Difference between revisions of "Part:BBa K3505007"

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Level 0 vector for Golden Braid assembly. The Level 0 modules are Interchangable parts in order everyone can use it as it is without any modification based on Golden Braid grammar.LacZa module as cargo for Blue White Screening between 2 BsmBI sites and 2 BtgzI sites. Resistance in Chloramphenicol and ori pMB1.
 
Level 0 vector for Golden Braid assembly. The Level 0 modules are Interchangable parts in order everyone can use it as it is without any modification based on Golden Braid grammar.LacZa module as cargo for Blue White Screening between 2 BsmBI sites and 2 BtgzI sites. Resistance in Chloramphenicol and ori pMB1.
  
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     [[Image:T--Thessaly--GBcloninggrammar.jpeg|900px|thumb|none|]]
 
     [[Image:T--Thessaly--GBcloninggrammar.jpeg|900px|thumb|none|]]
  
 
<b>Figure 1.</b> The GB2.0 grammar
 
<b>Figure 1.</b> The GB2.0 grammar
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    [[Image:T--Thessaly--pudp2.png|700px|thumb|none|]]
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<b>Figure 2.</b> pUPD2 map
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    [[Image:T--Thessaly--pudp2 map.png|700px|thumb|none|]]
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<b>Figure 3.</b> pUPD2
 
===Usage and Biology===
 
===Usage and Biology===
 
GoldenBraid (GB) is a DNA assembly strategy for Plant Synthetic Biology based on Type IIS enzymes. It is also compatible for MoClo assembly.
 
GoldenBraid (GB) is a DNA assembly strategy for Plant Synthetic Biology based on Type IIS enzymes. It is also compatible for MoClo assembly.

Revision as of 19:41, 13 October 2020


pUPD2 for Golden Braid

Level 0 vector for Golden Braid assembly. The Level 0 modules are Interchangable parts in order everyone can use it as it is without any modification based on Golden Braid grammar.LacZa module as cargo for Blue White Screening between 2 BsmBI sites and 2 BtgzI sites. Resistance in Chloramphenicol and ori pMB1.


T--Thessaly--GBcloninggrammar.jpeg

Figure 1. The GB2.0 grammar


T--Thessaly--pudp2.png

Figure 2. pUPD2 map

T--Thessaly--pudp2 map.png

Figure 3. pUPD2

Usage and Biology

GoldenBraid (GB) is a DNA assembly strategy for Plant Synthetic Biology based on Type IIS enzymes. It is also compatible for MoClo assembly. The sequences must not contain BsmBI and BsaI sites! Domestication may be done in order to vanish BsaI and BsmBI sites from the inner sequence.

>GB proposes an alternative view of modular cloning, and essentially the change is that you can infinitely build assemble new vectors by performing “braids”.[1] Using BsmBI another big advantage is the use of a single level 0 vector (pUPD and pUPD2, where pUPD2 is derived from iGEM-borne pSB1C3) for any GBpart one needs.

Then combining the desirable fragments from level 0; ‘level alpha’ (level a) cloning is succeeded creating Transcription Units (TU). Desirable TUs are combined to result in (Level Ω) cloning.

  • Using BsmBI for Level 0 modules and ‘level omega’ (Level Ω)
  • Using BsaI for ‘Level alpha’ (Level a)

But this is not the end, GoldenBraid outperforms GoldenGate (which everyone knows) because of the ability to continuously clone TUs in an “exponential” manner, compared to the linear progression GoldenGate. Bianry assembly of 2 Level Ω (Level 2 for MoClo) result in an alpha vector. Again Binary assembly of 2 alpha result in an omega vector. With this assembly you can insert step by step as many parts and as many TUs you want with high efficiency.


Sequence and Features


Assembly Compatibility:
  • 10
    INCOMPATIBLE WITH RFC[10]
    Illegal EcoRI site found at 260
    Illegal EcoRI site found at 2674
    Illegal XbaI site found at 233
    Illegal SpeI site found at 627
    Illegal PstI site found at 221
    Illegal PstI site found at 641
  • 12
    INCOMPATIBLE WITH RFC[12]
    Illegal EcoRI site found at 260
    Illegal EcoRI site found at 2674
    Illegal SpeI site found at 627
    Illegal PstI site found at 221
    Illegal PstI site found at 641
    Illegal NotI site found at 634
    Illegal NotI site found at 2680
  • 21
    INCOMPATIBLE WITH RFC[21]
    Illegal EcoRI site found at 260
    Illegal EcoRI site found at 2674
    Illegal BamHI site found at 239
    Illegal XhoI site found at 1658
    Illegal XhoI site found at 2550
  • 23
    INCOMPATIBLE WITH RFC[23]
    Illegal suffix found in sequence at 627
    Illegal EcoRI site found at 260
    Illegal EcoRI site found at 2674
    Illegal XbaI site found at 233
    Illegal PstI site found at 221
  • 25
    INCOMPATIBLE WITH RFC[25]
    Illegal EcoRI site found at 260
    Illegal EcoRI site found at 2674
    Illegal XbaI site found at 233
    Illegal SpeI site found at 627
    Illegal PstI site found at 221
    Illegal PstI site found at 641
  • 1000
    INCOMPATIBLE WITH RFC[1000]
    Illegal BsaI site found at 14
    Illegal BsaI.rc site found at 611


Functional Parameters

References

[1] Alejandro Sarrion-Perdigones, Marta Vazquez-Vilar, Jorge Palací, Bas Castelijns, Javier Forment, Peio Ziarsolo, José Blanca, Antonio Granell, Diego Orzaez (2013). “GoldenBraid 2.0: A Comprehensive DNA Assembly Framework for Plant Synthetic Biology.” Plant Physiology , 162 (3) 1618-1631; DOI: 10.1104/pp.113.217661