MetClo DNA Assembly Kit
(Kit #
1000000153
)
Depositing Lab: Christopher A. O'Callaghan
The MetClo DNA Assembly Kit is a 48 vector set allowing modular, hierarchical assembly of large DNA fragments using a single type IIS restriction enzyme. The kit includes the 3 E. coli strains necessary for BsaI, BpiI and LguI-based MetClo assembly vector preparation.
This kit will be sent as bacterial glycerol stocks in 96-well plate format.
Original Publication
MetClo: methylase-assisted hierarchical DNA assembly using a single type IIS restriction enzyme Lin D, O'Callaghan CA. Nucleic Acids Research. 2018 Jul 9. pii: 5050631. doi: 10.1093/nar/gky596. PubMed PMID: 29986052.
Description
MetClo allows fast and flexible modular assembly of large DNA fragments from multiple donor plasmids using a single type IIS restriction enzyme. It relies on methylation-switching to selectively block an outside flanking pair of restriction sites but allows cutting at the inner flanking sites to generate compatible adhesive ends. Any fragments that start with adhesive end ‘p’ and end with adhesive end ‘q’ can then be cloned into any of the assembly vectors with one of three types of adaptor sequence (types ‘Start’, ‘Middle’, ‘End’). This allows insert fragments to be preferentially assembled in a one-pot reaction.
Figure Legend: The standard MetClo vector set contains 30 low copy vectors with a p15a replication origin and either an ampicillin (Amp), kanamycin (Kan) or chloramphenicol (Cam) selection marker, and a LacZa negative selection marker flanked by different adaptor sequences sufficient for the assembly of up to 6 fragments in a single reaction. The vector set also contains 18 single copy vectors with an F replication origin, one of the three antibiotic selection markers, and a fragment containing a ColE1 replication origin and a LacZα negative selection marker flanked by different adaptor sequences sufficient for assembly of up to 4 fragments in a single reaction.
Kit-Specific Resources
Click below to download the MetClo Strain Verification protocol:
MetClo Strain Verification protocol 66.3Â 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 MetClo DNA Assembly Kit was a gift from Christopher A. O'Callaghan (Addgene kit #1000000153).”
For your Reference section:
MetClo: methylase-assisted hierarchical DNA assembly using a single type IIS restriction enzyme Lin D, O'Callaghan CA. Nucleic Acids Research. 2018 Jul 9. pii: 5050631. doi: 10.1093/nar/gky596. PubMed PMID: 29986052.
MetClo DNA Assembly Kit - #1000000153
- 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
| Chloramphenicol | |
| Kanamycin | |
| Ampicillin | |
| Bleocin (Zeocin) |
Inventory
| Well | Plasmid | Resistance |
|---|---|---|
| A / 1 | pMXLC_pFa | Chloramphenicol |
| A / 2 | pMXLC_aFb | Chloramphenicol |
| A / 3 | pMXLC_bFc | Chloramphenicol |
| A / 4 | pMXLC_cFd | Chloramphenicol |
| A / 5 | pMXLC_dFe | Chloramphenicol |
| A / 6 | pMXLC_aFq | Chloramphenicol |
| A / 7 | pMXLC_bFq | Chloramphenicol |
| A / 8 | pMXLC_cFq | Chloramphenicol |
| A / 9 | pMXLC_dFq | Chloramphenicol |
| A / 10 | pMXLC_eFq | Chloramphenicol |
| A / 11 | pMXBC_pFa | Chloramphenicol |
| A / 12 | pMXBC_aFb | Chloramphenicol |
| B / 1 | pMXBC_bFc | Chloramphenicol |
| B / 2 | pMXBC_aFq | Chloramphenicol |
| B / 3 | pMXBC_bFq | Chloramphenicol |
| B / 4 | pMXBC_cFq | Chloramphenicol |
| B / 5 | pMXLK_pFa | Kanamycin |
| B / 6 | pMXLK_aFb | Kanamycin |
| B / 7 | pMXLK_bFc | Kanamycin |
| B / 8 | pMXLK_cFd | Kanamycin |
| B / 9 | pMXLK_dFe | Kanamycin |
| B / 10 | pMXLK_aFq | Kanamycin |
| B / 11 | pMXLK_bFq | Kanamycin |
| B / 12 | pMXLK_cFq | Kanamycin |
| C / 1 | pMXLK_dFq | Kanamycin |
| C / 2 | pMXLK_eFq | Kanamycin |
| C / 3 | pMXBK_pFa | Kanamycin |
| C / 4 | pMXBK_aFb | Kanamycin |
| C / 5 | pMXBK_bFc | Kanamycin |
| C / 6 | pMXBK_aFq | Kanamycin |
| C / 7 | pMXBK_bFq | Kanamycin |
| C / 8 | pMXBK_cFq | Kanamycin |
| C / 9 | pMXLP_pFa | Ampicillin |
| C / 10 | pMXLP_aFb | Ampicillin |
| C / 11 | pMXLP_bFc | Ampicillin |
| C / 12 | pMXLP_cFd | Ampicillin |
| D / 1 | pMXLP_dFe | Ampicillin |
| D / 2 | pMXLP_aFq | Ampicillin |
| D / 3 | pMXLP_bFq | Ampicillin |
| D / 4 | pMXLP_cFq | Ampicillin |
| D / 5 | pMXLP_dFq | Ampicillin |
| D / 6 | pMXLP_eFq | Ampicillin |
| D / 7 | pMXBP_pFa | Ampicillin |
| D / 8 | pMXBP_aFb | Ampicillin |
| D / 9 | pMXBP_bFc | Ampicillin |
| D / 10 | pMXBP_aFq | Ampicillin |
| D / 11 | pMXBP_bFq | Ampicillin |
| D / 12 | pMXBP_cFq | Ampicillin |
| E / 1 | DH10B-M.Osp807II | Bleocin (Zeocin) |
| E / 2 | DH10B-M2.NmeMC58II | Bleocin (Zeocin) |
| E / 3 | DH10B-M.XmnI | Bleocin (Zeocin) |