Team:BIT-China/Parts
In 2018, BIT-China has submitted 16 parts. We divide our parts into 3 group, Regulator, Feedback and Output. Each group contains some parts with similar function or close relation. Different groups also connect with each other.
BBa_K2765026 and BBa_K2765021 is a collection of regulate components including coding protein and expression cassette, which can increase the accumulation of intracellular ROS through overexpressing yon1 induced by galactose.
BBa_K2765030 is 1000bp homologous arm of yca1 gene, which can knock out yca1 gene through homologous recombination so that the yeast's tolerance to H2O2 could be improved.
BBa_K2765000~BBa_K2765005 is a collection of feedback components that are downstream gene promoter of Yap1. We use them to turn on the expression of dCas9 which can inhibit the expression of yno1 gene. Considering the application of our system in the future, to realize different feedback effect, we verified different promoters, GLR1p, SOD2p, TRR1p, TRX2p and GSH1p, with different transcription level.
BBa_K2765040~BBa_K2765053 is a collection of output components including promoters, coding protein and expression cassette, which can express redox-sensitive green fluorescent protein roGFP2-Orp1 and display ROS content in cells by its fluorescence ratio.
According to this measurement principle, the concentration of whole roGFP2-Orp1 can influence the final signal output strength, even detect the same sample, so we verified four promoters, FBA1, TEF2, TEF1 and ENO2, with different transcription level to solve this interfere. Considering the application of our system in the future, we made this collection and wished these parts can be used in different antioxidant detection.
We performed codon optimization on team Connell 2017's previous part BBa_K2296006 and obtained our core part BBa_K2296006, which codes for roGFP2-Orp1 protein. Also we got As a result, it is more suitable to our chassis organisms---yeast and has better expression in yeast. Check here to know further information.
| No. | Biobrick | Name | Description | Length |
|---|
| Regulator | ||||
| 1 | BBa_K2765021 | yno1 | ROS Content Increaser | 1713 bp |
| 2 | BBa_K2765026 | GAL1p-yno1-CYC1t | Increase ROS Content | 2651 bp |
| 3 | BBa_K2765030 | homologous arm-ura(△yca1) | Knock out the yca1 gene | 3038 bp |
| Feedback | ||||
| 4 | BBa_K2765000 | GAL1p | Turn on the expression of dCas9 | 332 bp |
| 5 | BBa_K2765001 | SOD2p | Turn on the expression of dCas9 | 526 bp |
| 6 | BBa_K2765002 | TRR1p | Turn on the expression of dCas9 | 960 bp |
| 7 | BBa_K2765003 | TRX2p | Turn on the expression of dCas9 | 283 bp |
| 8 | BBa_K2765005 | GSH1p | Turn on the expression of dCas9 | 2027 bp |
| Output | ||||
| 9 | BBa_K2765040 | FBA1p | Turn on the expression of roGFP2-Orp1 | 631 bp |
| 10 | BBa_K2765042 | ENO2p | Turn on the expression of roGFP2-Orp1 | 512 bp |
| 11 | BBa_K2765043 | roGFP2-Orp1 | ROS Content Detector | 1317 bp |
| 12 | BBa_K2765045 | TEF1p | Turn on the expression of roGFP2-Orp1 | 402 bp |
| 13 | BBa_K2765050 | TEF2p-roGFP2-Orp1-CYC1t | Detect ROS Content | 1965 bp |
| 14 | BBa_K2765051 | TEF1p-roGFP2-Orp1- CYC1t | Detect ROS Content | 1964 bp |
| 15 | BBa_K2765052 | ENO2p-roGFP2-Orp1- CYC1t | Detect ROS Content | 2074 bp |
| 16 | BBa_K2765053 | FBA1p-roGFP2-Orp1- CYC1t | Detect ROS Content | 2193 bp |
