Difference between revisions of "Part:BBa K1399010"

(Created page with " == Plac-RFP(AAV) == Lactose/IPTG inducible promoter with RFP reporter tagged with AAV-ssrA degradation tag followed by terminator. The tagged RFP is actively degraded within ce...")
 
m (→‎Usage and Biology)
 
(6 intermediate revisions by 2 users not shown)
Line 1: Line 1:
−
 
 
== Plac-RFP(AAV) ==
 
== Plac-RFP(AAV) ==
  
 
Lactose/IPTG inducible promoter with RFP reporter tagged with AAV-ssrA degradation tag followed by terminator.
 
Lactose/IPTG inducible promoter with RFP reporter tagged with AAV-ssrA degradation tag followed by terminator.
−
The tagged RFP is actively degraded within cell, thus it provides better temporal resolution of red fluorescence and promoter activity.  
+
The tagged RFP is actively degraded within cell (Figure 1), thus it provides better temporal resolution of red fluorescence and promoter activity.
−
Part was assembled using BrickClip assembly (BBF RFC 104) using other biobrick parts as templates. BrickClip assembly is a special case of more general Paperclip assembly method.[1]
+
 
 +
[[File:EDiGEM14-SsrA degron mediated degradation.png|400px|thumb|center|'''Figure 1 SsrA degron mediated protein degradation.''' ('''A''') Any version of SsrA tags can be attached to any protein of interest (e.g. RFP or GFP). ('''B''') The tag is recognized by ClpX unfoldase forming a complex with ClpP protease and the tagged protein gets degraded.]]
 +
 
 +
== Usage and Biology ==
  
 +
[http://2017.igem.org/Team:Kingsborough_NY Team Kingsborough NY 2017] contributed to the characterization of this part by showing decreased fluorescence when expressed either in a higher salt media - such as LB with 3% sodium chloride - or E. coli that lacks tmRNA, the principal component of the cell's ribosome rescue system. Cells without tmRNA will have less native ssrA substrates, and this may lead to increased degradation of ssrA tagged RFP. For more information and data, see the experience page or [http://2017.igem.org/Team:Kingsborough_NY/RFP visit our Wiki]<br>
 +
(<small>--[[User:djcamenares|djcamenares]] 17:56, 27 October 2017 (UTC) </small>)
  
−
[1] Trubitsyna, M., Michlewski, G., Cai, Y., Elfick, A. & French, C. E. PaperClip: rapid multi-part DNA assembly from existing libraries. Nucleic Acids Res. 1–8 (2014). doi:10.1093/nar/gku829
+
<!-- -->
 +
<span class='h3bb'>Sequence and Features</span>
 +
<partinfo>BBa_K1399010 SequenceAndFeatures</partinfo>

Latest revision as of 19:55, 19 November 2017

Plac-RFP(AAV)

Lactose/IPTG inducible promoter with RFP reporter tagged with AAV-ssrA degradation tag followed by terminator. The tagged RFP is actively degraded within cell (Figure 1), thus it provides better temporal resolution of red fluorescence and promoter activity.

Figure 1 SsrA degron mediated protein degradation. (A) Any version of SsrA tags can be attached to any protein of interest (e.g. RFP or GFP). (B) The tag is recognized by ClpX unfoldase forming a complex with ClpP protease and the tagged protein gets degraded.

Usage and Biology

[http://2017.igem.org/Team:Kingsborough_NY Team Kingsborough NY 2017] contributed to the characterization of this part by showing decreased fluorescence when expressed either in a higher salt media - such as LB with 3% sodium chloride - or E. coli that lacks tmRNA, the principal component of the cell's ribosome rescue system. Cells without tmRNA will have less native ssrA substrates, and this may lead to increased degradation of ssrA tagged RFP. For more information and data, see the experience page or [http://2017.igem.org/Team:Kingsborough_NY/RFP visit our Wiki]
(--djcamenares 17:56, 27 October 2017 (UTC) )

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
    INCOMPATIBLE WITH RFC[25]
    Illegal AgeI site found at 781
    Illegal AgeI site found at 893
  • 1000
    COMPATIBLE WITH RFC[1000]