Part:BBa_K3002018
Sp20 8xHis tag for Chlamydomonas reinhardtii (Phytobrick)
This basic part contains the sequence of the sp20 8xHis-tag (B5) and was built as a part of the Kaiser Collection. Combined with a secretion signal of the Kaiser Collection like BBa_K3002007 (cCA (B2)), BBa_BBa_K3002008 (GLE (B2)) or BBa_3002009 (ARS (B2)), an appropriate promoter (BBa_K3002001 (PSAD promoter (A1-B1)) or BBa_K3002031 (PAR promoter (A1-B1))) and a terminator (e.g. BBa_BBa_K3002006(RPL23 terminator)) your target protein is secreted efficiently and can be purified via a nickel-NTA column.
The SP20 tag improves the secretion of the enzymes by increasing the glycosylation. It is a repetition of Prolin and Serin, whereby Prolin is converted into hydroxyproline and then glycolyzed. This was necessary to secrete a measurable quantity of PETase and to improve the secretion of the MHETase.
The Kaiser Collection
We are proud to present our very own MoClo part collection for C. reinhardtii - the Kaiser collection.
These 20 Parts are specifically designed and codon optimized for Chlamydomonas. Among them are regulatory elements, antibiotic resistances, resistance cassettes, secretion signals and tags. These parts were tested and optimized thoroughly and we can guarantee that they work 100%. With these, expression and secretion in Chlamy will be a success. Because this is a MoClo collection, the parts are highly standardized for worldwide application. The combination with other part collections works fast and easy. While in MoClo, nomenclature is a bit different from the iGEM BioBricks, it is quickly explained:
Level 0 parts are equivalent to basic parts, e.g. Promoters, coding sequences, etc.
Level 1 parts are combinations of basic parts and usually form functional transcription units.
Level 2 parts are combinations of Level 1 parts, in case you want to transfer multiple transcription units at once. For example, you can pair your gene of interest with a selection marker.
The great thing about the Kaiser Collection and MoClo is that the ligation works in a one pot, one step reaction, as the Type IIs restriction enzymes cut out their own recognition sites. This way, multiple constructs can be combined linearly in a fixed order to create complex structures. This is ensured by the standardized overlaps that assign the parts one of 10 positions in the final constructs. After trying MoClo once, you won’t go back to traditional ligation. It is incredibly easy and reliable. For this reason, we believe that our Kaiser Collection will strike a significant chord, as the future lies in standardized, easy to use methods such as MoClo. Visit our part collection site to get an overview over all parts of the Kaiser Collection
Sequence and Features
- 10COMPATIBLE WITH RFC[10]
- 12COMPATIBLE WITH RFC[12]
- 21COMPATIBLE WITH RFC[21]
- 23COMPATIBLE WITH RFC[23]
- 25COMPATIBLE WITH RFC[25]
- 1000COMPATIBLE WITH RFC[1000]
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