DNA

Part:BBa_K4621083

Designed by: Ruikang Chen   Group: iGEM23_SCUT-China   (2023-10-05)


CRISPRi-441

This CRISPRi fragment contains the dCas9 and sgRNA necessary for the CRISPRi system. The CRISPRi system is suitable for a variety of Streptomyces.[1] In this study, it was used to inhibit the expression of gabT gene in SCUT-3 to produce more GABA.


Usage and Biology

Due to the limited types of plasmids available for SCUT-3, we inserted the CRISPRi-441 fragment into the previously constructed plasmid by homologous recombination to achieve simultaneous stable expression of multiple target genes.

Reference

[1] Zhao Y, Li L, Zheng G, Jiang W, Deng Z, Wang Z, Lu Y. CRISPR/dCas9-Mediated Multiplex Gene Repression in Streptomyces. Biotechnol J. 2018 Sep;13(9):e1800121. doi: 10.1002/biot.201800121. Epub 2018 Jul 4. PMID: 29862648.


Sequence and Features


Assembly Compatibility:
  • 10
    INCOMPATIBLE WITH RFC[10]
    Illegal SpeI site found at 4530
    Illegal PstI site found at 2424
    Illegal PstI site found at 2658
    Illegal PstI site found at 3870
  • 12
    INCOMPATIBLE WITH RFC[12]
    Illegal NheI site found at 4507
    Illegal SpeI site found at 4530
    Illegal PstI site found at 2424
    Illegal PstI site found at 2658
    Illegal PstI site found at 3870
  • 21
    INCOMPATIBLE WITH RFC[21]
    Illegal BglII site found at 463
    Illegal BglII site found at 1258
    Illegal BglII site found at 4027
    Illegal BamHI site found at 203
    Illegal BamHI site found at 1552
    Illegal BamHI site found at 4495
    Illegal BamHI site found at 4700
    Illegal XhoI site found at 876
    Illegal XhoI site found at 3222
  • 23
    INCOMPATIBLE WITH RFC[23]
    Illegal SpeI site found at 4530
    Illegal PstI site found at 2424
    Illegal PstI site found at 2658
    Illegal PstI site found at 3870
  • 25
    INCOMPATIBLE WITH RFC[25]
    Illegal SpeI site found at 4530
    Illegal PstI site found at 2424
    Illegal PstI site found at 2658
    Illegal PstI site found at 3870
    Illegal NgoMIV site found at 1820
    Illegal NgoMIV site found at 2191
    Illegal NgoMIV site found at 2394
  • 1000
    INCOMPATIBLE WITH RFC[1000]
    Illegal BsaI.rc site found at 3402
    Illegal BsaI.rc site found at 4281
    Illegal SapI site found at 794
    Illegal SapI site found at 2084
    Illegal SapI.rc site found at 3281


[edit]
Categories
Parameters
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