Difference between revisions of "Part:BBa K2609026"
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<h2>Usage</h2>
 
<h2>Usage</h2>
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<h3> IISc-Bangalore iGEM 2018</h3>
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<h3> IISc-Bangalore 2018</h3>
 
<p>
 
<p>
 
     We used the this Lambda RED recombination system with a T4 Endolysin gene (<a href="https://parts.igem.org/Part:BBa_K2609017">BBa_K2609017</a>) to recombine Bacteriophage T4 and create a lysis deficient phage. This lysis deficient phage triggers its host to produce a monocyte chemoattractant <i>mcp-1</i> (<a href="https://parts.igem.org/Part:BBa_K2609000">BBa_K2609000</a>).
 
     We used the this Lambda RED recombination system with a T4 Endolysin gene (<a href="https://parts.igem.org/Part:BBa_K2609017">BBa_K2609017</a>) to recombine Bacteriophage T4 and create a lysis deficient phage. This lysis deficient phage triggers its host to produce a monocyte chemoattractant <i>mcp-1</i> (<a href="https://parts.igem.org/Part:BBa_K2609000">BBa_K2609000</a>).
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<span class='h3bb'>Sequence and Features</span>
 
<span class='h3bb'>Sequence and Features</span>
 
<partinfo>BBa_K2609026 SequenceAndFeatures</partinfo>
 
<partinfo>BBa_K2609026 SequenceAndFeatures</partinfo>
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===References===
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[1] Mosberg, J. A., Lajoie, M. J., & Church, G. M. (2010). Lambda Red Recombineering in Escherichia coli Occurs Through a Fully Single-Stranded Intermediate. Genetics, 186(3), 791–799. http://doi.org/10.1534/genetics.110.120782
  
 +
[2]Sharan S K, Thomason L C, Kuznetsov S G, et al. Recombineering: a homologous recombination-based method of genetic engineering[J]. Nature protocols, 2009, 4(2): 206-223.
 +
 +
[3]Datsenko K A, Wanner B L. One-step inactivation of chromosomal genes in Escherichia coli K-12 using PCR products[J]. Proceedings of the National Academy of Sciences, 2000, 97(12): 6640-6645.
  
 
<!-- Uncomment this to enable Functional Parameter display  
 
<!-- Uncomment this to enable Functional Parameter display  

Revision as of 15:46, 11 October 2018


Lambda Red Recombinases under T7 expression system

This part generates the three Lambda ReD recombinases - exo, gam, beta - when expressed in a T7 expression strain like E. coli BL21(DE3).

Usage and Biology

Biology

The genes exo, gam and beta are a part of Bacteriophage Lambda's genome and is used in recomibination of bacteriophage DNA. The gam protein binds to the RecBCD nuclease of the host, thus protecting linear viral DNA from degradation. The protein exo is a 5' to 3' exonuclease which exposes the ends of linear dsDNA. The protein beta promotes single strand annealing and hence promotes homologous recombination. It is also important in rollin circle DNA replication which comes late in the infective cycle of the lambda phage.

Usage

IISc-Bangalore 2018

We used the this Lambda RED recombination system with a T4 Endolysin gene (BBa_K2609017) to recombine Bacteriophage T4 and create a lysis deficient phage. This lysis deficient phage triggers its host to produce a monocyte chemoattractant mcp-1 (BBa_K2609000).

Sequence and Features


Assembly Compatibility:
  • 10
    COMPATIBLE WITH RFC[10]
  • 12
    INCOMPATIBLE WITH RFC[12]
    Illegal NheI site found at 1943
  • 21
    COMPATIBLE WITH RFC[21]
  • 23
    COMPATIBLE WITH RFC[23]
  • 25
    INCOMPATIBLE WITH RFC[25]
    Illegal AgeI site found at 1460
  • 1000
    COMPATIBLE WITH RFC[1000]

References

[1] Mosberg, J. A., Lajoie, M. J., & Church, G. M. (2010). Lambda Red Recombineering in Escherichia coli Occurs Through a Fully Single-Stranded Intermediate. Genetics, 186(3), 791–799. http://doi.org/10.1534/genetics.110.120782

[2]Sharan S K, Thomason L C, Kuznetsov S G, et al. Recombineering: a homologous recombination-based method of genetic engineering[J]. Nature protocols, 2009, 4(2): 206-223.

[3]Datsenko K A, Wanner B L. One-step inactivation of chromosomal genes in Escherichia coli K-12 using PCR products[J]. Proceedings of the National Academy of Sciences, 2000, 97(12): 6640-6645.