Plasmids for Stem Cell Research
Stem cells are proliferative, unspecialized cells with the capacity to differentiate into many different cell types in the body during development. Unlike other cell types, each stem cell has the potential to either remain a stem cell or become another type of cell with a more specialized function. They also have the unique potential to continuously divide while maintaining an undifferentiated state. Stem cells are classified as pluripotent or multipotent, based on their ability to generate different cell lineages. Pluripotent stem cells, such as embryonic cells, can potentially give rise to any other cell type in the body. Multipotent stem cells, such as adult (or somatic) stem cells, can differentiate into multiple cell types, but are typically limited to related cell lineages.
Pluripotent embryonic and multipotent adult stem cells have been studied for over 30 years. All of this research led to Kazutoshi Takahashi and Shinya Yamanaka’s breakthrough development of induced pluripotent stem cells (iPSCs) in both mouse and human cells in 2006 and 2007, respectively. To create iPSCs, fully-differentiated adult somatic cells are returned to a pluripotent state using a cocktail of factors (Oct3/4, Sox2, c-Myc, and Klf4) that are known to maintain pluripotency during embryonic development. This “reprogramming” of the cell to an embryonic state holds immense potential for drug development, disease modeling, and regenerative medicine.
iPSCs, much like their ES cell cousins, have the hallmarks of pluripotent cells--they are capable of differentiating into every other cell type in the body and have the capacity to give rise to an entire organism. iPSC technology removes the possible ethical concerns related to embryonic stem cell use and has the major advantage that iPSCs can potentially be derived from patient-specific or disease-specific cells. For this reason, iPSCs have become an excellent unlimited stem cell source for therapeutic use.
Since 2006, scientists have developed many approaches to generate iPSCs. The generation of iPSCs is relatively simple in concept: ectopically express a cocktail of stem cell reprogramming factors and wait for cells to de-differentiate. However it may be difficult to decide which method to use. Addgene's blog post Delivery Methods for Generating iPSCs provides an overview of several different reprogramming methods with the goal of helping readers choose a strategy suited to their research. Once you know which method you would like to use, browse Addgene's collection of Reprogramming Plasmids in the table below.
Once iPSCs have been created they can be directly differentiated into specific somatic stem cells or fully differentiated cell types. Browse Addgene's collection of Differentiation Plasmids below.
Alternatively, it is also possible to directly differentiate one differentiated somatic cell into another via transdifferentiation. This method of cell lineage conversion can be faster, bypasses intermediate pluripotent stages, and can occur without cell division, all of which lower the risk of mutation and minimize tumorigenicity, an inherent disadvantage of iPS cell technology. Browse Addgene's collection of Transdifferentiation Plasmids below.
Many of the techniques for reprogramming, differentiation, and transdifferentiation utilize plasmids to increase expression of key factors. The tables below will help you find plasmids available from Addgene that can be used to create iPSCs or another cell type. For more information on stem cells, visit the NIH Stem Cell information page.
Browse the table below for plasmids used to create induced pluripotent stem cells (iPSCs). Sort the table by delivery method, species, or PI and then select the article link to find more information.
|CRISPRa||Human||CRISPR activator system for reprogramming human cells to pluripotency by activating enogenous genes||Human pluripotent reprogramming with CRISPR activators. Nat Commun. 2018 Jul 6;9(1):2643.||Otonkoski|
|Lentivirus||Human||Expression of human Sox2, Nanog, Oct4, and Lin28 from four separate lentiviral plasmids||Induced Pluripotent Stem Cell Lines Derived from Human Somatic Cells. Science. 2007 Dec 21;318(5858):1917-20.||Thomson|
|Lentivirus||Human||Doxycycline-inducible expression of human Oct4, Sox2, Klf4, and c-Myc from four separate lentiviral plasmids||A drug-inducible system for direct reprogramming of human somatic cells to pluripotency. Cell Stem Cell. 2008 Sep 11. 3(3):346-53.||Jaenisch|
|Lentivirus||Human||Single polycistronic lentiviral vector for the expression of human Oct4, Sox2, and Klf4||Polycistronic lentiviral vector for "hit and run" reprogramming of adult skin fibroblasts to induced pluripotent stem cells. Stem Cells. 2009 May . 27(5):1042-9.||Townes|
|Lentivirus||Human||Single polycistronic lentiviral vector for the expression of human Oct4, Klf4, Sox2, and c-Myc||Human Induced Pluripotent Stem Cells Produced Under Xeno-Free Conditions. Stem Cells Dev. 2010 Aug;19(8):1221-9.||Cibelli|
|Lentivirus||Human||Single polycistronic, doxycycline-inducible lentiviral vector for the expression of human Oct4, Klf4, c-Myc, and Sox2||Integrative Analyses of Human Reprogramming Reveal Dynamic Nature of Induced Pluripotency. Cell. 2015 Jul 16;162(2):412-24.||Mikkelsen|
|Lentivirus||Human||Expression of human Klf4, Oct4, c-Myc, and Sox2 as VP-16 transcriptional activating fusions in single lentiviral vectors for human iPSC generation||OCT4 and SOX2 Work as Transcriptional Activators in Reprogramming Human Fibroblasts. Cell Rep. 2017 Aug 15;20(7):1585-1596.||Ptashne|
|Minicircle||Human||Non-integrating mini-intronic plasmid (MIP) polycistronic expression of human Oct4, Klf4, Sox2, c-Myc and hairpin RNA p53 for single plasmid reprogramming||Novel codon-optimized mini-intronic plasmid for efficient, inexpensive, and xeno-free induction of pluripotency. Sci Rep. 2015 Jan 28;5:8081.||Kay and Wu|
|MMLV-derived Retrovirus||Human||Yamanaka factors for generating human iPS cells; retroviral expression of human Sox2, Oct3/4, Klf4, and c-Myc from four separate plasmids||Induction of pluripotent stem cells from adult human fibroblasts by defined factors. Cell. 2007 Nov 30. 131(5):861-72.||Yamanaka|
|Plasmid||Human||Non-integrating transient expression of human L-Myc, Lin28, Sox2, Klf4, Oct3/4, Glis1, and EBNA1 from separate plasmids||An Efficient Non-viral Method to Generate Integration-Free Human iPS Cells from Cord Blood and Peripheral Blood Cells. Stem Cells. 2012 Nov 29.||Yamanaka|
|Replicating EBNA1 episome||Human||Non-integrating EBNA1-mediated polycistronic expression of human Sox2, KLF4, L-Myc, Lin28, OCT3/4, and shRNA against p53 in different gene/insert combinations.||A more efficient method to generate integration-free human iPS cells. Nat Methods. 2011 May;8(5):409-12.||Yamanaka|
|Replicating EBNA1 episome||Human||Non-integrating polycistronic expression of human Oct4, Klf4, Sox2, c-Myc, Nanog, Lin28, NR5A2, and microRNA 302/367 in three different combinations of reprogramming factors||Efficient germ-line transmission obtained with transgene-free induced pluripotent stem cells. Proc Natl Acad Sci U S A. 2014 Jul 22;111(29):10678-83.||Capecchi|
|Replicating EBNA1 episome||Human||Fluorescent-tagged EBNA1-mediated expression of human Oct3/4, shp53, Klf4, Sox2, L-Myc, Lin28 from separate non-integrating episomes to allow sorting and dosage tracking of reprogramming factors||Fluorescent tagged episomals for stoichiometric induced pluripotent stem cell reprogramming. Stem Cell Res Ther. 2017 Jun 5;8(1):132.||Zovein|
|Replicating EBNA1 episome||Human||Non-integrating EBNA1-mediated polycistronic expression of human Oct4, Sox2, Myc, Klf4, BCL2L1, and EBNA1 from three separate plasmids||A Facile Method to Establish Human Induced Pluripotent Stem Cells From Adult Blood Cells Under Feeder-Free and Xeno-Free Culture Conditions: A Clinically Compliant Approach. Stem Cells Transl Med. 2015 Mar 5. pii: sctm.2014-0214.||Cheng|
|RNA||Human||Reprogramming factors for delivering synthetic mRNAs encoding human Klf4, Oct4, Sox2, c-Myc, and Lin28A to somatic cells from separate plasmids||Highly Efficient Reprogramming to Pluripotency and Directed Differentiation of Human Cells with Synthetic Modified mRNA. Cell Stem Cell. 2010 Nov 5;7(5):618-30.||Rossi|
|RNA||Human||Non-integrating, polycistronic, self-replicating VEE RNA species expressing human Oct4, Klf4, Sox2, with either c-Myc or Glis1||Efficient generation of human iPSCs by a synthetic self-replicative RNA. Cell Stem Cell. 2013 Aug 1;13(2):246-54.||Dowdy|
|Adenovirus||Mouse||Non-integrating expression of mouse Sox2, Oct4, c-Myc, and Klf4 from four separate adenoviral plasmids||Induced Pluripotent Stem Cells Generated Without Viral Integration. Science. 2008 Nov 7;322(5903):945-9.||Hochedlinger|
|Lentivirus||Mouse||Doxycycline-inducible expression of mouse Oct4, Sox2, Klf4, and c-Myc from four separate lentiviral plasmids||Sequential expression of pluripotency markers during direct reprogramming of mouse somatic cells. Cell Stem Cell. 2008 Feb 7. 2(2):151-9.||Jaenisch|
|Lentivirus||Mouse||Polycistronic, doxycycline-inducible lentiviral vectors for the expression of mouse Oct4, Klf4, c-Myc, and Sox2||Reprogramming of murine and human somatic cells using a single polycistronic vector. Proc Natl Acad Sci U S A. 2009 Jan 6. 106(1):157-62.||Jaenisch|
|Lentivirus||Mouse||Doxycycline-inducible lentiviral expression of mouse Oct4, Sox2, Klf4, and c-Myc from four separate plasmids||Defining molecular cornerstones during fibroblast to iPS cell reprogramming in mouse. Cell Stem Cell. 2008 Mar 6. 2(3):230-40.||Hochedlinger|
|MMLV-derived Retrovirus||Mouse||Original set of Yamanaka factors for generating mouse iPS cells; retroviral expression of mouse Sox2, Oct3/4, Klf4, and c-Myc from separate plasmids||Induction of pluripotent stem cells from mouse embryonic and adult fibroblast cultures by defined factors. Cell. 2006 Aug 25. 126(4):663-76.||Yamanaka|
|PiggyBac||Mouse||Doxycycline-inducible piggyBac transposon reprogramming system; expression of mouse Oct4, Sox2, Klf4, and c-Myc from polycistronic cassettes||KLF4 N-terminal variance modulates induced reprogramming to pluripotency. Stem Cell Reports. 2015 Apr 14;4(4):727-43.||Woltjen|
|Plasmid||Mouse||Non-integrating polycistronic expression of mouse Oct4, Klf4, Sox2, and c-Myc from two separate plasmids||Generation of Mouse Induced Pluripotent Stem Cells Without Viral Vectors. Science. 2008 Nov 7;322(5903):949-53.||Yamanaka|
|Replicating EBNA1 episome||Mouse||Non-integrating EBNA1-mediated polycistronic expression of mouse Oct4, Sox2, Klf4, c-Myc, and Lin28||Efficient human iPS cell derivation by a non-integrating plasmid from blood cells with unique epigenetic and gene expression signatures. Cell Res. 2011 Mar;21(3):518-29.||Cheng|
Differentiation and Transdifferentiation Plasmids
Browse the table below for plasmids used for differentiation of iPSCs or transdifferntion of a somatic cell to a new type of somatic cell. Sort the table by cell type, delivery system, species, or PI and then select the article link to find more information.