Team:MIT phage background

From 2010.igem.org

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<img style="float: left" src="https://static.igem.org/mediawiki/2010/e/ed/P3_image.jpg" height=400px title="Image from: Arap, Phage Display Technology – Applications and Innovations, 2005">
<img style="float: left" src="https://static.igem.org/mediawiki/2010/e/ed/P3_image.jpg" height=400px title="Image from: Arap, Phage Display Technology – Applications and Innovations, 2005">
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<b>PHAGE DISPLAY</b>
 
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M13 bacteriophage is a long, filamentous virus that infects bacteria via interactions with the F-pilus.  By genetically modifying the phage, fusion proteins can be created that become integrated into the phage coat in a defined manner.  Usually these new proteins are linked to the pIII or pVIII coat proteins (though PVII or PIX can be used).  Thus, novel proteins can be displayed on the phage coat.  This has historically been used as a mechanism for screening libraries of proteins or peptides (e.g., antibodies) for binding to a specific substrate or ligand of interest.  In a process called panning, a population of phage is iteratively enriched for those that bind a substrate.  Phage with proteins that do not bind are washed away and the remaining phage can then be amplified through infection.
 
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<b>HYPERPHAGE</b>
 
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Hyperphage is a plasmid with the gene for pIII truncated.  The pIII protein is required for phage exit from the host cell membrane; phage without a proper pIII grow into long fibril-like structures called polyphage.  The goal is to crosslink these polyphage.
 
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<b>M13 ANATOMY</b>
<b>M13 ANATOMY</b>
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M13 bacteriophage is a long, filamentous virus that infects bacteria via interactions with the F-pilus.  Below is a list of genes:
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PV – dimerizes and binds ssDNA, packaging it into a rod for assembly</li></ul>
PV – dimerizes and binds ssDNA, packaging it into a rod for assembly</li></ul>
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List of M13 genes:
 
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<img src="https://static.igem.org/mediawiki/2010/b/be/M13_parts.png" title="Image from: Barbas III et al. Phage Display: a Laboratory Manual. 2004." height=200px>
<img src="https://static.igem.org/mediawiki/2010/b/be/M13_parts.png" title="Image from: Barbas III et al. Phage Display: a Laboratory Manual. 2004." height=200px>
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Termination: PIII/PXI are added to the end of the particle
Termination: PIII/PXI are added to the end of the particle
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<br>
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<b>PHAGE DISPLAY</b>
 +
<br>
 +
By genetically modifying the phage, fusion proteins can be created that become integrated into the phage coat in a defined manner.  Usually these new proteins are linked to the pIII or pVIII coat proteins (though PVII or PIX can be used).  Thus, novel proteins can be displayed on the phage coat.  This has historically been used as a mechanism for screening libraries of proteins or peptides (e.g., antibodies) for binding to a specific substrate or ligand of interest.  In a process called panning, a population of phage is iteratively enriched for those that bind a substrate.  Phage with proteins that do not bind are washed away and the remaining phage can then be amplified through infection.
 +
<br><br>
 +
<b>HYPERPHAGE</b>
 +
<br>
 +
Hyperphage is a plasmid with the gene for pIII truncated.  The pIII protein is required for phage exit from the host cell membrane; phage without a proper pIII grow into long fibril-like structures called polyphage.  The goal is to crosslink these polyphage.
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<div style="text-align:center">
<div style="text-align:center">
&larr; <a href="https://2010.igem.org/Team:MIT_phage">Introduction</a>
&larr; <a href="https://2010.igem.org/Team:MIT_phage">Introduction</a>

Revision as of 00:49, 25 October 2010

hairy cells and polymerizing phage - background

M13 ANATOMY
M13 bacteriophage is a long, filamentous virus that infects bacteria via interactions with the F-pilus. Below is a list of genes:
  • PIII/PVI (pointed end) – mediate termination of assembly and particle release from the membrane. About five copies.
  • PVII/PIX (blunt end) – small coat proteins. About five copies.
  • PVIII – major coat protein. About 2700 copies.
  • PII – nicking enzyme, allows for replication to occur
  • PX – last third of PII; may inhibit PII in a regulatory manner
  • PIV – channel for phage exit
  • PI – may hydrolyze ATP to promote assembly
  • PXI – last third of PI, may interact with PI to form a channel that works with PIV
  • PV – dimerizes and binds ssDNA, packaging it into a rod for assembly




M13 ASSEMBLY

  • Initiation: PVII/PIX and PVII interaction with the DNA packaging signal. This is mediated by PI.
  • Elongation: DNA is extruded with PV being replaced by PVIII. This is mediated by PI. The particle elongates through the PIV channel.
  • Termination: PIII/PXI are added to the end of the particle

PHAGE DISPLAY
By genetically modifying the phage, fusion proteins can be created that become integrated into the phage coat in a defined manner. Usually these new proteins are linked to the pIII or pVIII coat proteins (though PVII or PIX can be used). Thus, novel proteins can be displayed on the phage coat. This has historically been used as a mechanism for screening libraries of proteins or peptides (e.g., antibodies) for binding to a specific substrate or ligand of interest. In a process called panning, a population of phage is iteratively enriched for those that bind a substrate. Phage with proteins that do not bind are washed away and the remaining phage can then be amplified through infection.

HYPERPHAGE
Hyperphage is a plasmid with the gene for pIII truncated. The pIII protein is required for phage exit from the host cell membrane; phage without a proper pIII grow into long fibril-like structures called polyphage. The goal is to crosslink these polyphage.

← Introduction       Design →

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