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Team:Tokyo Tech/Projects/PHAs/detail/index.htm
From 2012.igem.org
Production trial of PHAs by past teams
We introduce some attempts in the past iGEM to show how great our work is in iGEM.
| Part number | Description | States | experience |
| BBa_K338003 | PHA Synthase Composite, Part 1/2 | planning | |
| BBa_K338004 | PHA Synthase Composite, Part 1/2 | Available |
They couldn’t prove that their engineered bacteria produced PHB according to the team wiki.
| Part number | Description | States | experience |
| BBa_K342001 | Pha C (poly-β-hydroxybutyrate polymerase) | available |
They registered and submitted a part, which contains only phaC sequence, as worked. But the team wiki shows that the part doesn’t meet the standard which is required in iGEM because they couldn’t cut their part with Xba1.
| Part number | Description | States | experience |
| BBa_K282000 | phaAB | available |
| Part number | Description | States | experience |
| BBa_K089001 | phaA gene | planning | |
| BBa_K089002 | phaB gene | planning | |
| BBa_K089003 | phaC gene | planning |
| Part number | Description | States | experience |
| BBa_K125801 | RBS-phaA | available | |
| BBa_K125802 | RBS-phaB | Planning | |
| BBa_K125803 | RBS-phaC | Available | |
| BBa_K125804 | RBS-phaE | Available | |
| BBa_K125501 | phaA BioPlastic polyhydroxybutyrate synthesis pathway (origin PCC6803 slr1994) | Available | |
| BBa_K125502 | phaB BioPlastic polyhydroxybutyrate synthesis pathway (origin PCC6803 slr1994) | Available | |
| BBa_K125503 | phaC BioPlastic polyhydroxybutyrate synthesis pathway (origin PCC6803 slr1994) | Available | |
| BBa_K125504 | phaE BioPlastic polyhydroxybutyrate synthesis pathway (origin PCC6803 slr1994) | Available |
| Part number | Description | States | experience |
| BBa_K156031 | RBS + phaA + double terminator | available | |
| BBa_K156033 | RBS + phaB1 + double terminator | available | |
| BBa_K156034 | RBS + phaC1 + double terminator | available | |
| BBa_K156012 | phaA (acetyl-CoA acetyltransferase) | available | |
| BBa_K156013 | phaB1 (acetyacetyl-CoA reductase) | available | |
| BBa_K156014 | phaC1 (Poly(3-hydroxybutyrate) polymerase) | available | |
| BBa_K156021 | Promoter + RBS + phaA + double terminator | available | |
| BBa_K156018 | Promoter + RBS + phaA | planning | |
| BBa_K156019 | Promoter + RBS + phaB1 | planning | |
| BBa_K156020 | Promoter + RBS + phaC1 | planning | |
| BBa_K156022 | Promoter + RBS + phaB1 + double terminator | planning | |
| BBa_K156023 | Promoter + RBS + phaC1 + double terminator | planning |
Construction of pha-C1-A-B1 in Biobrick format
To construct a part that meets Biobrick format, we have modified the phaC1-A-B1 operon not to contain forbidden restriction enzyme sites. First, we cloned the wild type gene phaC1-A-B1 from R.eutropha H16 by using PCR and inserted the gene into pSB1C3. However, wild type phaC1-A-B1 gene sequence contains one NotI and three PstI recognition sites that are not allowed in Biobrick format. To get phaC1-A-B1 sequence without these recognition sites, we ordered the chemically synthesized DNA from IDT/MBL. In this chemically synthesized DNA, coding is optimized for E.coli. We used restriction enzyme XbaI (on pSB1C3) and BsrGI (on phaC1-A-B1) to insert sequence. That is to say, we got Poly[(R)-3-hydroxybutyrate] synthesizing gene in Biobrick format (BBa_K934001). Fig3:construction of phaC1-A-B1
