Team:Linkoping Sweden/Parts

Novosite

• Home

• Project

  • Description
  • Design
  • Parts
  • Model
  • Demonstrate

• Lab

  • Notebook
  • Experiments
  • Results
  • Characterization
  • Improvement
  • Safety

• People

  • Team Members
  • Collaborations
  • Attributions
  • Sponsors

• Parts

  • Parts Overview
  • Basic Parts
  • Composite Parts
  • Part Collection
  • Primers

• Human Practices

  • Human Practices
  • Public Engagement
• Awards

Back to the top Composit Parts Basic Parts Other Parts

Parts overview

Parts overview

Down below all parts that were used in our project can be seen. To quickly generate our parts we utilized the offer from IDT, and let them synthesize most of the sequences used. The characterized parts were instead used from the BioBrick kits. The base vector, BBa_K3182200 (pT7-CBD_cipA), were first cloned into pSB1C3 along with BBa_K1033933 (AsPink). Later pCons-AsPink (BBa_J23100, BBa_K1033933) were cloned into that vector and "pink-white screening" was utilized. After our base vector had been established the rest of our basic parts was cloned into it with the pink-white screening method (see BBa_K3182100). Our characterization of the binding domain and antimicrobial peptides and enzymes will hopefully reduce the need for antibiotics.

The use of Vibrio natriegens in our work

The team also used the newly found host, Vibrio natriegens (V. natriegens). To enable expression of our composite parts parts needed to be moved from pSB1C3 (unable to replicate in V. natriegens) to another vector, for this pUC19 was chosen, since it has already been proved to replicate in V. natriegens. The vector pUC19 was chosen due to its similarities with pSB1C3, being a small plasmid. It was slightly modified with termination sequences outside of the insert and the pLac and LacZ' was removed. The team then utilized the same pink-white screening method as before and successfully cloned in all of our basic parts into the pUC19-CBD- vector as well. However, we had much work to do with our agents already and did not fully explore V. natriegens.

Please note that none of the parts were designed with a termination sequence, this is because this would add one to all basic parts which would turn into 1000 extra base pairs for IDT to synthesize. Termination sequences are also troublesome to handle sometimes. We instead used vectors with termination sequences outside of the insert, to avoid adding complexity to our ligation reactions.

Composite parts

Table 1. All the composite parts was used in the wet lab part of our project. All team members worked with our composite parts.

Biobrick ID	Short name	Function	Size	Designed by
BBa_K3182000	pT7-CBDcipA-AsPink	Characterization of the carbohydrate binding domain	1287 bp	Oliver Hild Walett
BBa_K3182100	pT7-CBDcipA-pCons-AsPink	Carbohydrate binding domain assembly vector	1334 bp	Oliver Hild Walett
BBa_K3182200	pT7-CBDcipA	Base fusion vector for peptides and enzymes	585 bp	Oliver Hild Walett
BBa_K3182103	pT7-CBDcipA-PlyF307-SQ8C	CBD fusion - Acinetobacter genus lysin	1029 bp	Oliver Hild Walett
BBa_K3182104	pT7-CBDcipA-CHAP(K)	CBD fusion - Staphylococcus genus lysin	1074 bp	Oliver Hild Walett, Leo Juhlin
BBa_K3182105	pT7-CBDcipA-LL-37	CBD fusion - Antibacterial peptide LL-37	702 bp	Oliver Hild Walett
BBa_K3182106	pT7-CBDcipA-Magainin 2	CBD fusion - Antimicrobial peptide MG2	660 bp	Oliver Hild Walett
BBa_K3182107	pT7-CBDcipA-Pln1	CBD fusion - Antibacterial peptide Pln1	702 bp	Oliver Hild Walett
BBa_K3182108	pT7-CBDcipA-sfGFP	Characterization of the carbohydrate binding domain	1305 bp	Oliver Hild Walett

Basic parts

Table 2. Basic parts used in the wet lab part of our project. All basic parts used was designed to fit for assembly with BBa_K3182001.

Biobrick ID	Short name	Function	Size	Designed by
BBa_K3182001	CBDcipA	Carbohydrate binding domain with a C-terminal linker	516 bp	Oliver Hild Walett, Johan Larsson
BBa_K3182003	PlyF307-SQ8C	Acinetobacter genus lysin	444 bp	Oliver Hild Walett
BBa_K3182004	CHAP(K)	Staphylococcus genus lysin	489 bp	Leo Juhlin
BBa_K3182005	LL-37	Antibacterial peptide	117 bp	Oliver Hild Walett
BBa_K3182006	Magainin 2	Antimicrobial peptide	75 bp	Oliver Hild Walett
BBa_K3182007	Pln1	Antibacterial peptide	117 bp	Oliver Hild Walett
BBa_K3182009	Parseqα	Synthetic antimicrobial peptide	60 bp	Johan Larsson, Jonatan Baggman
BBa_K3182010	Parseqβ	Synthetic antimicrobial peptide	60 bp	Johan Larsson, Jonatan Baggman
BBa_K3182013	mNeonGreen	Fluorescent protein	771 bp	Oliver Hild Walett, iGEM Linköping 2018

Improved parts

Table 3. The improved version of iGEM Linköping 2018s reporter mNeonGreen. This fluorescent protein is also one of the brightest and fastest to mature.

Biobrick ID	Short name	Function	Size	Desgined by
BBa_K3182300	pT7-mNG	Improvement of iGEM Linköping 2018s biobrick	840 bp	Oliver Hild Walett, iGEM Linköping 2018

Verified parts

Table 4. Parts which were verified by our team members. Where EforRed gained more attention due to its chromo/fluorescent properties.

Biobrick ID	Short name	Function	Size	Designed by
BBa_K864402	pCons-eforRed	Verified function in E. coli	742 bp	Erik Lundin, iGEM Uppsala 2012
BBa_K1033933	AsPink	Verified function in E. coli	702 bp	Erik Lundin, iGEM Uppsala 2013
BBa_I746916	sfGFP	Verified function in E. coli	720 bp	Stefan Milde, iGEM Cambridge 2007

• 
• 
•