Difference between revisions of "Part:BBa K2984000"
Line 28: Line 28:
 
<partinfo>BBa_K2984000 parameters</partinfo>
 
<partinfo>BBa_K2984000 parameters</partinfo>
 
<!-- -->
 
<!-- -->
 +
 +
 +
 +
==References==
 +
<ol>
 +
<li>
 +
Rasala, B. A., Lee, P. A., Shen, Z., Briggs, S. P., Mendez, M., & Mayfield, S. P. (2012). Robust expression and secretion of Xylanase1 in Chlamydomonas reinhardtii by fusion to a selection gene and processing with the FMDV 2A peptide. PloS one, 7(8), e43349.
 +
</li>
 +
</ol>

Revision as of 20:15, 18 October 2019


3'UTR, Secretion signal of Arylsulfatase ARS B2-B2

This vector is a part of the Chlamy-HUB-Collection. For the secretion of a protein in Chlamydomonas reinhardtii it is necessary that this protein is attached to a signal sequence that targets it to the secretory pathway and eventually into the medium. It has been demonstrated that the Arylsulfatase 1 (ARS1) is not only able to direct protein synthesis to the secretory pathway, it was also shown that it is able to secrete heterologous proteins (Rasala et al., 2012). The amino acid sequence of the signal peptide of ARS1 is MHARKMGALAVLAVACLAAVASVAHA.



Usage and Biology

Sequence and Features


Assembly Compatibility:
  • 10
    COMPATIBLE WITH RFC[10]
  • 12
    COMPATIBLE WITH RFC[12]
  • 21
    COMPATIBLE WITH RFC[21]
  • 23
    COMPATIBLE WITH RFC[23]
  • 25
    COMPATIBLE WITH RFC[25]
  • 1000
    COMPATIBLE WITH RFC[1000]

Blue & Green Screening

The effect of the arylsulfatase secretion signal is used for screening of postive Cas9 incision in the SNRK locus in C.reinhardtii. The target site lies within a regultory unit, inhibiting the expression of the arylsulfatse. When cut, expression of the enzyme rises. Outside the cell the acitivity of the enzyme turns a supplemented color into blue proving the functionallity of the signal.

colonies_total
Fig.1 - C. reinahrdtii colonies on TAP-Agar plate. The blue colonies indicate a successfull incision inside the SNRK locus which regulates the expression of the arylsulfatase. The blue color is generated through the activity of the arylsulfatase outside the cells, verifiying the functionality of the ARS secretion signal.



References

  1. Rasala, B. A., Lee, P. A., Shen, Z., Briggs, S. P., Mendez, M., & Mayfield, S. P. (2012). Robust expression and secretion of Xylanase1 in Chlamydomonas reinhardtii by fusion to a selection gene and processing with the FMDV 2A peptide. PloS one, 7(8), e43349.