Team:Montpellier/WetLab Peptides

Design

General design

Each of our constructions contained RpsU promoter [1] which is a Lactobacillus jensenii strong promoter. This RpsU sequence also contains the putative sequence for the RBS. We added spacers to all of our constructions to unable easier use of the sequence and separation of the different genes of the sequences. We used two Terminators to our sequences :BBa_B0014 & BBa_B0015 to ensure the stopping of the transcription. Our constructions were assembled in the pLEM415 vector by Gibson Assembly method. pLEM415 is a plasmid that works in Lactobacilli species but it’s not specific to L. jensenii [2].

Figure 1: Caption of the designs.

LL-37

For the LL-37 sequence we used the RpsU promoter and the sequence coding the LL-37 peptide.

Figure 2: Design of the sequence coding the LL-37 protein with the RpsU promoter.

Figure 3: Design of the sequence coding the LL-37 protein with the pHyperSpank promoter.

SubtilosinA

Figure 4: Design of the sequence coding the Subtilosin protein with the RpsU promoter.

Figure 5: Design of the sequence coding the Iron Sulfur Cluster with the RpsU promoter.

Lacticin 3147

This circuit was made from 2 native genes of Lactococcus lactis ltA1 and ltnA that express Lacticin peptide. Also, the design contains Lacticin-post-transcriptional regulator ltM1 and M2. A promoter orthogonal was used : ptsH and differents spacer taken from igem_parts.

Figure 6: Design of the sequence coding the Lacticin 3147

References
[1]	Xiaowen Liu, et al,. 2006. Engineered vaginal lactobacillus strain for mucosal delivery of the human immunodeficiency virus inhibitor cyanovirin-N. Antimicrobial agents and chemotherapy 50(10), 3250-3259.
[2]	Bao, Sujin, et al.2013 "Distribution dynamics of recombinant Lactobacillus in the gastrointestinal tract of neonatal rats." PloS one 8.3 (2013): e60007.