Difference between revisions of "Part:BBa K1963010"

 
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<partinfo>BBa_K1963010 short</partinfo>
 
<partinfo>BBa_K1963010 short</partinfo>
  
This is a multi-sequence system for small RNA (sRNA) production in an E. coli host - or especially with co-expression of E. coli Hfq BBa_K1963000. The system is based on the BBa_K1963002 template and has the proD strong promoter, followed by a short antisense sequence targeting the RBS and 5' end of the fliC transcript, followed by the E. coli micC hairpin, followed by the strong terminator sequence T1/TE. The anti fliC sRNA is placed after C-193. The system should impair motility of E. coli.
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This is a multi-sequence system for small RNA (sRNA) production in an <i>E. coli</i> host - or especially with co-expression of <i>E. coli</i> Hfq <partinfo>BBa_K1963000</partinfo>. The system is based on the <partinfo>BBa_K1963002</partinfo> template and has the <i>proD</i> strong promoter, followed by a short antisense sequence targeting the RBS and 5' end of the <i>E. coli</i> <i>fliC</i> transcript, followed by the <i>E. coli micC</i> hairpin, followed by the strong terminator sequence T1/TE. The anti RBS-<i>fliC</i> sRNA is placed after C-193. The system should impair motility of <i>E. coli</i> as a proof-of-concept.
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<b>Note that this part differs from <partinfo>BBa_K1963004</partinfo> slightly in that the sRNA for this one covers the <i>fliC</i> RBS</b>
  
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===Usage and Biology===
 
===Usage and Biology===
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The motile <i>E. coli</i> host strain MG1655 was tranformed with <partinfo>BBa_K1963010</partinfo> and subjected to a plate-based swimming assay (Figure 1). The <partinfo>BBa_K1963010</partinfo> contains a sRNA that will target the RBS and initial codons of the <i>fliC</i> transcript that encodes flagellin - the principle component of the flagellum. The data related to this biobrick are labelled <b><i>spiRNA-fliC-RBS-CDS</i></b> in Figure 1.
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Experimental procedures needed.
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https://static.igem.org/mediawiki/parts/f/f6/Ecoliflicfrulhuq2.png
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<b>Figure 1. sRNA Devices can Inhibit Swimming in an <i>E. coli</i> Test System.</b>
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Photographs representative of motility assays with MG1655 transformed with <partinfo>BBa_K1963002</partinfo> (control), <partinfo>BBa_K1963004</partinfo> (<i>spiRNA-fliC-CDS</i>)and <partinfo>BBa_K1963010</partinfo> (<i>spiRNA-fliC-RBS-CDS</i>). Bar graph showing average swimming/spreading area and standard deviation of replicates.
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Latest revision as of 15:20, 23 October 2016


An sRNA system for down-regulation of E. coli fliC encoding flagellin

This is a multi-sequence system for small RNA (sRNA) production in an E. coli host - or especially with co-expression of E. coli Hfq BBa_K1963000. The system is based on the BBa_K1963002 template and has the proD strong promoter, followed by a short antisense sequence targeting the RBS and 5' end of the E. coli fliC transcript, followed by the E. coli micC hairpin, followed by the strong terminator sequence T1/TE. The anti RBS-fliC sRNA is placed after C-193. The system should impair motility of E. coli as a proof-of-concept.

Note that this part differs from BBa_K1963004 slightly in that the sRNA for this one covers the fliC RBS

Usage and Biology

The motile E. coli host strain MG1655 was tranformed with BBa_K1963010 and subjected to a plate-based swimming assay (Figure 1). The BBa_K1963010 contains a sRNA that will target the RBS and initial codons of the fliC transcript that encodes flagellin - the principle component of the flagellum. The data related to this biobrick are labelled spiRNA-fliC-RBS-CDS in Figure 1.

Experimental procedures needed.



Ecoliflicfrulhuq2.png

Figure 1. sRNA Devices can Inhibit Swimming in an E. coli Test System. Photographs representative of motility assays with MG1655 transformed with BBa_K1963002 (control), BBa_K1963004 (spiRNA-fliC-CDS)and BBa_K1963010 (spiRNA-fliC-RBS-CDS). Bar graph showing average swimming/spreading area and standard deviation of replicates.


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
    INCOMPATIBLE WITH RFC[1000]
    Illegal BsaI.rc site found at 101