DNA

Part:BBa_K4621079

Designed by: Ruikang Chen   Group: iGEM23_SCUT-China   (2023-10-05)
Revision as of 11:43, 10 October 2023 by Conerrr (Talk | contribs)


CRISPRi-62

This CRISPRi fragment contains four parts : BBa _ K4621070, BBa _ K3875019, BBa _ K4621029, and BBa _ K3081104, covering 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 GQS52 _ 08040 gene in SCUT-3 to produce more ectoine and hydroxyectoine.

Usage and Biology

mechanism-of-crispri.png

Fig.1 CRISPRi system used in the study

Due to the limited types of plasmids available for SCUT-3, we inserted the CRISPRi-62 fragment into the previously constructed plasmid by homologous recombination to achieve simultaneous stable expression of multiple target genes. Subsequently, we verified the effectiveness of the CRISPRi-62 system (Pi2 in the figure below) in the fermentation of ordinary LB and shrimp shells.

fermentation-under-high-salt-environment.png Fig.2 Performance test on CRISPRi strains

fermentation-using-shrimp-shells-p.png Fig. 3 Ectoine/hydroxyectoine production using shrimp shell waste of CRISPRi strains

Sequence and Features


Assembly Compatibility:
  • 10
    INCOMPATIBLE WITH RFC[10]
    Illegal XbaI site found at 15
    Illegal SpeI site found at 4534
    Illegal PstI site found at 3
    Illegal PstI site found at 2428
    Illegal PstI site found at 2662
    Illegal PstI site found at 3874
  • 12
    INCOMPATIBLE WITH RFC[12]
    Illegal NheI site found at 4511
    Illegal SpeI site found at 4534
    Illegal PstI site found at 3
    Illegal PstI site found at 2428
    Illegal PstI site found at 2662
    Illegal PstI site found at 3874
  • 21
    INCOMPATIBLE WITH RFC[21]
    Illegal BglII site found at 467
    Illegal BglII site found at 1262
    Illegal BglII site found at 4031
    Illegal BamHI site found at 207
    Illegal BamHI site found at 1556
    Illegal BamHI site found at 4499
    Illegal XhoI site found at 880
    Illegal XhoI site found at 3226
  • 23
    INCOMPATIBLE WITH RFC[23]
    Illegal XbaI site found at 15
    Illegal SpeI site found at 4534
    Illegal PstI site found at 3
    Illegal PstI site found at 2428
    Illegal PstI site found at 2662
    Illegal PstI site found at 3874
  • 25
    INCOMPATIBLE WITH RFC[25]
    Illegal XbaI site found at 15
    Illegal SpeI site found at 4534
    Illegal PstI site found at 3
    Illegal PstI site found at 2428
    Illegal PstI site found at 2662
    Illegal PstI site found at 3874
    Illegal NgoMIV site found at 1824
    Illegal NgoMIV site found at 2195
    Illegal NgoMIV site found at 2398
  • 1000
    INCOMPATIBLE WITH RFC[1000]
    Illegal BsaI.rc site found at 3406
    Illegal BsaI.rc site found at 4285
    Illegal SapI site found at 798
    Illegal SapI site found at 2088
    Illegal SapI.rc site found at 3285


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Categories
Parameters
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