Composite

Part:BBa_K4368004

Designed by: Carmen Jimenez Amores, Alonso Molina Calvo, Paola Morales Gonzalez   Group: iGEM22_UMA_MALAGA   (2022-09-26)
Revision as of 22:43, 6 October 2022 by Alonso (Talk | contribs)


pcI + rbs + glgC16 + terminator

Description

GlgC16 encodes for the ADP-glucose pyrophosphorylase gene of Escherichia coli (BBa_K118016). This enzyme is responsible for the synthesis of starch-like starch using glucose as a substrate. In addition, this part includes the composition used by the team, which includes a strong rbs (BBa_B0030), a double terminator (BBa_B0015) as well as a promoter inducible by the cI protein of the phage lambda (BBa_R1051). The gene has been placed under the control of this promoter to build the glucose concentration-based gene regulatory circuit that integrates all our parts.

Characterization

The expression cassette sequence was digested with EcoRI and PstI enzymes and subsequently ligated with a chloramphenicol resistant plasmid backbone (Cm). Transforming bacteria were created with this plasmid and seeded on LB-Agar+Cm plates. After growth, colonies were selected based on their color (white) and DNA extraction was performed using the Promega PureYield Plasmid Miniprep System kit. The resulting DNA is used for further digestion with EcoRI and PstI. The digests are then run on a 0.75% agarose gel at 90 mV voltage and constant amperage. BioRad brand RedSafe is used as an intercalating developing agent.

Sequence and Features


Assembly Compatibility:
  • 10
    COMPATIBLE WITH RFC[10]
  • 12
    COMPATIBLE WITH RFC[12]
  • 21
    INCOMPATIBLE WITH RFC[21]
    Illegal BamHI site found at 272
  • 23
    COMPATIBLE WITH RFC[23]
  • 25
    COMPATIBLE WITH RFC[25]
  • 1000
    COMPATIBLE WITH RFC[1000]

Contribution

  • Group: [1]
  • Author: Jiménez Amores, Carmen


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Categories
Parameters
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