Team:Slovenia/nanoBRICKs.html

From 2009.igem.org

Development of the BioBRICKS

The main goal of iGEM project was to create versatile two or three dimensional protein structures. Shapes of these structures depend not only on protein domains characteristics but also on the length of the linker connecting protein domains as schematically presented in Figure1.

Figure 1

First challenge of our iGEM team was a development of a simple and efficient cloning system for extending a linker between interconnected protein domains. The need for that is the fact that almost all parts were ordered by gene synthesis adapted for cloning into BioBrick standards. To avoid ordering same protein domain with different linker extensions, the NEW BioBrick standard, named the linker-extension standard was designed. As basics, the BBF RFC 25 (fusion protein (Freiburg) BioBrick assembly) standard was used. The introduced modifications in multiple-cloning site enable (i) in-frame fusion, (ii) friendly scar and (iii) simple extension of linker between parts.
A comprehensive description of the linker-extension standard is under [http://dspace.mit.edu/bitstream/handle/1721.1/46705/BBFRFC37.pdf?sequence=1 BBF RFC37].

Functionalised linker-extension standard, nanoBRICKs[PRO]

Although most promoters, ribosomal binding sites, tags and terminators are currently specified as separate parts in the Registry, we are now moving to a new design in which these elements are included within the vector. Our working hypothesis is that the new design will reduce the likelihood of unexpected functional composition problems between promoter / terminator and coding sequence.
Several types of nanoBRICKs[PRO] vectors were constructed (Figure 2) with different 5' and 3' sequences coding specific promoters, terminators and coding sequences: KSI domain, his-tag, etc.

Figure 2

Table of functionalised nanoBRICKs[PRO] vectors