Difference between revisions of "Part:BBa K3806016:Design"
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===Design Notes===
 
===Design Notes===
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This biobrick represents the DNA template for the expression of the <i>lacZ</i> reporter gene in a T7 based cell-free expression system. Expression is regulated by a theophylline binding aptazyme. The dsRNA sequence was PCR amplified from BBa_K3806014, with a primer that changes the C and T (U in the aptazyme) nucleotides of the antisense strand into G and A, respectively, resulting in a partial mismatch to the RBS (Fig. 1).
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This biobrick represents the DNA template for the expression of the <i>lacZ</i> reporter gene in a T7 based cell-free expression system. Expression is regulated by a theophylline binding aptazyme [1]. The dsRNA sequence was PCR amplified from BBa_K3806014, with a primer that changes the C and T (U in the aptazyme) nucleotides of the antisense strand into G and A, respectively, resulting in a partial mismatch to the RBS (Fig. 1).
  
 
Fwd primer: 5’ AATTTAATACGACTCACTATAGGGAAACAAACAAA<b>GA</b>CCTTGCTGTCACCGGAATG 3’
 
Fwd primer: 5’ AATTTAATACGACTCACTATAGGGAAACAAACAAA<b>GA</b>CCTTGCTGTCACCGGAATG 3’
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Rv primer: 5’ CACACAGGAAACAGCTATGACCATG 3’
 
Rv primer: 5’ CACACAGGAAACAGCTATGACCATG 3’
  
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===References===
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==References==
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[1] Townshend, B., Xiang, J. S., Manzanarez, G., Hayden, E. J. and Smolke, C. (2021). A multiplexed, automated evolution pipeline enables scalable discovery and characterization of biosensors. Nat Commun, 12, 1437.
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Revision as of 00:34, 21 October 2021


Theophylline-binding aptazyme regulating lacZ expression (semi-cRBS). With T7 promoter.


Assembly Compatibility:
  • 10
    COMPATIBLE WITH RFC[10]
  • 12
    COMPATIBLE WITH RFC[12]
  • 21
    INCOMPATIBLE WITH RFC[21]
    Illegal BamHI site found at 3204
  • 23
    COMPATIBLE WITH RFC[23]
  • 25
    INCOMPATIBLE WITH RFC[25]
    Illegal NgoMIV site found at 2321
  • 1000
    COMPATIBLE WITH RFC[1000]


Design Notes

This biobrick represents the DNA template for the expression of the lacZ reporter gene in a T7 based cell-free expression system. Expression is regulated by a theophylline binding aptazyme [1]. The dsRNA sequence was PCR amplified from BBa_K3806014, with a primer that changes the C and T (U in the aptazyme) nucleotides of the antisense strand into G and A, respectively, resulting in a partial mismatch to the RBS (Fig. 1).

Fwd primer: 5’ AATTTAATACGACTCACTATAGGGAAACAAACAAAGACCTTGCTGTCACCGGAATG 3’

Rv primer: 5’ CACACAGGAAACAGCTATGACCATG 3’



References

  1. [1] Townshend, B., Xiang, J. S., Manzanarez, G., Hayden, E. J. and Smolke, C. (2021). A multiplexed, automated evolution pipeline enables scalable discovery and characterization of biosensors. Nat Commun, 12, 1437.