Novakovic A, Nenezic D, Stojanovic I, Milojevic P, Jovic M, Ugresic N, Kanjuh V, Yang Q, He GW: Nicorandil directly and cyclic GMP-dependently opens K channels in human bypass grafts. Journal of pharmacological sciences 2015.