Translational_Unit

Part:BBa_K4879009

Designed by: Anuvind Pramod   Group: iGEM23_IISER-Pune-India   (2023-10-08)
Revision as of 10:35, 11 October 2023 by Sauron 3791 (Talk | contribs)


Yarrowia lipolytica HygR construct

The transcriptional unit for HygR, designed for expression in Y. lipolytica in order to delete the faa1 gene from our chassis.

Characterization

Cloning: The construct, along with homologous flanking sequences corresponding to the faa1 gene locus, were synthesized by iGEM’s DNA synthesis partner IDT. The resulting gene fragment was then PCR amplified with the respective primers to attain a higher copy number of the fragment. The amplification of the fragment was verified by running a 0.8% agarose gel, with the fragments expected to show at around the 3000 bp region.

The amplified fragment was then gel-extracted and subsequently transformed into our chassis.

Validation: The transformed yeast culture was then plated onto the selection plates containing about 200 µg/ml of hygromycin B and were incubated at 30°C. The growth of the yeast on the selection plates confirms the functional validity of the expression construct.

Sequence and Features


Assembly Compatibility:
  • 10
    INCOMPATIBLE WITH RFC[10]
    Illegal prefix found in sequence at 1561
    Illegal suffix found in sequence at 1831
    Illegal EcoRI site found at 684
    Illegal EcoRI site found at 778
    Illegal EcoRI site found at 1825
  • 12
    INCOMPATIBLE WITH RFC[12]
    Illegal EcoRI site found at 684
    Illegal EcoRI site found at 778
    Illegal EcoRI site found at 1561
    Illegal EcoRI site found at 1825
    Illegal SpeI site found at 1832
    Illegal PstI site found at 1846
    Illegal NotI site found at 1567
    Illegal NotI site found at 1839
  • 21
    INCOMPATIBLE WITH RFC[21]
    Illegal EcoRI site found at 684
    Illegal EcoRI site found at 778
    Illegal EcoRI site found at 1561
    Illegal EcoRI site found at 1825
  • 23
    INCOMPATIBLE WITH RFC[23]
    Illegal prefix found in sequence at 1561
    Illegal suffix found in sequence at 1832
    Illegal EcoRI site found at 684
    Illegal EcoRI site found at 778
    Illegal EcoRI site found at 1825
  • 25
    INCOMPATIBLE WITH RFC[25]
    Illegal prefix found in sequence at 1561
    Illegal EcoRI site found at 684
    Illegal EcoRI site found at 778
    Illegal EcoRI site found at 1825
    Illegal XbaI site found at 1576
    Illegal SpeI site found at 1832
    Illegal PstI site found at 1846
  • 1000
    COMPATIBLE WITH RFC[1000]


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Categories
Parameters
None