Skip to main content
This website uses cookies to ensure you get the best experience. By continuing to use this site, you agree to the use of cookies.

Please note: Your browser does not support the features used on Addgene's website. You may not be able to create an account or request plasmids through this website until you upgrade your browser. Learn more

Please note: Your browser does not fully support some of the features used on Addgene's website. If you run into any problems registering, depositing, or ordering please contact us at [email protected]. Learn more

Addgene
plasmid kit icon

AspFlex Kit
(Kit # 1000000217 )

Depositing Lab:   Veronica Godoy-Carter

This collection is a set of Modular Cloning (MoClo) plasmids for Acinetobacter spp. and Acinetobacter baumannii for protein expression analyses and CRISPRi experiments.

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

Add to Cart
$ 400 USD + shipping

Available to academics and nonprofits only.

Original Publication

AspFlex: Molecular Tools to Study Gene Expression and Regulation in Acinetobacter baumannii. Brychcy M, Kokodynski A, Lloyd D, Godoy VG. ACS Synth Biol. 2023 Aug 16. doi: 10.1021/acssynbio.3c00167. PubMed (Link opens in a new window) Article (Link opens in a new window).

Description

This collection is a set of MoClo plasmids for both Acinetobacter spp. and Acinetobacter baumannii. The plasmids support easy Golden Gate cloning for protein expression analyses in Acinetobacter spp., including the construction of transcriptional reporters or fusion proteins. The collection is an expansion pack for the EcoFlex collection for E. coli (Addgene Kit#: #1000000080). The kit introduces a new plasmid based CRISPRi Golden Gate system to A. baumannii, with plasmids for cloning sgRNAs and plasmids containing a catalytically deficient Cas9 (dCas9).

Several accessory plasmids are also available, which contain an additional Tellurite resistance cassette for multi-drug resistance strains. These plasmids are not part of the kit but can be ordered separately (see the Accessory Plasmids tab).

Two schematics of molecular parts represented as blocks and how they can be connected. Part A shows general MoClo approach. In part B, the Level 0 unit is a custom sgRNA, which are assembled into Level 2 and 3 CRISPRi units.
  • Assembly mechanism of molecular elements in AspFlex. (A) A DNA recombinant approach to clone any transcriptional unit (for example for protein expression, transcriptional reporters, etc.). The system is organized in the standardized modular level based MoClo fashion. Here, a Level 0 plasmid consists of the elements necessary to form a transcriptional unit, a Level 1 plasmid places the elements from Level 0 plasmids to create a single transcriptional unit, while a Level 2 plasmid combines multiple transcriptional units from Level 1 plasmids to create a plasmid with two to five independent transcriptional units. Lastly, a Level 3 plasmid allows for the combination of six or more independent transcriptional units. (B) Workflow to assemble a CRISPRi unit. Here, a Level 0 allows the possibility to create a custom sgRNA, which can then be transferred into a shuttle vector in a Level 1 plasmid containing a (inducible) dCas9 provided with the kit. A Level 0 plasmid containing dCas9, which can be moved into different level plasmids is also provided. Up to four independent sgRNAs can be used in Level 2 plasmids. Level 3 then allows for five or more sgRNAs.

Kit Documentation

AspFlex Kit Protocols (PDF, 263 KB)

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 AspFlex Kit was a gift from Veronica Godoy-Carter (Addgene kit #1000000217).”

For your Reference section:

AspFlex: Molecular Tools to Study Gene Expression and Regulation in Acinetobacter baumannii. Brychcy M, Kokodynski A, Lloyd D, Godoy VG. ACS Synth Biol. 2023 Aug 16. doi: 10.1021/acssynbio.3c00167. PMID: 37587063 (Link opens in a new window)

AspFlex Kit - #1000000217

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 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

Ampicillin
Kanamycin
Chloramphenicol

Inventory

Well Plasmid Resistance
A / 1 pMB1-A
Ampicillin
A / 2 pMB1-B
Ampicillin
A / 3 pMB1-C
Ampicillin
A / 4 pMB1-D
Ampicillin
A / 5 pMB1-D1
Ampicillin
A / 6 pMB1-E
Ampicillin
A / 7 pMB3-A
Ampicillin
A / 8 pMB3-B
Ampicillin
A / 9 pMB1-A-ptet-dCas9
Ampicillin
A / 10 pMB1-A-ptet-FLAG-dCas9
Ampicillin
A / 11 pMB1-A-pBAD-dCas9
Ampicillin
A / 12 pMB2a
Kanamycin
B / 1 pMB2b
Kanamycin
B / 2 pMB2-A
Kanamycin
B / 3 pMB2-B
Kanamycin
B / 4 pMB2-C
Kanamycin
B / 5 pMB2-D
Kanamycin
B / 6 pBP_sgRNA
Chloramphenicol
B / 7 pBP_dCas9
Chloramphenicol
Data calculated @ 2024-04-18

Kit Plate Map - #1000000217

Plate #1

  1 2 3 4 5 6 7 8 9 10 11 12
A
B
C
D
E
F
G
H

Accessory Plasmids

The following related plasmids are also available with additional Tellurite resistance cassette for multi-drug resistance strains. These plasmids are not included in the kit, but can be ordered separately:

Content blocked, you may need to disable your ad-blocker.