Difference between revisions of "Part:BBa K823054"
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For N-terminal fusion proteins with cgeA (display on B. subtilis spores)
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For generating N-terminal fusion proteins with CgeA, displayed on the surface of ''Bacillus subtilis'' endospores.
  
 
=='''Sporo'''vector==
 
=='''Sporo'''vector==

Revision as of 01:27, 27 September 2012

For generating N-terminal fusion proteins with CgeA, displayed on the surface of Bacillus subtilis endospores.

Sporovector

Our Sporovector is specially designed to give you the opportunity to easily create [http://2012.igem.org/Team:LMU-Munich/Spore_Coat_Proteins Sporobeads] with any proteins you can think of. It is available as BioBrick in pSB1C3 and can be cloned in any of our empty B. subtilis vectors. We also offer it “precloned” in pSBBs4S which in this version lacks NgoMIV restriction sites.

Sporovectorcloning.png

The Sporovector was created by ligation of three cut PCR-products. It is designed to allow N-terminal fusion of genes in Freiburg standard ([http://dspace.mit.edu/bitstream/handle/1721.1/45140/BBF_RFC%2025.pdf?sequence=1 RCF25]). Therefore, the gene to be inserted must be cut with XbaI and AgeI and the vector must be digested with XbaI and NgoMIV. By this digest, the RFP cassette will be removed from this vector to allow easy selection for the integration of the new insert. After ligation, there is a scar of six nucleotides between both genes but no restriction site left. The gene fusion is controlled by the PcotYZ-promoter which was the strongest one in our promoter evaluations. The C-terminal gene part is cgeA, one of the two known spore crust proteins.

The vector is cloned but has not been tested with an insert, yet. We are working on further Sporovectors to offer also fusion with cotZ as well as C-terminal fusions.

This vector is part of the [http://www.2012.igem.org/Team:LMU-Munich Beadzillus Project].