Existing Parts (used in our parts)
Basic Parts
| Part | Name | Length (bp) | Description |
|---|---|---|---|
| BBa_K106005 | mCherry | 708 | Red fluorescent protein that helps in protein detection |
| BBa_K4344066 | T7 promoter | 18 | IPTG inducible promoter |
| BBa_J23100 | J23100 | 35 | Constitutive promoter from the J23119 family |
| BBa_K1123000 | FNR promoter | 120 | Anaerobic inducible promoter |
| BBa_K1624002 | LacO | 23 | Lac operator sequence, involved in transcriptional regulation of the lac operon |
| BBa_K5369112 | KanR | 816 | Kanamycin resistance gene |
| BBa_B0016 | T7 terminator | 48 | T7 gene 1 specific transcriptional terminator |
| BBa_K4941006 | pBR322 ori | 589 | pBR322 plasmid-derived ColE1 replicon |
| BBa_K4514038 | f1 ori | 456 | Bacteriophage origin of replication |
New Parts
Basic Parts
| Part | Name | Length (bp) | Description |
|---|---|---|---|
| BBa_25VE6X4F | EcTI | 519 | Enterolobium contortisiliquum trypsin inhibitor |
| BBa_25LGVMRT | CYP450 | 3147 | Cytochrome P450-BM3 enzyme found in Bacillus megaterium |
| BBa_25G9KW20 | CYP450DM | 3147 | Cytochrome P450-BM3 enzyme with double mutation at A82F and A328F. Improves efficiency and accepts phenol as initial substrate to produce HQ |
| BBa_25I3G9UB | Hecate | 75 | Synthetic lytic peptide |
| BBa_2551OCR8 | Melittin | 84 | Lytic peptide from bee venom |
| BBa_25ECQV2O | Modified NSP4-K | 63 | NSP4 secretion signal peptide modified to include K-K trypsin cleavage site |
| BBa_25UNFLT8 | AmpR | 861 | Ampicillin resistance gene |
Composite Parts
| Part | Name | Length (bp) | Description |
|---|---|---|---|
| BBa_25N3Q748 | EcTI-His | 555 | EcTI attached to His tag for detection/purification |
| BBa_25GIB12Q | CYP450-Flag | 3174 | CYP450 fused with Flag tag for detection |
| BBa_25FQMYFL | CYP450DM-Flag | 3174 | CYP450DM fused with Flag tag for detection |
| BBa_25WGKZEX | CYP450-3×(GGGGS)-mCherry | 3903 | CYP450 fused with mCherry using GGGGS linker |
| BBa_25V45BQK | CYP450DM-3×(GGGGS)-mCherry | 3903 | CYP450DM fused with mCherry using GGGGS linker |
| BBa_251SSAZQ | NSP4-K-Hecate | 135 | Hecate fused with modified NSP4 signal peptide, allowing extracellular secretion and trypsin cleavage |
| BBa_25TLAX5C | NSP4-K-Melittin | 141 | Melittin fused with modified NSP4 signal peptide, allowing extracellular secretion and trypsin cleavage |
| Name | Part | Simple name | Length (bp) | Description | Sequence |
|---|---|---|---|---|---|
| pET28a-FNRpromoter>mCherry:rrnB_TT | BBa_25E58ZN2 | FNR-mCherry | 5104 | The mCherry reporter gene is inserted in the pET28a backbone; the gene expression is regulated by anaerobic promoter FNR. The purpose of this design is for verify the FNR promoter functionality since mCherry activity can be easily detected. | link |
| pET28a-FNRpromoter>NSP4-K-K-H(CO_Ec):rrnB_TT | BBa_25G76T6V | FNR-hecate | 4528 | The fused Hecate peptide with modified NSP4-K secretion is inserted in the pET28a backbone; the gene expression is regulated by the anaerobic promoter FNR. The Kanamycin resistance (BBa_K5369112) helps in screening for successful transformation. | link |
| pET28a-FNRpromoter>NSP4-K(CO_Ec)K-Melittin(Co_Ec):rrnB_TT | BBa_25UVLJOJ | FNR-Melittin | 4537 | The fused Melittin peptide with modified NSP4-K secretion is inserted in the pET28a backbone; the gene expression is regulated by the anaerobic promoter FNR. The Kanamycin resistance (BBa_K5369112) helps in screening for successful transformation. | link |
| pET-J23100>EcTl(co_Ec/GGGGS/6×His | BBa_251SK4RC | J23100-EcTI | 5273 | The fused protein EcTI-His is inserted in the pET plasmid. We used the J23100 promoter to express the EcTI-His protein because it provides a moderate level of expression. While a stronger promoter like J23119 exists, it could lead to high levels of EcTI that are toxic to the E. coli cells. | link |
| pET-CYP450/Flag(co-Ec)* | BBa_25HS5FHK | T7-CYP450-flag | 8810 | The fused protein CYP450-Flag is inserted into pET plasmid, where T7 promoter regulates the gene expression in the presence of IPTG. The Ampicillin resistance gene (BBa_25UNFLT8) allows screening successful transformation. | link |
| pET-CYP450/Flag(co-Ec)*(A83FA329F) | BBa_25MIYL3S | T7-CYP450 DM-flag | 8810 | The fused protein CYP450DM-Flag is inserted into pET plasmid, where T7 promoter regulates the gene expression in the presence of IPTG. The Ampicillin resistance gene (BBa_25UNFLT8) allows screening successful transformation. | link |
| pET-{CYP450(co_Ec)*}:3xGGGGS:mCherry | BBa_25H2URHN | T7-CYP450-mCherry | 9539 | The fused protein CYP450-mCherry is inserted into pET plasmid, where T7 promoter regulates the gene expression in the presence of IPTG. The Ampicillin resistance gene (BBa_25UNFLT8) allows screening successful transformation. | link |
| pET-{CYP450(co_Ec)*(A83FA329F)}:3xGGGGS:mCherry | BBa_25MRBB84 | T7-CYP450 DM-mCherry | 9539 | The fused protein CYP450DM-mCherry is inserted into pET plasmid, where T7 promoter regulates the gene expression in the presence of IPTG. The Ampicillin resistance gene (BBa_25UNFLT8) allows screening successful transformation. | link |