Composite

Part:BBa_K4368006

Designed by: Carmen Jimenez Amores, Alonso Molina Calvo, Paola Morales Gonzalez   Group: iGEM22_UMA_MALAGA   (2022-09-26)
Revision as of 18:26, 7 October 2022 by Carmeenjmnezz (Talk | contribs) (→‎Enzyme digestion)


pcstA + rbs + GFP + terminator

Description

GFP (BBa_E0040) is the gene coding green fluorescent protein which is a protein synthesized by the jellyfish Aequorea victoria. 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 glucose concentration (BBa_K118011).

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.

Enzyme digestion

UMA GFP.png

Sequence and Features


Assembly Compatibility:
  • 10
    COMPATIBLE WITH RFC[10]
  • 12
    COMPATIBLE WITH RFC[12]
  • 21
    COMPATIBLE WITH RFC[21]
  • 23
    COMPATIBLE WITH RFC[23]
  • 25
    COMPATIBLE WITH RFC[25]
  • 1000
    INCOMPATIBLE WITH RFC[1000]
    Illegal BsaI.rc site found at 804

Contribution

  • Group: [1]
  • Author: Molina Calvo, Alonso


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