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. 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.