Difference between revisions of "Part:BBa K404108"
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{|width='80%'
 
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|width=35% valign='top'|
 
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[[Image:Terminator.png]]
 
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<partinfo>BBa_K404108 parameters</partinfo>
 
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| width=50% valign='top' style='border: 1px solid black'|
 
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<partinfo>BBa_K404108 short</partinfo>
 
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* x
 
  
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[[Image:Freiburg10_VectorplasmidBricks 5.png|thumb|center|480px]]<br>
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<h3>Usage and Biology</h3>
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{| style="margin: 0px 0px 350px 20px; color: black; float: right;" cellpadding="6" cellspacing="1" border="2"
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! colspan="2" style="background:#66bbff;"|[https://parts.igem.org/Part:BBa_K404107 <i>beta-globin</i> intron]
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|-
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! colspan="2"|[[Image:Freiburg10 VectorplasmidBricks 5.png|200px]]
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|-
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|'''BioBrick Nr.'''
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|[https://parts.igem.org/Part:BBa_K404108 BBa_K404108]
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|-
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|'''RFC standard'''
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|[https://parts.igem.org/Help:Assembly_standard_10 RFC 10]
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|-
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|'''Requirement'''
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|pSB1C3
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|-
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|'''Source'''
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|pAAV_MCS
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|-
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|'''Submitted by'''
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|[http://2010.igem.org/Team:Freiburg_Bioware FreiGEM 2010]
 
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|}
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<html>
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<p style="margin-right:100px" align="justify">
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The iGEM team Freiburg provides an hGH plolyadenylation sequence within the ‘Virus Construction Kit’ due to the fact that almost every eukaryotic mRNA is processed at their 3´ and 5´end except for histone mRNAs (Millevoi et al., 2006). Pre-mRNAs contain two canonical conserved sequences. First, the polyadenylation signal “AATAAA” which is recognized by the multiprotein complex and second the GT-rich region (downstream sequence element, DSE) which is located 30 nucleotides downstream of the cleavage site. The assembled 3´end-processing machinery cleaves the mRNA transcript immediately after a CA-nucleotide therefore defining the cleavage site (Danckwardt, Hentze, & Kulozik, 2008).
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<br />
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<img src="https://static.igem.org/mediawiki/parts/f/fb/Freiburg10_Nucleotide_sequence_hGH_terminator.png" width="660"
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height="auto"/>
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<br />
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</p>
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</html>
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<br />
  
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<h3>Characterization</h3>
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[[Image:Freiburg10_VectorplasmidBricks 5.png|thumb|center|480px]]<br>
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<p style="margin-right:100px" align="justify">
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Recombinant AAV genomes were engineered containing the inverted terminal repeats (ITRs), a strong eukaryotic promoter and mVenus as gene of interest with and without the hGH terminator signal. Transduction of HT1080 cells with viral particles containing the rAAV genomes and measuring mVenus expression 24-hours post infection by flow cytometry demonstrated that transgene expression of the constructs lacking the hGH termination signal is significantly reduced. The iGEM team Freiburg_Bioware 2010 therefore suggests using the provided hGH termination signal within the Virus Construction Kit for optimal gene expression.
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</p>
  
  

Revision as of 02:42, 23 October 2010


Freiburg10 VectorplasmidBricks 5.png

Usage and Biology

beta-globin intron
Freiburg10 VectorplasmidBricks 5.png
BioBrick Nr. BBa_K404108
RFC standard RFC 10
Requirement pSB1C3
Source pAAV_MCS
Submitted by [http://2010.igem.org/Team:Freiburg_Bioware FreiGEM 2010]

The iGEM team Freiburg provides an hGH plolyadenylation sequence within the ‘Virus Construction Kit’ due to the fact that almost every eukaryotic mRNA is processed at their 3´ and 5´end except for histone mRNAs (Millevoi et al., 2006). Pre-mRNAs contain two canonical conserved sequences. First, the polyadenylation signal “AATAAA” which is recognized by the multiprotein complex and second the GT-rich region (downstream sequence element, DSE) which is located 30 nucleotides downstream of the cleavage site. The assembled 3´end-processing machinery cleaves the mRNA transcript immediately after a CA-nucleotide therefore defining the cleavage site (Danckwardt, Hentze, & Kulozik, 2008).


Characterization

Recombinant AAV genomes were engineered containing the inverted terminal repeats (ITRs), a strong eukaryotic promoter and mVenus as gene of interest with and without the hGH terminator signal. Transduction of HT1080 cells with viral particles containing the rAAV genomes and measuring mVenus expression 24-hours post infection by flow cytometry demonstrated that transgene expression of the constructs lacking the hGH termination signal is significantly reduced. The iGEM team Freiburg_Bioware 2010 therefore suggests using the provided hGH termination signal within the Virus Construction Kit for optimal gene expression.


Secondary Structure

File:Mfold-K404108-1.png

Measurement

  • [http://openwetware.org/wiki/Cconboy:Terminator_Characterization/Results How these parts were measured]