Difference between revisions of "Part:BBa K2460008"
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This is a recombination site attB for TG1 integrase ([[Part:BBa_K2460007|BBa_K2460007]]). When used with phiBT1 attP ([[Part:BBa_K2460009|BBa_K2460009]]) and TG1 integrase, it can flip a DNA sequence flanked by attB and attP sites, as well as integrating a circular DNA with a attB/P site into a linear/circular DNA with a attP/B site.
 
This is a recombination site attB for TG1 integrase ([[Part:BBa_K2460007|BBa_K2460007]]). When used with phiBT1 attP ([[Part:BBa_K2460009|BBa_K2460009]]) and TG1 integrase, it can flip a DNA sequence flanked by attB and attP sites, as well as integrating a circular DNA with a attB/P site into a linear/circular DNA with a attP/B site.
 
For more information about kinetic characterization and the orthogonality between the integrase and some other serine recombinase, go to [[Part:BBa_K2460007|TG1 integrase]].
 
For more information about kinetic characterization and the orthogonality between the integrase and some other serine recombinase, go to [[Part:BBa_K2460007|TG1 integrase]].
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=GENAS_China 2019=
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*'''Group:''' GENAS_China 2019
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We use this part and [[Part:BBa_K2460009|BBa_K2460009]](placed in the same deriction) as funcitional elements in [[Part:BBa_K3254005|BBa_K3254005]], and they worked as expected, see below.
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==Genetic design==
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The composition and principle of the experimental system are indicated below. More details can be seen on the pages of [[Part:BBa_K3254013|BBa_K3254013]] and [[Part:BBa_K3254005|BBa_K3254005]].
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[[File:T--GENAS_China--excision_with_backbone.PNG|200px|thumb|left| ]]
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==Experimental Setup==
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The reporter plasmid contained this part and [[Part:BBa_K2460009|BBa_K2460009]] were co-transferred into E.coli DH5α host with 6 integrase generator plasmids.
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Then single colonies were inoculated into M9 supplemented medium for overnight growth. Then, the cell cultures were diluted 1000-fold with M9 supplemented medium with 500 μM IPTG inducer and growth for another 20 hours. All incubations were carried out using a Digital Thermostatic Shaker maintained at 37 °C and 1000 rpm, using flat-bottom 96-well plates sealed with sealing film. Finally, The cultures were sampled for genotype PCR testing.
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==Results==
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IBR-G22 was the plasmid with TG1 attB and attP sites.
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The result indicates that the TG1 att sites can only be recombined by TG1 integrase.
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The recombined sequence is GATCAGCTCCGCGGGCAAGACCGTGCTCTTACCCAGTTGGGCGGGA.
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[[File:T--GENAS_China--orthogonality_test.png]]
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Revision as of 11:10, 19 October 2019


TG1 attB

This is a recombination site attB for TG1 integrase (BBa_K2460007). When used with phiBT1 attP (BBa_K2460009) and TG1 integrase, it can flip a DNA sequence flanked by attB and attP sites, as well as integrating a circular DNA with a attB/P site into a linear/circular DNA with a attP/B site. For more information about kinetic characterization and the orthogonality between the integrase and some other serine recombinase, go to TG1 integrase.

GENAS_China 2019

  • Group: GENAS_China 2019

We use this part and BBa_K2460009(placed in the same deriction) as funcitional elements in BBa_K3254005, and they worked as expected, see below.

Genetic design

The composition and principle of the experimental system are indicated below. More details can be seen on the pages of BBa_K3254013 and BBa_K3254005.

T--GENAS China--excision with backbone.PNG

Experimental Setup

The reporter plasmid contained this part and BBa_K2460009 were co-transferred into E.coli DH5α host with 6 integrase generator plasmids. Then single colonies were inoculated into M9 supplemented medium for overnight growth. Then, the cell cultures were diluted 1000-fold with M9 supplemented medium with 500 μM IPTG inducer and growth for another 20 hours. All incubations were carried out using a Digital Thermostatic Shaker maintained at 37 °C and 1000 rpm, using flat-bottom 96-well plates sealed with sealing film. Finally, The cultures were sampled for genotype PCR testing.

Results

IBR-G22 was the plasmid with TG1 attB and attP sites. The result indicates that the TG1 att sites can only be recombined by TG1 integrase. The recombined sequence is GATCAGCTCCGCGGGCAAGACCGTGCTCTTACCCAGTTGGGCGGGA.

T--GENAS China--orthogonality test.png


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
    COMPATIBLE WITH RFC[1000]