Difference between revisions of "Part:BBa K5102072"

 
 
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<partinfo>BBa_K5102072 short</partinfo>
 
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The composite part is designed for the ProgRAM molecular recording system and features a recording tape composed of a series of START codons arranged in three forward open reading frames (ORFs). Downstream of the tape, a sequence of fluorescent proteins enables visualization of the current recorded state. The design incorporates sequentially modifiable adenosine sites within the RNA tape, creating a dynamic "traffic light" system that allows for precise in vivo monitoring of recording events without disrupting cellular functions. Each adenosine corresponds to a START codon; upon deamination, this modification disrupts the codon, shifting the open reading frame by one base pair and triggering the expression of one of three distinct fluorescent proteins: miRFP670nano3, mScarlet3, mTagBFP2.
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To ensure proper protein folding, each fluorescent protein is encoded downstream of the RNA tape and preceded by a 2A peptide, which promotes ribosomal skipping during translation. This design guarantees the efficient production of fluorescent proteins without interference from upstream sequences. Additionally, eUnaG (Truong et al., 2024), a small green fluorescent protein codon-optimized for all three ORFs, is included to provide a translational-level control of total protein expression throughout the system. The composite part also contains elements to enhance mRNA expression and stability, such as the 5' CMV UTR, human beta-globin 3' UTR, and WPRE.
  
 
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Latest revision as of 00:14, 1 October 2024


pRAM_ProgRAM-recording-tape1.0

The composite part is designed for the ProgRAM molecular recording system and features a recording tape composed of a series of START codons arranged in three forward open reading frames (ORFs). Downstream of the tape, a sequence of fluorescent proteins enables visualization of the current recorded state. The design incorporates sequentially modifiable adenosine sites within the RNA tape, creating a dynamic "traffic light" system that allows for precise in vivo monitoring of recording events without disrupting cellular functions. Each adenosine corresponds to a START codon; upon deamination, this modification disrupts the codon, shifting the open reading frame by one base pair and triggering the expression of one of three distinct fluorescent proteins: miRFP670nano3, mScarlet3, mTagBFP2.

To ensure proper protein folding, each fluorescent protein is encoded downstream of the RNA tape and preceded by a 2A peptide, which promotes ribosomal skipping during translation. This design guarantees the efficient production of fluorescent proteins without interference from upstream sequences. Additionally, eUnaG (Truong et al., 2024), a small green fluorescent protein codon-optimized for all three ORFs, is included to provide a translational-level control of total protein expression throughout the system. The composite part also contains elements to enhance mRNA expression and stability, such as the 5' CMV UTR, human beta-globin 3' UTR, and WPRE.

Sequence and Features


Assembly Compatibility:
  • 10
    INCOMPATIBLE WITH RFC[10]
    Illegal SpeI site found at 18
  • 12
    INCOMPATIBLE WITH RFC[12]
    Illegal SpeI site found at 18
  • 21
    INCOMPATIBLE WITH RFC[21]
    Illegal BamHI site found at 2494
    Illegal BamHI site found at 3554
    Illegal XhoI site found at 2563
  • 23
    INCOMPATIBLE WITH RFC[23]
    Illegal SpeI site found at 18
  • 25
    INCOMPATIBLE WITH RFC[25]
    Illegal SpeI site found at 18
    Illegal NgoMIV site found at 4899
    Illegal AgeI site found at 3645
  • 1000
    COMPATIBLE WITH RFC[1000]