Overview
New Composite Parts
- We have designed two composite parts for the homologous recombination of the membrane protein of Shewanella: BBa_K4134066, BBa_K4134077 and proved they worked as we expected. Since the two parts function in the form of linear DNA, we designed corresponding plasmids: BBa_K4134067 , BBa_K4134078 which could amplify in E.coli. cells.
New Basic Parts
We also registered and uploaded basic parts BBa_K4134001, BBa_K4134002, BBa_K4134003, BBa_K4134004, BBa_K4134011 , BBa_K4134012 .
We expand the applications of Part: BBa_K1980007 as silver ions indicator through reference support and our experimental results.
We improved the composite part BBa_K1795024 turn it into a new homologous recombination device BBa_K4134066 with reporter(Kanamycin resistance).
The correct transformation of E. coli. strain for plasmids BBa_K4134067 , BBa_K4134078.
The correct transformation and homologous recombination of Shevanella oneidensis MR-1 strain for part BBa_K4134066.
All Parts We Made
| Part Number | Part Name | Part Type | Abstract |
|---|---|---|---|
| BBa_K4134001 | AtoxⅠ | Coding | AtoxⅠ(Homo sapiens antioxidant Ⅰcopper chaperone) has been reported to combine silver ions in form of a tetrasilver cluster in its dimer. Atox1 captures silver ions and paves the way for a new approach to synthesizing silver nanoparticles. |
| BBa_K4134002 | AgBP2 | Coding | AgBP2 is a typical silver-binding peptide consisting of 12 amino acids. We read about the silver-binding peptide AgBP2 selected from a combinatorial peptide display library and engineered it onto the Shewanella membrane protein (BpfA-AgBP2). |
| BBa_K4134003 | KanR | Coding | KanR stands for the Kanamycin resistance gene. We kept the coding region and removed its initiation codon so that KanR shares the same initiation codon with BpfA. Then Kanamycin resistance can be used as the basis to judge whether the strain is recombinant that displays silver-binding protein. |
| BBa_K4134004 | Flexible Linker | DNA | As an integral part of fusion protein recombination, Linker plays an important role in the construction of stable and biologically active fusion proteins. Linker is an amino acid chain that acts as a link between two fusion proteins and is flexible enough to allow the proteins on both sides to perform their independent functions. |
| BBa_K4134011 | BpfA(C-terminal 1000bp) | Coding | BpfA stands for the biofilm-promoting protein A, a large surface protein. This 1000bp-long fragment of BpfA is the head of our homologous recombination device, which fuses the destination fragment to the C-terminus of BpfA. |
| BBa_K4134012 | AggC(N-terminal 1000bp) | Coding | AggC is a gene downstream of the BpfA. This 1000bp-long fragment of AggC is the tail of our homologous recombination device. |
| BBa_K1980007 | pCusC mKate2 | Reporter | This composite part consists of the promoter pCusC (BBa_K1980004), a copper-responsive promoter activated by the CusS/R two-component system, with a downstream RFP variant (mKate). We expand its applications as silver ions indicator through reference support and our experimental results. |
| BBa_K4134066 | BpfA-Atox1-Linker-KanR-loxP-AggC | Device | We designed a device for fusion protein recombination. With this device, you can display any small protein to the C-terminus of BpfA, a large membrane protein of S. oneidensis. Considering the addition of KanR, you can easily confirm whether homologous recombination succeeds or not, since only the recombinants can grow into colonies on the medium supplemented with kanamycin. The part number for the existing part we improved: BBa_K1795024 |
| BBa_K4134067 | pUC57mini-BpfA-Atox1-Linker-KanR-loxP-AggC | Plasmid | This linear device(BBa_K4134066) is attached to pUC57mini on both ends and cyclized into a plasmid. When introduced to E.coli(DH 5alpha), it amplifies. Moreover, pUC57mini consists of the replication origin, AmpR, and its promoter. |
| BBa_K4134077 | BpfA-AgBP2-Linker-KanR-loxP-AggC | Device | It has the same design as the device BBa_K4134066 for fusion protein recombination. The only difference is that we replace Atox1 with Agbp2. |
| BBa_K4134078 | pUC57mini-BpfA-AgBP2-Linker-KanR-loxP-AggC | Plasmid | This linear device(BBa_K4134077) is attached to pUC57mini on both ends and cyclized into a plasmid. When introduced to E.coli(DH 5alpha), it amplifies. Moreover, pUC57mini consists of the replication origin, AmpR, and its promoter. |