Part:BBa_K3645011

Target-AID (CBE)

Contains the full CDS of Target-AID, whose Cas9 part was replace with our lab's dCas9.

Sequence and Features

Assembly Compatibility:

• 10
  COMPATIBLE WITH RFC[10]
• 12
  INCOMPATIBLE WITH RFC[12]

  Illegal NheI site found at 1099
  
• 21
  INCOMPATIBLE WITH RFC[21]

  Illegal BglII site found at 4775
  Illegal BamHI site found at 3378
  Illegal XhoI site found at 4384
  
• 23
  COMPATIBLE WITH RFC[23]
• 25
  COMPATIBLE WITH RFC[25]
• 1000
  COMPATIBLE WITH RFC[1000]

Contribution From NNU-China 2021

Group: iGEM Team NNU-China 2021

Author: Yan Xu

Summary: Testing its gene editing efficiency in BL21 (DE3)

Characterization from iGEM21-NNU-China

        Cytosine base editors (CBEs) enable targeted C•G-to-T•A conversions in genomic DNA, consisting of dSpCas9, CDA, and UGI [1]. It was first registered in 2020. In order to test the editing efficiency of this composite part, we construct the dual plasmid system based on the (BBa_K3645011). We selected the cadA, maeA, and maeB genes as the testing sites, and the related pTarget plasmids were constructed. Results showed that the (BBa_K3645011) can successfully work in the BL21 (DE3), and the editing efficiency of single gene editing, double genes editing and triple genes editing can reach 85%, 56% and 25%, respectively (Fig. 1). These results provide references for future iGEM teams to choose gene-editing tools in E.coli.

Fig.1 The gene editing efficiency of the part of dCas9-CDA-UGI.

Reference

[1] Zhao D, Li J, Li S, Xin X, Hu M, Price MA, Rosser SJ, Bi C, Zhang X. Glycosylase base editors enable C-to-A and C-to-G base changes. Nature Biotechnology. 2021; 39: 35–40.