How to Deposit Your Plasmids with Addgene

By Multiple Authors

At Addgene, it is our mission to make it easy for you to share plasmids. To achieve this goal, we will archive any plasmids you've deposited with us and distribute them to scientists worldwide. What's more, depositing is free! We've written this post as a step-by-step guide to the online deposit process.

To deposit plasmids online, you will need to create an account with Addgene before starting the process.

After you have logged in to your account, you can begin the deposit process by clicking “Online Submission” (for online submission) or “Spreadsheet Submission” (for emailed submission) in the Deposit menu:

A screenshot of the Addgene home page with arrows pointing at Online Submission and Spreadsheet Submission on the Deposit drop down menu.

Figure 1: Begin the deposit process by clicking “Online Submission” or “Spreadsheet Submission.”

Enter article information

You have three options when starting the deposit:

A screenshot of the “Start Your Plasmid Deposit” page with three options. The first option is “Submit Plasmids from a Published Article”. The second option is “Submit Pre-Publication or Unpublished Plasmids”. The third option is “Submit Plasmids Using a Spreadsheet”.

Figure 2: Choose to submit published or unpublished plasmid(s) online
or submit plasmid(s) using a spreadsheet.

  1. You can “Submit Plasmids from a Published Article”: This option allows you to quickly associate your plasmids with the original publication where you described them. Search for your article as you would search for it in PubMed and choose the appropriate publication. This publication will be listed at the top of the final plasmid pages and scientists will be asked to cite this article when using your plasmids.

  2. You can “Submit Pre-Publication or Unpublished Plasmids”: Unpublished plasmids or plasmids that will not be published are accepted. Addgene also encourages submitting pre-publication plasmids so they can be available when the paper is published. Plasmids can be linked to a peer-reviewed manuscript OR a preprint. In the case of preprint submission, we are happy to update the publication information once the manuscript is published in a peer-reviewed journal. To start this process, simply enter your preprint title or a general description of the plasmids and select your desired distribution status.
    • Hold for publication: we will initiate our MTA process and perform standard quality control but will not make the plasmids available until you update us with publication information or permission to distribute.

    • Distribute pending QC: your plasmids become available for distribution as soon as they pass our standard QC process and MTA has been approved.

  3. You can "Submit Plasmids Using Our Spreadsheet": The spreadsheet option is recommended for depositing 10 or more plasmids. (If you believe you need to deposit a collection that is greater than 75 plasmids, please let us know by contacting deposit@addgene.org. These deposits are considered large deposits and are exceptions to our standard workflow.) Our spreadsheet contains columns that correspond to the plasmid data we need; you can copy and paste your information directly into the file. This file can be emailed to deposit@addgene.org along with your GenBank files, plasmid sequences, and/or maps. If available, we welcome submission of QUEEN-generated GenBank files (Mori and Yachie, 2021) but can accept sequence files in any format.

Learn more about the Deposit Spreadsheet


Add plasmids to your article

After you’ve selected your publication or entered your description, the next page will allow you to add plasmids to your article:

 A screenshot of the “Add Plasmids” page. Instructions are provided for naming your plasmid, choosing the type of plasmid, and adding a description/experimental purpose. Additional plasmids can be entered by clicking the “Add” button. Additional plasmids from a new article can be entered by clicking the “Submit plasmids from a new article” button.

Figure 3: Add plasmids to the article by entering a plasmid name,
plasmid type and plasmid description/experimental purpose.

On the article page, enter the name, type, and description of each plasmid associated with this deposit as explained below:

Pro Tip!

Add all the plasmid names and descriptions first before entering the data. This step associates an Addgene ID with your plasmids, and adding all the plasmids at once increases the likelihood that the plasmids are given sequential IDs.

 

Name – we encourage the use of descriptive plasmid names and using the exact names listed in your publication. Aliases for your plasmid can be added on subsequent pages.

Type - please choose from one of the following descriptions for your plasmid:

  • Encodes One Insert:

A plasmid with one gene or protein coding sequence cloned into it. 

  • Encodes gRNA/shRNA

A plasmid with one gRNA or shRNA sequence cloned into it. gRNAs are used for directing CRISPR/Cas9 to the appropriate genomic location, while shRNAs are used to knockdown expression of mRNAs with complementarity to the shRNA sequence.

  • Empty Backbone

A plasmid designed for cloning a gene/insert into it. Empty backbones often contain epitope tags or fusion proteins that are appended to new inserts through cloning.

  • Encodes Multiple Inserts

A plasmid encoding multiple inserts can contain multiple genes or small non-coding RNAs. Only enter as a multiple insert if you want to indicate separate promoters, tags, or cloning information for the inserts.

*Note  If you will be depositing multiple plasmids and would prefer to send us your plasmid information in spreadsheet format, e-mail us at  deposit@addgene.org

Special cases

Please contact us at deposit@addgene.org or +1 (617)-ADDGENE (233-4363) if your deposit falls into one of the special cases below:

 

  • Pooled Library

A pooled library is a set of plasmids all built with the same backbone and only differing in the gene/insert. Pooled libraries are normally supplied as liquid DNA with all of the plasmids in the library in a single tube. The number of unique plasmids in a pooled library can range from a few hundred to millions. Pooled libraries must be verified by next-generation sequencing. Please see the help center article, “Can I deposit a pooled DNA library with Addgene?” or contact us at deposit@addgene.org or +1 (617)-ADDGENE (233-4363) with additional questions.

  • Bacterial Strain

Addgene can distribute your bacterial strains if they:

  • are E. coli that can be used under BSL1 conditions
  • were generated in your lab
  • are necessary for use of a plasmid you are depositing
  • can be grown using standard conditions (37 °C, LB, antibiotics, etc.) and stored as glycerol stocks

We will need information on how to verify any genomic alterations unique to your strain. Please contact us at deposit@addgene.org or +1 (617)-ADDGENE (233-4363) if you’d like to distribute your bacterial strain(s) through Addgene.

Description – please indicate the gene cloned into your plasmid, any tags or fusion proteins contained in the plasmid, any mutations, and the intended purpose of the plasmid. A descriptive purpose focuses on the practical application of the plasmid and will help a scientist outside your field understand if this plasmid will work for them.

Once you've named and described all of the plasmids in your deposit, you should click the "Enter Data" button to begin entering the specifics of your plasmids.

A screenshot of the “Add Plasmids” page with an arrow pointing to the “Enter Data” button which leads to the pages where plasmid data can be added.

Figure 4: Click the “Enter Data” button to add your plasmid information.

Enter plasmid information

During the remainder of the process, you will enter plasmid data in the pages described below.

You can complete these pages sequentially by entering the appropriate data and pressing “Save and Continue to Next Step” (green button at the bottom of Figure 5) or you can click the save button and jump around to the different pages by clicking on the appropriate boxes at the top of the page.

Exclamation.jpgPro Tip!

Be sure to press the blue “Save” button at the bottom of the page before moving onto a different page so that any data you enter is not lost.

 

Step 1: Sequences, Maps, and Files

Please provide any full or partial plasmid sequences and any vector maps. The more sequence data available, the better. We highly encourage you to upload full sequence data whenever possible. Note that the full plasmid sequence can be assembled or theoretical — it does not have to be entirely verified by sequencing.

Upload any support files that will make it easier for scientists to best utilize your plasmids (these can be protocols, GenBank files, etc). We can accept sequence files in any format. We encourage submission of QUEEN-generated GenBank files (Mori and Yachie, 2021).

A screenshot of the “Sequences, Maps, and Files” page. There are “Yes” and “No” buttons for “Plasmid Sequence”, “Plasmid Map”, and “Support Files” that when “Yes” is selected, fields become visible allowing addition of files and/or sequence.

Figure 5: Add sequence, maps, and files.

Step 2: Gene and Insert

*Note: This page will not appear for empty backbones

Please enter the name of any gene contained within your insert and select the species that the gene or insert sequence came from. Entering this information will populate a list of possible Entrez Gene matches to your insert. Select the appropriate gene if it appears in the Entrez Gene list, as this will make it easier for other scientists to find information related to your plasmid. It is optional to include the total size of the insert and the GenBank ID associated with your insert.

Exclamation.jpgPro Tip!

To get the Entrez Gene list to auto-populate with the correct gene, entering the Aliases or “Also known as” names in the Gene/Insert box works best. Once you have selected the appropriate gene from the list, save your progress and you can then modify the gene/insert and alternative name boxes as needed.

Use the “Relevant mutations/deletions” box to describe any mutations within your insert using standard mutation notation. Please indicate the amino acid change as well as any functional consequences of that change. For example, “Mutated aspartate 123 to Lysine (D123K), lowers biosensor affinity for calcium.”

Please describe any tags or fusions (e.g. His, FLAG, EGFP, etc) on your insert. Use the drop-down menu to indicate whether the tag is on the N- or C- terminus and whether it was present in the backbone or was a part of the insert (when cloned into the vector backbone).

As part of our quality control process, we will perform NGS sequencing to confirm features you annotate on this page.

A screenshot of the “Gene and/or insert” information page. Fields for Gene/Insert, Alternative names, Insert size (bp), Species of gene, GenBank ID, Entrez gene, relevant mutations/deletions, and fusion proteins or tags can be populated.

Figure 6: Enter gene and/or insert information.

Step 3: Cloning Information

Please enter the name of the vector backbone you cloned your insert into and fill out the appropriate boxes with the vector backbone manufacturer, vector backbone size, total size of your vector + insert, any modifications you made to the backbone, and the vector type.

Exclamation.jpgPro Tip!

Reference the Addgene ID in the “backbone manufacturer” box for backbones requested through Addgene. For example, you could enter “Feng Zhang (Addgene plasmid # 52961)”.

 

The vector type should describe how your plasmid is intended to be used. For instance, if your plasmid is designed to allow you to produce large amounts of your insert in E. coli, then this is a Bacterial Expression vector. If your plasmid can be classified in multiple categories, please check all of the appropriate categories (e.g., for a lentiviral vector to be used in mammalian cells, check Lentiviral and Mammalian Expression). This information will make it easier for scientists to find your plasmid by type.

A screenshot of the top section of the “Cloning” page. Fields for Vector backbone, backbone manufacturer, backbone size without insert, total vector size, modifications to backbone, and vector type can be populated.

Figure 7: Add vector backbone information.

Input cloning information to describe how your gene was inserted in the plasmid backbone and what primers can be used to verify it, as shown below:

 

A screenshot of the “Cloning information” at the bottom of the “Cloning” page. Fields for promoter, cloning method, and 5’ and 3’ sequencing primers can be populated.

Figure 8: Add cloning information for the gene or insert.

Step 4: Growth and Distribution

Please indicate which strain Addgene should propagate your plasmids in: DH5α, NEB Stable, or ccdB Survival.­ Whenever possible, we propagate plasmids in the standard cloning strain DH5α. Please indicate if your plasmid cannot be propagated in DH5α. For plasmids with highly repetitive sequences (which can be prone to recombination in bacteria) such as lentiviral, retroviral, and AAV plasmids, we recommend the NEB Stable strain. For plasmids containing the ccdB gene, such as Gateway vectors, we recommend the ccdB Survival strain.

A screenshot of the “growth and distribution” page. Fields for bacterial resistance, high or low copy number, growth strain, growth temperature, additional growing instructions, selectable marker, safety, legal, gene origin, and comments can be populated.

Figure 9: Enter the growth and distribution information

Step 5: Verification

On this page you will see a preview of the final plasmid page that will appear on Addgene’s website once your plasmids are available online. Please review all of the information on this page to make sure it is correct. You can easily update the information by navigating to the previous pages using the links at the top of the page.

A screenshot of a completed plasmid page. The top left shows the SnapGene-generated image of the plasmid sequence and/or uploaded plasmid maps. The top right shows the purpose and publication. The lower part of the page lists the backbone information including, vector backbone, backbone manufacturer, backbone size without insert, total vector size, and vector type; growth in bacteria information including bacterial resistance, growth temperature, growth strain, and copy number; gene/insert information including gene/insert name, species, insert size, mutation, promoter, and tag/fusion protein; and cloning information including cloning method, 5’ and 3’ cloning sites, and 5’ and 3’ sequencing primers.

Figure 10: A completed plasmid page for review before submission.

Once you are satisfied with the plasmid information, click on the green “Save Verified Data and Return to Article” button to return to the table listing all of the plasmids for this deposit. You can then enter the information for the next plasmids in your deposit.

Exclamation.jpgPro Tip!

If you have multiple plasmids with the same gene or vector backbone, you can copy some information from a previous plasmid in your deposit by selecting the plasmid to copy from the drop-down menu and clicking the blue “Copy” button.

*Note: The “Copy” button will only appear after you have entered information for your first plasmid and can be used on each individual page (Gene and insert, Cloning Information, Growth and Distribution).

 

A screenshot of the “Cloning” page with a red arrow pointing at the “Copy” bottom located near the top right of the page.

Figure 11: The “Copy” button is located near the top right of each data entry page.

Once you have completed the data entry process for all of the plasmids in your deposit, return to the Add Plasmids page and click the green “I am done entering data. Request Deposit Kit” button.

 A screenshot of the “Add Plasmids” page with a red arrow pointing at the “I am done entering data. Request deposit kit.” Button at the bottom right of the page.

Figure 12: Click the “I am done entering data. Request Deposit Kit.”
button to complete the data entry.

Clicking this button will allow you to choose which plasmids you wish to deposit, associate the plasmids with a Principal Investigator (PI), provide us with your shipping address for deposit materials, and agree to our terms of submission:

  • I hereby certify that I am affiliated with the institution ___. If this is no longer true, please modify your profile before continuing.
  • I hereby certify that the material I am submitting to Addgene qualifies as BL1 or BL2 as defined by NIH guidelines.
  • I hereby certify that I have disclosed all known hazardous and infectious properties of the submitted materials.

  • As the Provider Scientist, I hereby consent to the distribution of the submitted material under the terms deemed necessary by my institution or other third party rights holders.

Once this process is complete, you will receive a Deposit Confirmation email from us with a unique Deposit ID. Addgene will send you a package containing instructions on how to prepare your plasmids and a prepaid label with shipping materials to send your plasmids to us. Essentially, plasmids can be submitted as 15 µL of DNA in a 1.5 mL microfuge tube at a concentration of 0.1 -1µg/µL (or bacterial streaks, if you prefer).

If you wish to edit your plasmid data after submission, please email us at deposit@addgene.org. Include your deposit number and which data needs to be updated.

When your plasmids arrive at Addgene, we will notify you that we have received them and they will undergo our quality control process to verify your plasmids. Our friendly Addgene scientists will notify you once your plasmids are online and available to the research community.

This article was originally written by Tyler Ford in January 2016. It was updated by Angela Holmes and Christina Mork in February of 2024. 


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