Part:BBa_K1111009:Experience

Sequencing of the part revealed the presence of an unexpected STOP codon before the superfolder GFP sequence. This could be due to a mistake in the design of the primers or in their fabrication. Isolation of the expressed protein should still be possible since a His tag was added to its CDS together with the Gibson assembly primers. Bacteria were transformed, inoculated and grown overnight so that an assay could be performed. 50ml of 20% arabinose was then added to the 5ml of their LB+Chl growth medium, to activate the pBAD/araC promoter and induce gelE expression. Western blots with anti-His antibodies were then performed on the culture medium and cell lysates but revealed themselves negative. As a last resort, Ni-NTA spin kit purification (for His tags) of bacteria prepared in the same way was done.

Applications of BBa_K1111009

This part can be used to induce the expression of a gelatin-degrading protein upon a desired signal. Another promoter, responding to the specific requirements if the user, can be incorporated into the plasmid instead of the pBAD/araC promoter by Gibson assembly.

User Reviews

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