Difference between revisions of "Part:BBa K1807007"

Latest revision as of 01:09, 19 September 2015

Synthetic Phosphate-specific Transporter Operon

This part enables the inducible expression of a synthetically designed (codon optimized and re-factored) Accumulibacter Phosphate-specific transporter (the amino acid sequence has been derived from the Accumulibacter phosphatis SK-1 strain).

Usage and Biology

Phosphate-specific transporters (Pst) are comprised of four domains. A periplasmic substrate-binding protein PstS, two permease proteins PstC and PstA (providing a translocation pathway) and PstB (an ATP-ase that energizes phosphate transport). Normally Pst's expresssion is induced under low concentrations of phosphate in the environment. This construct is IPTG-inducible.

The known Candidatus Accumulibacter phosphatis strains are considered Polyphosphate-Accumulating Organisms (PAOs). They were first discovered in Activated Sludge performing Enhanced Biological Phosphorus Removal in wastewater treatment facilities. Those organisms are capable of accumulating more than two times higher percentage of their dry weight in phosphorus than their non-PAO counterparts. Unfortunately, Accumulibacter has so far proven unculturable which makes it difficult to examine its genetics and biochemistry. However, metagenomic sequences from enriched cultures have been obtained in recent years and this has enabled the production of phylogenetic analysis and mostly-complete genomic sequence assemblies. iGEM York 2015 decided to tap into the PAOs potential and systematically test Accumulibacter phosphate-uptake and storage related enzymes. Rather than attempt to clone the genes from Activated sludge, iGEM York 2015 designed chemically synthesized DNA based on the metagenomic sequence samples available.

Sequence and Features

Assembly Compatibility:

10
COMPATIBLE WITH RFC[10]
12
INCOMPATIBLE WITH RFC[12]
Illegal NotI site found at 2133
21
INCOMPATIBLE WITH RFC[21]
Illegal BglII site found at 529
23
COMPATIBLE WITH RFC[23]
25
INCOMPATIBLE WITH RFC[25]
Illegal NgoMIV site found at 655
Illegal NgoMIV site found at 716
Illegal NgoMIV site found at 1732
Illegal NgoMIV site found at 2305
Illegal AgeI site found at 164
Illegal AgeI site found at 2762
1000
INCOMPATIBLE WITH RFC[1000]
Illegal BsaI site found at 1417
Illegal BsaI site found at 2383
Illegal SapI site found at 1223
Illegal SapI.rc site found at 3169

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+Phosphate-specific transporters (Pst) are comprised of four domains. A periplasmic substrate-binding protein PstS, two permease proteins PstC and PstA (providing a translocation pathway) and PstB (an ATP-ase that energizes phosphate transport). Normally Pst's expresssion is induced under low concentrations of phosphate in the environment. This construct is IPTG-inducible.
+The known Candidatus ''Accumulibacter phosphatis'' strains are considered Polyphosphate-Accumulating Organisms (PAOs). They were first discovered in Activated Sludge performing Enhanced Biological Phosphorus Removal in wastewater treatment facilities. Those organisms are capable of accumulating more than two times higher percentage of their dry weight in phosphorus than their non-PAO counterparts. Unfortunately, ''Accumulibacter'' has so far proven unculturable which makes it difficult to examine its genetics and biochemistry. However, metagenomic sequences from enriched cultures have been obtained in recent years and this has enabled the production of phylogenetic analysis and mostly-complete genomic sequence assemblies. iGEM York 2015 decided to tap into the PAOs potential and systematically test ''Accumulibacter'' phosphate-uptake and storage related enzymes. Rather than attempt to clone the genes from Activated sludge, iGEM York 2015 designed chemically synthesized DNA based on the metagenomic sequence samples available.
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