biotINK project overview

General part design using RFC[10] & RFC[25]

All of our BioBricks were assembled according to the RFC[10] or RFC[25] standard. For all basic parts the RFC[10] standard was used. Because the RFC[10] scar codes for a stop codon this assembly standard does not allow the generation of fusion proteins. Therefore all fusion proteins were assembled by RFC[25] standard. Thus all RFC[10] and RFC[25] digestion sites were eliminated from the BioBricks either directly when planing the genesynthesis or by quick-change PCR. Only one of the BioBricks which we supply for the community could not be freed from all digestion sites, because one part of the BioBrick

Featured parts

Parts we improved

GMK_TK90: BBa_K404113 & BBa_K782063 were improved as both parts were wrong as they both contained a forbidden PstI restriction site that made it impossible to ligate BioBricks downstream of the BioBrick. A corrected BioBrick was provided (BBa_K2170060)

Tet-Operator: BBa_I739001 was improved by confirming functionality, adding a Tet-Operator together with an OmpA secretion signal yielding the part BBa_K2170141 which can be used by teams for the SEC-dependent secretion in the bacterial periplasm.

The collection of eukaryotic signal peptides in RFC[25] was expanded significantly by identifying non-working signal peptides (BBa_I712009, as not in RFC[25]), characterizing working signal peptides (BBa_K157001) and designing, constructing and testing two additional signal peptides with significantly higher functionality.

Impressions from the packaging

Fig.2 Finally it's done. Ready to send our 2016 BioBricks

Sendbox

<groupparts>iGEM2016 LMU-TUM_Munich</groupparts>