Team:Bologna

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= Project Summary =
= Project Summary =
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'''Which is our idea?'''
 
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The project aims to implement a protein synthesis regulation system in ''Escherichia coli'' that acts at translational level regardless of the target gene to be downregulated. This "general-purpose" device could allow a faster control of protein expression. Our device was named '''T-Rex''' ('''T'''rans '''R'''epressor of '''Ex'''pression). It consists of two new BioBricks, i.e. the '''Trans-repressor''' and the '''Cis-repressing'''.
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'''Our idea'''
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The aim of our project is the design of a standard device to control the synthesis of any protein of interest. This "general-purpose" device, implemented in <i>E. coli</i>, acts at the translational level to allow silencing of protein expression faster than using regulated promoters. We named this device <b>T-REX</b> (<b>T</b>rans <b>R</b>epressor of <b>Ex</b>pression).
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'''How T-REX works'''
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'''How can we achieve our goal?'''
 
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The TRANS-repressor is a non-coding DNA sequence that acts as a silencer of the CIS-repressing mRNA. In fact, the Cis-repressing sequence includes a TRANS-repressor complementary region ending with a ribosome binding site (RBS). Moreover, it is assembled upstream of the target gene coding sequence. When the TRANS-repressor and the CIS-repressing mRNAs bind together, the RBS recognition by the ribosome is prevented. Thus, the TRANS-repressor amount regulates the gene mRNA translation rate.
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The device consists of two new BioBricks:
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The TRANS-repressor sequence was determined by a computational analysis performed to minimize the interference with the genomic mRNAs and to maximize the base-pairing interaction to the CIS-repressing RNA.  
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<li><font color="#000080"><b>CIS-repressing</b></font>, to be assembled upstream of the target coding sequence.
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<li><font color="#000080"><b>TRANS-repressor</b></font>, complementary to the CIS-repressing and placed under the control of a different promoter.
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CIS-repressing and TRANS-repressor sequences were designed by [[Team:Bologna/Software#1|BASER]] software.
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Transcription of the target gene yields a mRNA strand - containing the CIS-repressing sequence at its 5' end - available for translation into protein by ribosomes (<i>see Fig. 1, left panel</i>). When the promoter controlling the TRANS coding sequence is active, it drives the transcription of an oligoribonucleotide complementary to the CIS mRNA sequence. The TRANS/CIS <b>RNA duplex</b> prevents ribosomes from binding to RBS on target mRNA, thus <b>silencing protein synthesis</b>. The amount of the TRANS-repressor regulates the rate of translation of the target mRNA (<i>see Fig. 1, right panel</i>)
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[[Image:project3b.png|center|950px|thumb|<center>Figure 1 - T-REX device</center>]]
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'''How we can test the device'''
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We developed the following genetic circuit in order to test and characterize our T-Rex device:
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In order to test and characterize our T-REX device, we developed the following genetic circuit (Fig 2):
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The T-REX device is proposed as a universal and fast switch in synthetic gene circuits.
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[[Image:circuit2OK.jpg|center|900px|thumb|<center>Figure 2 - Genetic Circuit to test CIS and TRANS' mRNA affinity</center>]]
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More details about our work in the [[Team:Bologna/Project|Project]] section.
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More details about our work are reported in the [[Team:Bologna/Project|Project]] section.
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= Acknowledgements =
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* ''' [http://www.unibo.it/Portale/default.htm University of Bologna] '''
* ''' [http://www.unibo.it/Portale/default.htm University of Bologna] '''
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* '''Cultural Association San Sebastiano'''  
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* <font color=#0000cd>'''Cultural Association San Sebastiano'''</font>
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Latest revision as of 03:12, 22 October 2009

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Project Summary


Our idea

The aim of our project is the design of a standard device to control the synthesis of any protein of interest. This "general-purpose" device, implemented in E. coli, acts at the translational level to allow silencing of protein expression faster than using regulated promoters. We named this device T-REX (Trans Repressor of Expression).


How T-REX works


The device consists of two new BioBricks:

  • CIS-repressing, to be assembled upstream of the target coding sequence.
  • TRANS-repressor, complementary to the CIS-repressing and placed under the control of a different promoter.

CIS-repressing and TRANS-repressor sequences were designed by BASER software.

Transcription of the target gene yields a mRNA strand - containing the CIS-repressing sequence at its 5' end - available for translation into protein by ribosomes (see Fig. 1, left panel). When the promoter controlling the TRANS coding sequence is active, it drives the transcription of an oligoribonucleotide complementary to the CIS mRNA sequence. The TRANS/CIS RNA duplex prevents ribosomes from binding to RBS on target mRNA, thus silencing protein synthesis. The amount of the TRANS-repressor regulates the rate of translation of the target mRNA (see Fig. 1, right panel)

Figure 1 - T-REX device



How we can test the device


In order to test and characterize our T-REX device, we developed the following genetic circuit (Fig 2):

Figure 2 - Genetic Circuit to test CIS and TRANS' mRNA affinity




More details about our work are reported in the Project section.


Acknowledgements


  • [http://www.unibo.it/Portale/default.htm University of Bologna]


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  • [http://serinar.criad.unibo.it Ser.In.Ar. Cesena]


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  • Cultural Association San Sebastiano
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