Part:BBa_K338004:Design
PHA Synthase Composite, Part 2/2
- 10COMPATIBLE WITH RFC[10]
- 12COMPATIBLE WITH RFC[12]
- 21INCOMPATIBLE WITH RFC[21]Illegal BglII site found at 1348
- 23COMPATIBLE WITH RFC[23]
- 25INCOMPATIBLE WITH RFC[25]Illegal AgeI site found at 697
Illegal AgeI site found at 1047 - 1000COMPATIBLE WITH RFC[1000]
Design Notes
When ligated downstream of BBa_K338003, the completed construct was designed to express all three PHA synthase genes required to make PHB oligomers from soybean oil. The three genes would be transcribed polycistronically on a single mRNA transcript under the IPTG-inducible control of the BBa_K215000 promoter. Naturally, each gene is preceded by a standard RBS (BBa_B0034) and the transcript finishes with a strong terminator (BBa_B0015), for a total size of about 3500bp.
Note that these three genes should only cause the production of PHB oligomers in cells, not hardened plastic. A crosslinking agent is required to link the oligomers and form the final plastic product. Over-expression of the phaC1 gene could cause some crosslinking, but this has not been experimentally verified.
SY Lee describes how a similar gene construct (pSYL105) was used to produce very large amounts of PHB, up to 80-90% of the dry cell weight, under certain conditions. Synthesis of PHB is related to the amount of acetyl-CoA available - synthesis was bolstered in the presence of complex nitrogen sources, amino acids, or oleic acid. He also mentions that PHB production was highly dependent on the particular bacterial strain used. [1]
Source
Registry of Standard Biological Parts
References
- Lee, SY. Bacterial polyhydroxyalkanoates. Biotechnology and Bioengineering 49, 1-14 (1996).