Coding

Part:BBa_K2876005

Designed by: Isaac Justice   Group: iGEM18_Stanford   (2018-06-22)


A fusion of a RNAPolIII alpha subunit and antiAFB1-ScFv1

A fusion of a RNAPolIII alpha subunit and an aflatoxin sensing single chain antibody variable region (antiAFB1-ScFv1). Used in conjunction with BBa_K2876003 (https://parts.igem.org/Part:BBa_K2876005) and BBa_K2876000 (https://parts.igem.org/Part:BBa_K2876000) to create for an E. coli two hybrid system capable of detecting aflatoxin.

For our small molecule test case we chose to detect aflatoxin, a small molecule produced by Aspergillus fungi species that infects many agricultural crops, including cannabis.2,3 Aflatoxin also causes a number of health problems in people when inhaled or ingested by immunocompromised patients.7 We fused two single chain antibodies targeting aflatoxin (BBa_K2876005 and BBa_K2876003) to the lambda repressor and alpha subunit of RNA-polymerase III (BBa_K2876002) described in Dove, Joung, and Hochschild in order to create two fusion proteins capable of activating transcription in the presence of aflatoxin (Figure 4 small molecule). For our reporter we used the previously described pOL2-62 from Dove, Joung, and Hochschild fused to a RFP. See more information about our reporter below.

Sequence and Features


Assembly Compatibility:
  • 10
    COMPATIBLE WITH RFC[10]
  • 12
    INCOMPATIBLE WITH RFC[12]
    Illegal NotI site found at 8
    Illegal NotI site found at 766
  • 21
    COMPATIBLE WITH RFC[21]
  • 23
    COMPATIBLE WITH RFC[23]
  • 25
    INCOMPATIBLE WITH RFC[25]
    Illegal NgoMIV site found at 480
  • 1000
    COMPATIBLE WITH RFC[1000]


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Categories
Parameters
None