Part:BBa_K4883010
Ptef1-RIB1-ERBV-1-RIB7-PTV-ADE4-Tcyc1
This is a complete expression cassette, constructed by inserting RIB1-ERBV-1-RIB7-PTV-ADE4 (BBa_K4883009) between a strong promoter Ptef1 and a CYC1 terminator.
Sequence and Features
- 10COMPATIBLE WITH RFC[10]
- 12INCOMPATIBLE WITH RFC[12]Illegal NheI site found at 1743
- 21INCOMPATIBLE WITH RFC[21]Illegal BglII site found at 4090
Illegal BamHI site found at 836 - 23COMPATIBLE WITH RFC[23]
- 25INCOMPATIBLE WITH RFC[25]Illegal AgeI site found at 856
- 1000INCOMPATIBLE WITH RFC[1000]Illegal BsaI.rc site found at 176
Illegal BsaI.rc site found at 1675
Usage and Biology
To mitigate vitamin B2 deficiency, we planned to make vitamin B2-enriched bread with engineered yeast, Saccharomyces cerevisiae. Co-overexpression of RIB1 (BBa_K4883001), RIB7 (BBa_K4883002), and ADE4 (BBa_K4883000) should lead to the overproduction of vitamin B2 in S. cerevisiae. Ptef1 (BBa_K2407300) has been proven one of the strongest promoters of S. cerevisiae, so we used it for overexpression (Partow et al., 2010). A CYC1 terminator was also included to form the complete expression cassette.
Characterization
2023 Hangzhou-SDG Team characterized this part with vitamin B2 production
To find the strain with the highest vitamin B2 production, we constructed different strains overexpressing different combinations of RIB1, RIB7, and ADE4 (BBa_K4883011, BBa_K4883012, BBa_K4883013, BBa_K4883004, BBa_K4883006, BBa_K4883008, BBa_K4883010).
Vitamin B2 Production Tests in Liquid YPD Media
The WT S. cerevisiae S288C and the engineered strains were inoculated into YPD media, and cultured at 30 ℃, 180 rpm. The riboflavin concentrations in the supernatants were measured using high-performance liquid chromatography (HPLC).
| None |