Coding

Part:BBa_K2560260:Design

Designed by: Daniel Marchal   Group: iGEM18_Marburg   (2018-09-30)
Revision as of 12:32, 30 September 2018 by Marchal (Talk | contribs)


mcr gene for Malonyl-CoA Reductase from Chloroflexus aurantiacus


Assembly Compatibility:
  • 10
    COMPATIBLE WITH RFC[10]
  • 12
    INCOMPATIBLE WITH RFC[12]
    Illegal NheI site found at 1932
    Illegal NheI site found at 3334
  • 21
    INCOMPATIBLE WITH RFC[21]
    Illegal BglII site found at 885
    Illegal BamHI site found at 213
  • 23
    COMPATIBLE WITH RFC[23]
  • 25
    INCOMPATIBLE WITH RFC[25]
    Illegal NgoMIV site found at 1188
    Illegal NgoMIV site found at 2005
    Illegal NgoMIV site found at 3619
    Illegal AgeI site found at 1270
    Illegal AgeI site found at 3168
  • 1000
    COMPATIBLE WITH RFC[1000]


Design Notes

The part contains the whole coding region from the mcr gene of C. aurantiacus (AY530019) without the startcodon. This region is flanked by overhangs which are Phytobrick- and MoClo-compatible and by two BsaI recognition sites (Weber et al., 2011).

It was shown that a splitted version of Mcr with separated C- and N-terminal domains increases enzyme activity (Liu et al., 2013). If these splitted genes shall be used take BBa_K2560261 and BBa_K2560262.

GGTCTCGAATG-coding_region-GCTTTGAGACC


The sequence was codonoptimized for V. natriegens ATCC 14048.

Source

Source of the part:

Genome: Chloroflexus aurantiacus strain OK-70-fl

Accession number of gene: AY530019

Accession number of encoded protein: AAS20429

mcr was codonoptimized for V. natriegens and then synthetisized and integrated into the vector BBa_K2560002 via BsmBI

References

Liu, C., Wang, Q., Xian, M., Ding, Y., Zhao, G., 2013. Dissection of Malonyl-Coenzyme A Reductase of Chloroflexus aurantiacus Results in Enzyme Activity Improvement. PLoS One 8, 1–8. https://doi.org/10.1371/journal.pone.0075554

Strauss, G., Fuchs, G., 1993. Enzymes of a novel autotrophic CO2 fixation pathway in the phototrophic bacterium Chloroflexus aurantiacus, the 3-hydroxypropionate cycle. Eur. J. Biochem. 215, 633–43.

Weber, E., Engler, C., Gruetzner, R., Werner, S., Marillonnet, S., 2011. A Modular Cloning System for Standardized Assembly of Multigene Constructs. PLoS One 6, e16765. https://doi.org/10.1371/journal.pone.0016765