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Shen Lab DdCBE assembly kit
(Kit # 1000000212 )

Depositing Lab:   Bin Shen

The DddA-derived cytosine base editor (DdCBE) assembly kit is a collection of 192 modules harboring repeat variable di-residue (RVD) arrays for any desired target. With the components of this kit, DdCBE expression plasmids for a desired target can be obtained within 36 hours using Golden Gate assembly.

This kit will be sent as bacterial glycerol stocks in 96-well plate format.

Add to Cart
$ 640 USD + shipping

Available to academics and nonprofits only.

Original Publication

Precision modeling of mitochondrial diseases in zebrafish via DdCBE-mediated mtDNA base editing. Guo J, Zhang X, Chen X, Sun H, Dai Y, Wang J, Qian X, Tan L, Lou X, Shen B. Cell Discov. 2021 Sep 3;7(1):78. doi: 10.1038/s41421-021-00307-9. PubMed (Link opens in a new window) Article (Link opens in a new window)

Description

DdCBEs, composed of split interbacterial cytidine deaminase toxin, DddA, transcription activator-like effector (TALE), and uracil glycosylase inhibitor (UGI), targets gene editing either to nuclear genes or mitochondrial DNA (mtDNA) based on the presence of a nuclear localization signal (NLS) or mitochondrial targeting signal (MTS).

Several approaches have been developed to build TALEs using assemble custom repeat variable di-residues (RVD) arrays, most of which are time-consuming, costly, and unfriendly to ordinary laboratories. To establish a cost-effective, robust, and rapid solution suitable for mitochondria base editing, an RVD plasmid library was constructed that includes 192 modules (Figure 1a): 160 dimer RVD, 16 monomer RVD, and 16 half RVD modules.Two sets of DdCBE backbone plasmids, MTS-DdCBE and NLS-DdCBE, with mitochondrial or nuclear localization signal peptide (Figure 1b and 1c) are available as accessory plasmids. DdCBE expression plasmids, containing the desired RVD array, can be obtained within 36 hours by one-step Golden Gate assembly (Figure 1d).

Four separate diagrams labeled (a) through (d) that show the structure of the RVD library, the backbone plasmids into which the modules are cloned, and the one-step assembly of the modules into a DdCBE expression plasmid. Figure (a) shows RVD modules that are included in the kit and Figures (b) and (c) depict eight backbone plasmids into which the modules can be cloned. These are not included in the kit but are available as accessory plasmids. Figure (d) depicts how the BsaI cloning sites flanking each RVD module allow one-step assembly of multiple RVD modules into a backbone plasmid to generate a DdCBE expression vector.

Figure 1: Architecture of RVD library and DdCBE assembly. (a) The RVD plasmid library, in which each RVD module is flanked by BsaI restriction sites and cloned into a vector containing the tetracycline resistance gene. A total of 192 RVD plasmids were constructed: 160 dimer RVD, 16 monomer RVD, and 16 half RVD modules. (b, c) Backbone plasmids MTS-DdCBE and NLS-DdCBE (available as accessory plasmids) with half DddAtox split at G1333 and G1397 with mitochondrial localization signal (b) or nuclear localization signal (c). (d) One-step assembly of DdCBE. Top, plasmids containing intended RVD are cleaved using BsaI to produce sticky ends. Middle, RVD modules with sticky ends are allowed to ligate to the backbones sequentially. Bottom, the assembled DdCBE vectors contain 13.5–20.5 RVDs. Twenty-seven amino acids (aa) of the C-terminal TALE are from the backbone and 14 aa are from a half RVD module, resulting in 41 aa C-terminal TALE after assembly.

How to Cite this Kit

These plasmids were created by your colleagues. Please acknowledge the Principal Investigator, cite the article in which they were created, and include Addgene in the Materials and Methods of your future publications.

For your Materials and Methods section:

“The Shen Lab DdCBE assembly kit was a gift from Bin Shen (Addgene kit #1000000212).”

For your Reference section:

Precision modeling of mitochondrial diseases in zebrafish via DdCBE-mediated mtDNA base editing. Guo J, Zhang X, Chen X, Sun H, Dai Y, Wang J, Qian X, Tan L, Lou X, Shen B. Cell Discov. 2021 Sep 3;7(1):78. doi: 10.1038/s41421-021-00307-9. PubMed (Link opens in a new window) Article (Link opens in a new window)

Additional References

Precision modeling of mitochondrial disease in rats via DdCBE-mediated mtDNA editing. Qi X, Chen X, Guo J, Zhang X, Sun H, Wang J, Qian X, Li B, Tan L, Yu L, Chen W, Zhang L, Ma Y, Shen B. Cell Discov. 2021 Oct 19;7(1):95. doi: 10.1038/s41421-021-00325-7. PubMed (Link opens in a new window) Article (Link opens in a new window)

DdCBE mediates efficient and inheritable modifications in mouse mitochondrial genome. Guo J, Chen X, Liu Z, Sun H, Zhou Y, Dai Y, Ma Y, He L, Qian X, Wang J, Zhang J, Zhu Y, Zhang J, Shen B, Zhou F. Mol Ther Nucleic Acids. 2021 Nov 19;27:73-80. doi: 10.1016/j.omtn.2021.11.016. PubMed (Link opens in a new window) Article (Link opens in a new window)

DdCBE-mediated mitochondrial base editing in human 3PN embryos. Chen X, Liang D, Guo J, Zhang J, Sun H, Zhang X, Jin J, Dai Y, Bao Q, Qian X, Tan L, Hu P, Ling X, Shen B, Xu Z. Cell Discov. 2022 Feb 1;8(1):8. doi: 10.1038/s41421-021-00358-y. PubMed (Link opens in a new window) Article (Link opens in a new window)

Shen Lab DdCBE assembly kit - #1000000212

Resistance Color Key

Each circle corresponds to a specific antibiotic resistance in the kit plate map wells.

Inventory

Searchable and sortable table of all plasmids in kit. The Well column links to its kit plate map on the right, as well as lists the plasmid well location in its plate. The Plasmid column links to a plasmid's individual web page.

Kit Plate Map

96-well plate map for plasmid layout. Hovering over a well reveals the plasmid name, while clicking on a well opens the plasmid page.

Resistance Color Key

Tetracycline

Inventory

Well Plasmid Resistance
Plate 1 / A / 1 RVD-AA1
Tetracycline
Plate 1 / A / 2 RVD-AA2
Tetracycline
Plate 1 / A / 3 RVD-AA3
Tetracycline
Plate 1 / A / 4 RVD-AA4
Tetracycline
Plate 1 / A / 5 RVD-AA5
Tetracycline
Plate 1 / A / 6 RVD-AA6
Tetracycline
Plate 1 / A / 7 RVD-AA7
Tetracycline
Plate 1 / A / 8 RVD-AA8
Tetracycline
Plate 1 / A / 9 RVD-AA9
Tetracycline
Plate 1 / A / 10 RVD-AA10
Tetracycline
Plate 1 / A / 11 RVD-pA7
Tetracycline
Plate 1 / A / 12 RVD-pA8
Tetracycline
Plate 1 / B / 1 RVD-AC1
Tetracycline
Plate 1 / B / 2 RVD-AC2
Tetracycline
Plate 1 / B / 3 RVD-AC3
Tetracycline
Plate 1 / B / 4 RVD-AC4
Tetracycline
Plate 1 / B / 5 RVD-AC5
Tetracycline
Plate 1 / B / 6 RVD-AC6
Tetracycline
Plate 1 / B / 7 RVD-AC7
Tetracycline
Plate 1 / B / 8 RVD-AC8
Tetracycline
Plate 1 / B / 9 RVD-AC9
Tetracycline
Plate 1 / B / 10 RVD-AC10
Tetracycline
Plate 1 / B / 11 RVD-pA9
Tetracycline
Plate 1 / B / 12 RVD-pA10
Tetracycline
Plate 1 / C / 1 RVD-AG1
Tetracycline
Plate 1 / C / 2 RVD-AG2
Tetracycline
Plate 1 / C / 3 RVD-AG3
Tetracycline
Plate 1 / C / 4 RVD-AG4
Tetracycline
Plate 1 / C / 5 RVD-AG5
Tetracycline
Plate 1 / C / 6 RVD-AG6
Tetracycline
Plate 1 / C / 7 RVD-AG7
Tetracycline
Plate 1 / C / 8 RVD-AG8
Tetracycline
Plate 1 / C / 9 RVD-AG9
Tetracycline
Plate 1 / C / 10 RVD-AG10
Tetracycline
Plate 1 / C / 11 RVD-hA8
Tetracycline
Plate 1 / C / 12 RVD-hA9
Tetracycline
Plate 1 / D / 1 RVD-AT1
Tetracycline
Plate 1 / D / 2 RVD-AT2
Tetracycline
Plate 1 / D / 3 RVD-AT3
Tetracycline
Plate 1 / D / 4 RVD-AT4
Tetracycline
Plate 1 / D / 5 RVD-AT5
Tetracycline
Plate 1 / D / 6 RVD-AT6
Tetracycline
Plate 1 / D / 7 RVD-AT7
Tetracycline
Plate 1 / D / 8 RVD-AT8
Tetracycline
Plate 1 / D / 9 RVD-AT9
Tetracycline
Plate 1 / D / 10 RVD-AT10
Tetracycline
Plate 1 / D / 11 RVD-hA10
Tetracycline
Plate 1 / D / 12 RVD-hA11
Tetracycline
Plate 1 / E / 1 RVD-CA1
Tetracycline
Plate 1 / E / 2 RVD-CA2
Tetracycline
Plate 1 / E / 3 RVD-CA3
Tetracycline
Plate 1 / E / 4 RVD-CA4
Tetracycline
Plate 1 / E / 5 RVD-CA5
Tetracycline
Plate 1 / E / 6 RVD-CA6
Tetracycline
Plate 1 / E / 7 RVD-CA7
Tetracycline
Plate 1 / E / 8 RVD-CA8
Tetracycline
Plate 1 / E / 9 RVD-CA9
Tetracycline
Plate 1 / E / 10 RVD-CA10
Tetracycline
Plate 1 / E / 11 RVD-pC7
Tetracycline
Plate 1 / E / 12 RVD-pC8
Tetracycline
Plate 1 / F / 1 RVD-CC1
Tetracycline
Plate 1 / F / 2 RVD-CC2
Tetracycline
Plate 1 / F / 3 RVD-CC3
Tetracycline
Plate 1 / F / 4 RVD-CC4
Tetracycline
Plate 1 / F / 5 RVD-CC5
Tetracycline
Plate 1 / F / 6 RVD-CC6
Tetracycline
Plate 1 / F / 7 RVD-CC7
Tetracycline
Plate 1 / F / 8 RVD-CC8
Tetracycline
Plate 1 / F / 9 RVD-CC9
Tetracycline
Plate 1 / F / 10 RVD-CC10
Tetracycline
Plate 1 / F / 11 RVD-pC9
Tetracycline
Plate 1 / F / 12 RVD-pC10
Tetracycline
Plate 1 / G / 1 RVD-CG1
Tetracycline
Plate 1 / G / 2 RVD-CG2
Tetracycline
Plate 1 / G / 3 RVD-CG3
Tetracycline
Plate 1 / G / 4 RVD-CG4
Tetracycline
Plate 1 / G / 5 RVD-CG5
Tetracycline
Plate 1 / G / 6 RVD-CG6
Tetracycline
Plate 1 / G / 7 RVD-CG7
Tetracycline
Plate 1 / G / 8 RVD-CG8
Tetracycline
Plate 1 / G / 9 RVD-CG9
Tetracycline
Plate 1 / G / 10 RVD-CG10
Tetracycline
Plate 1 / G / 11 RVD-hC8
Tetracycline
Plate 1 / G / 12 RVD-hC9
Tetracycline
Plate 1 / H / 1 RVD-CT1
Tetracycline
Plate 1 / H / 2 RVD-CT2
Tetracycline
Plate 1 / H / 3 RVD-CT3
Tetracycline
Plate 1 / H / 4 RVD-CT4
Tetracycline
Plate 1 / H / 5 RVD-CT5
Tetracycline
Plate 1 / H / 6 RVD-CT6
Tetracycline
Plate 1 / H / 7 RVD-CT7
Tetracycline
Plate 1 / H / 8 RVD-CT8
Tetracycline
Plate 1 / H / 9 RVD-CT9
Tetracycline
Plate 1 / H / 10 RVD-CT10
Tetracycline
Plate 1 / H / 11 RVD-hC10
Tetracycline
Plate 1 / H / 12 RVD-hC11
Tetracycline
Plate 2 / A / 1 RVD-GA1
Tetracycline
Plate 2 / A / 2 RVD-GA2
Tetracycline
Plate 2 / A / 3 RVD-GA3
Tetracycline
Plate 2 / A / 4 RVD-GA4
Tetracycline
Plate 2 / A / 5 RVD-GA5
Tetracycline
Plate 2 / A / 6 RVD-GA6
Tetracycline
Plate 2 / A / 7 RVD-GA7
Tetracycline
Plate 2 / A / 8 RVD-GA8
Tetracycline
Plate 2 / A / 9 RVD-GA9
Tetracycline
Plate 2 / A / 10 RVD-GA10
Tetracycline
Plate 2 / A / 11 RVD-pG7
Tetracycline
Plate 2 / A / 12 RVD-pG8
Tetracycline
Plate 2 / B / 1 RVD-GC1
Tetracycline
Plate 2 / B / 2 RVD-GC2
Tetracycline
Plate 2 / B / 3 RVD-GC3
Tetracycline
Plate 2 / B / 4 RVD-GC4
Tetracycline
Plate 2 / B / 5 RVD-GC5
Tetracycline
Plate 2 / B / 6 RVD-GC6
Tetracycline
Plate 2 / B / 7 RVD-GC7
Tetracycline
Plate 2 / B / 8 RVD-GC8
Tetracycline
Plate 2 / B / 9 RVD-GC9
Tetracycline
Plate 2 / B / 10 RVD-GC10
Tetracycline
Plate 2 / B / 11 RVD-pG9
Tetracycline
Plate 2 / B / 12 RVD-pG10
Tetracycline
Plate 2 / C / 1 RVD-GG1
Tetracycline
Plate 2 / C / 2 RVD-GG2
Tetracycline
Plate 2 / C / 3 RVD-GG3
Tetracycline
Plate 2 / C / 4 RVD-GG4
Tetracycline
Plate 2 / C / 5 RVD-GG5
Tetracycline
Plate 2 / C / 6 RVD-GG6
Tetracycline
Plate 2 / C / 7 RVD-GG7
Tetracycline
Plate 2 / C / 8 RVD-GG8
Tetracycline
Plate 2 / C / 9 RVD-GG9
Tetracycline
Plate 2 / C / 10 RVD-GG10
Tetracycline
Plate 2 / C / 11 RVD-hG8
Tetracycline
Plate 2 / C / 12 RVD-hG9
Tetracycline
Plate 2 / D / 1 RVD-GT1
Tetracycline
Plate 2 / D / 2 RVD-GT2
Tetracycline
Plate 2 / D / 3 RVD-GT3
Tetracycline
Plate 2 / D / 4 RVD-GT4
Tetracycline
Plate 2 / D / 5 RVD-GT5
Tetracycline
Plate 2 / D / 6 RVD-GT6
Tetracycline
Plate 2 / D / 7 RVD-GT7
Tetracycline
Plate 2 / D / 8 RVD-GT8
Tetracycline
Plate 2 / D / 9 RVD-GT9
Tetracycline
Plate 2 / D / 10 RVD-GT10
Tetracycline
Plate 2 / D / 11 RVD-hG10
Tetracycline
Plate 2 / D / 12 RVD-hG11
Tetracycline
Plate 2 / E / 1 RVD-TA1
Tetracycline
Plate 2 / E / 2 RVD-TA2
Tetracycline
Plate 2 / E / 3 RVD-TA3
Tetracycline
Plate 2 / E / 4 RVD-TA4
Tetracycline
Plate 2 / E / 5 RVD-TA5
Tetracycline
Plate 2 / E / 6 RVD-TA6
Tetracycline
Plate 2 / E / 7 RVD-TA7
Tetracycline
Plate 2 / E / 8 RVD-TA8
Tetracycline
Plate 2 / E / 9 RVD-TA9
Tetracycline
Plate 2 / E / 10 RVD-TA10
Tetracycline
Plate 2 / E / 11 RVD-pT7
Tetracycline
Plate 2 / E / 12 RVD-pT8
Tetracycline
Plate 2 / F / 1 RVD-TC1
Tetracycline
Plate 2 / F / 2 RVD-TC2
Tetracycline
Plate 2 / F / 3 RVD-TC3
Tetracycline
Plate 2 / F / 4 RVD-TC4
Tetracycline
Plate 2 / F / 5 RVD-TC5
Tetracycline
Plate 2 / F / 6 RVD-TC6
Tetracycline
Plate 2 / F / 7 RVD-TC7
Tetracycline
Plate 2 / F / 8 RVD-TC8
Tetracycline
Plate 2 / F / 9 RVD-TC9
Tetracycline
Plate 2 / F / 10 RVD-TC10
Tetracycline
Plate 2 / F / 11 RVD-pT9
Tetracycline
Plate 2 / F / 12 RVD-pT10
Tetracycline
Plate 2 / G / 1 RVD-TG1
Tetracycline
Plate 2 / G / 2 RVD-TG2
Tetracycline
Plate 2 / G / 3 RVD-TG3
Tetracycline
Plate 2 / G / 4 RVD-TG4
Tetracycline
Plate 2 / G / 5 RVD-TG5
Tetracycline
Plate 2 / G / 6 RVD-TG6
Tetracycline
Plate 2 / G / 7 RVD-TG7
Tetracycline
Plate 2 / G / 8 RVD-TG8
Tetracycline
Plate 2 / G / 9 RVD-TG9
Tetracycline
Plate 2 / G / 10 RVD-TG10
Tetracycline
Plate 2 / G / 11 RVD-hT8
Tetracycline
Plate 2 / G / 12 RVD-hT9
Tetracycline
Plate 2 / H / 1 RVD-TT1
Tetracycline
Plate 2 / H / 2 RVD-TT2
Tetracycline
Plate 2 / H / 3 RVD-TT3
Tetracycline
Plate 2 / H / 4 RVD-TT4
Tetracycline
Plate 2 / H / 5 RVD-TT5
Tetracycline
Plate 2 / H / 6 RVD-TT6
Tetracycline
Plate 2 / H / 7 RVD-TT7
Tetracycline
Plate 2 / H / 8 RVD-TT8
Tetracycline
Plate 2 / H / 9 RVD-TT9
Tetracycline
Plate 2 / H / 10 RVD-TT10
Tetracycline
Plate 2 / H / 11 RVD-hT10
Tetracycline
Plate 2 / H / 12 RVD-hT11
Tetracycline
Data calculated @ 2024-04-14

Kit Plate Map - #1000000212

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