Part
We found a high expression level of promoter FBA1 when we reviewed the paper, and a team has expressed plasmids containing this promoter in S. cerevisiae in the past years. (Part number: BBa_K2765044,link: http://parts.igem.org/Part:BBa_K2765044) We further searched the literature to determine its basic promote strength.
Seung-pyo Hong et al[1] constructed an expression vector containing FBA1 in Yarrowia Lipolytica (pDMW212 & pDMW214), and measured its GUS Reportor activity and mRNA expression.
Figure 2. Histochemical and fluorometric GUS reporter assays to measure promoter activity. Fluorometric determination of GUS activity. GUSspecific activity was expressed as nM 4-methylumbelliferone released/min/mg protein Figure 2. Relative quantities of GUS mRNA. The relative transcript levels were calculated using the GUS mRNA level from strains with TEF1::GUS as the reference. Assays were done in quadruplicate.Troubleshooting
We have summarized the lessons learned from building the team until now, and have written a document to help future Igemer to avoid these problems.
- (1) Experiment
- Starting from the design of primers for dry experiments, the screening of wild strains, and the modeling of proteins for wet experiments,
- (2) Modeling
- There are some common models, such as the logistic equation most commonly used in the lab. For teams that prefer industrial tracks, Monod and Luedeking-Piret equations are also available. In addition, for the team that needs to express the enzyme, The Miesian equation is also a more commonly used model.
- (3) HP
- HP is a unique aspect of the IGEM.The judged Handbook is different every year, so ou need to keep an eye on what's happening to HP every year In addition to practical application research and interviews, it is difficult but significant for human practice to lead educational development and popularize synthetic biology concepts.
- (4) Team Building
- The international Genetic Engineering Competition is a biology discipline competition with high difficulty and high gold content. Faced with limited economic support, limited experimental period and limited experimental research, especially under the impact of COVID-19 this year, the negative impact of these problems will become more severe.
- For public school students, you need to deal with school resources in case you drop out due to lack of funds
- (5) Team management
- It is strongly not recommended that free-Rider participate in this competition and abandon the mentality of not contributing anything but waiting for the award. A truly united team is one that coordinates, develops together and moves toward the same goal. In this case, the team members should be highly enthusiastic about the international genetic engineering competition and have the ability to arrange and plan tasks and time coordination with certain intensity.
Open source blueprint
When we were designing the fermentation tank, we found that many models of the fermentation tank could be used universally, but the average student had no access to get these drawings and had to buy them in the store. Here we have uploaded some CAD drawings of the open source fermentor we found this year for future Igemer to refer to when designing hardware
download CAD fileReference:
[1] Hong S P , Seip J , Walters-Pollak D , et al. Engineering Yarrowia lipolytica to express secretory invertase with strong FBA1IN promoter[J]. Yeast, 2012, 29(2).