Addgene Research Program

At Addgene, our mission is to accelerate science by making key research tools available to the global community. In addition to supporting thousands of individual depositors, we now proactively identify and fill critical gaps in the plasmid catalog through the Addgene Research Program.

This initiative is guided by depositor input, user feedback, and our own assessment of emerging research needs and aims to ensure that scientists have reliable access to the plasmids they need to advance their work without having to duplicate efforts.

To assist researchers Addgene has created:

• Empty backbones in a variety of expression systems to provide flexible starting points for cloning and expression experiments.
• Control backbones to help confirm experiments are working as expected.
• Viral packaging plasmids to support gene delivery experiments.
• Lentiviral and AAV expression plasmids for commonly studied genes to enable efficient gene delivery.

Beyond providing the plasmids themselves, we go further to ensure reliability and reproducibility:

• Each plasmid is validated in its intended expression system, so users can be confident the construct functions as designed.
• On each material page, we supply data demonstrating functionality to guide expectations and support experimental planning.
• Protocols are provided so that researchers can reproduce our results and build on them effectively in their own labs.

This collection is just the beginning. We will continue to expand the collection to empower scientists to pursue new questions without unnecessary barriers.

Empty and Control Backbones for Mammalian Expression

Addgene’s pAG CMV plasmids have a CMV promoter for ubiquitous transient transgene expression. pAG CMV backbones are available untagged or with an N- or C-terminal hemagglutinin (HA) tag to aid with protein detection or purification. Key features of the pAG CMV series, including the HA tag, CMV promoter, P2A/puromycin, WPRE and SV40 polyadenylation signal, are flanked with unique restriction sites allowing users to easily replace existing features with alternative features, if desired.

To confirm functionality of the backbone, Addgene cloned EGFP into the empty backbones, transfected cells, and confirmed transient EGFP expression. The plasmids expressing pAG CMV EGFP are available as control plasmids.

Empty and Control Backbones for Bacterial Expression

Addgene’s pAG GST plasmid supports IPTG-inducible expression of N-terminal GST-tagged proteins for bacterial expression and purification. The backbone contains a thrombin site for enzymatic cleavage of the GST tag, if desired.

To confirm functionality of the backbone, Addgene cloned EGFP into the empty backbone, expressed the GST-tagged protein in E. coli, and purified the GST-EGFP with glutathione spin columns. The pAG GST EGFP-expressing plasmid is available as a control plasmid.

Empty and Control Backbones for Antibody Expression

Addgene’s pAG antibody expression series provides plasmids containing mouse (mIgG2a heavy chain, mKappa light chain) constant regions. Researchers can clone in their antibody variable region of choice to enable mammalian expression.

To generate a full-length antibody, plasmids must be used in matched species pairs; for example, a mouse IgG2a heavy chain plasmid with a mouse Kappa light chain plasmid.

To confirm functionality, Addgene cloned variable regions from well-characterized epitope tag antibodies (Anti-c-Myc [9E10] for mouse and Anti-HA [12CA5] for rat) into the pAG Ab plasmids. The paired heavy and light chains were co-expressed in HEK293 cells, antibodies were purified by affinity chromatography, and specificity was verified.

Empty and Control Backbones for AAV Expression

Addgene’s pAG AAV plasmids have a CMV promoter for ubiquitous transgene expression. pAG AAV backbones are available untagged or with an N- or C-terminal hemagglutinin (HA) tag to aid with protein detection or purification. Key features of the pAG AAV series, including the HA tag, CMV promoter, and SV40 polyadenylation signal, are flanked with unique restriction sites allowing users to easily replace existing features with alternative features, if desired. These plasmids lack the adenoviral helper genes required for AAV particle assembly and must be co-transfected with both an adenoviral helper plasmid and a capsid plasmid, such as pAdDeltaF6 (Plasmid #112867) and pAAV2/6 (Plasmid #240485). Find additional helper and capsid plasmids on the Adeno-Associated Viral (AAV) Plasmids page.

To confirm functionality of the backbone, Addgene cloned EGFP into the empty backbones, packaged the transgene into AAV particles, and confirmed transduction and expression in mammalian cells.

AAV Packaging Plasmids

Addgene cloned the AAV capsid genes for AAV serotypes 6 and 11 into packaging plasmids. These packaging plasmids can be transfected in parallel with an adenoviral helper plasmid and an AAV transfer plasmid, such as pAdDeltaF6 (Plasmid #112867) and pAG AAV CMV EGFP (Plasmid #242959) to create AAV particles. Find additional helper and transfer plasmids on the Adeno-Associated Viral (AAV) Plasmids page.

To confirm functionality, Addgene used these plasmids to package fluorescent transgenes into AAV particles and confirmed transduction and expression in mammalian cells.

AAV Expression Plasmids

Addgene has cloned common research tools into an AAV expression backbone for efficient gene delivery. These plasmids lack the adenoviral helper genes required for AAV particle assembly and must be co-transfected with a compatible adenoviral helper plasmid and the desired capsid plasmid, such as pAdDeltaF6 (Plasmid #112867) and pAAV2/6 (Plasmid #240485). Find additional helper and capsid plasmids on the Adeno-Associated Viral (AAV) Plasmids page.

To confirm functionality, Addgene packaged the transgene into AAV particles and confirmed transduction and expression in mammalian cells.

Empty and Control Backbones for Lentiviral Expression

Addgene’s pAG Lenti plasmids have a CMV promoter and puromycin selection for ubiquitous transgene expression and rapid selection of resistant cells during stable cell line selection. pAG Lenti backbones are available untagged or with an N- or C-terminal hemagglutinin (HA) tag to aid with protein detection or purification. Key features of the pAG Lenti series, including the HA tag, CMV promoter, P2A/puromycin, WPRE and SV40 polyadenylation signal, are flanked with unique restriction sites allowing users to easily replace existing features with alternative features, if desired. These backbones do not contain all of the genes necessary to generate lentivirus and must be transfected with lentiviral packaging and envelope plasmids, such as pMD2.G (Plasmid #12259) and psPAX2 (Plasmid #12260), to make lentiviral particles. Find additional packaging and envelope plasmids on the Lentiviral Plasmids page.

To confirm functionality of the backbone, Addgene cloned EGFP into the empty backbones, produced lentiviral particles, generated stable cell lines, and confirmed stable EGFP expression. The pAG Lenti EGFP expressing plasmids are available as control plasmids.

Lentiviral Expression Plasmids

Addgene has cloned common genes into a N-terminally HA-tagged lentiviral expression backbone for CMV-driven stable transgene expression. These backbones do not contain all of the genes necessary to generate lentivirus and must be transfected with lentiviral packaging and envelope plasmids, such as pMD2.G (Plasmid #12259) and psPAX2 (Plasmid #12260), to make lentiviral particles. Find additional packaging and envelope plasmids on the Lentiviral Plasmids page.

To confirm functionality of the plasmids, Addgene produced lentiviral particles, generated stable cell lines, and confirmed stable HA expression.

Empty and Control Backbones for Retroviral Expression

The pAG Retro vectors are designed for stable gene delivery and expression in mammalian cells. pAG Retro plasmids use a Mo-MuLV LTR promoter to drive long-term, transgene expression across a wide range of cell types. Plasmids are available with puromycin, hygromycin, or neomycin resistance for stable cell line generation. These backbones do not contain all of the genes necessary to generate lentivirus and must be transfected with lentiviral packaging and envelope plasmids, such as pMD2.G (Plasmid #12259) and psPAX2 (Plasmid #12260), to make lentiviral particles. Find additional packaging and envelope plasmids on the Lentiviral Plasmids page.

To confirm functionality of the backbone, Addgene cloned EGFP into the empty backbones, produced retroviral particles, generated stable cell lines, and confirmed stable EGFP expression. The pAG retro EGFP-expressing plasmids are available as control plasmids.

Additional Resources

• Choosing Your Perfect Plasmid Backbone
• Viral Vector Guides and Plasmids
• Fluorescent Proteins: Plasmids and Resources
• Protocols