Team:Caltech/BBa K338003

<partinfo>BBa_K338003 short</partinfo>

This is half of a planned part which would contain all three PHA synthase genes required to produce [http://en.wikipedia.org/wiki/Polyhydroxybutyrate polyhydroxybutyrate] (PHB) in cells: phaA, phaB1, phaC1. This half contains only phaA: <partinfo>BBa_K156012</partinfo>. It was designed to be ligated upstream of part BBa_K338004.

Usage and Biology

Design

When ligated upstream of BBa_K338004, 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 <partinfo>BBa_K215000</partinfo> promoter. Naturally, each gene is preceded by a standard RBS (<partinfo>B0034</partinfo>) and the transcript finishes with a strong terminator (<partinfo>B0015</partinfo>), 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.

Literature

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. [10]