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http://www.addgene.org/63802

Species: Drosophila melanogaster
Genetic Insert: SP-dCas9-VPR
Vector Backbone Description: Vector Backbone:pAWG; Vector Types:Insect Expression, CRISPR, Synthetic Biology; Bacterial Resistance:Ampicillin
Defining Citation: PMID:25730490
Comments: Please note that this plasmid contains a stop codon between dCas9-VPR and eGFP and does NOT produce a GFP fusion protein. Cas9 contains D839A and N863A mutations. Depositor states that these mutations should not affect function.

Proper citation: RRID:Addgene_63802 Copy   

•   Source: Addgene

http://www.addgene.org/63800

Species: Homo sapiens
Genetic Insert: dCas9-VPR
Vector Backbone Description: Vector Backbone:PB-TRE; Vector Types:Mammalian Expression, CRISPR, Synthetic Biology, Piggybac; Bacterial Resistance:Ampicillin
Defining Citation: PMID:25730490

Proper citation: RRID:Addgene_63800 Copy   

•   Source: Addgene

http://www.addgene.org/63798

Species: Homo sapiens
Genetic Insert: SP-dCas9-VPR
Vector Backbone Description: Vector Backbone:pcDNA3.3 TOPO; Vector Types:Mammalian Expression, CRISPR, Synthetic Biology; Bacterial Resistance:Ampicillin
Defining Citation: PMID:25730490
Comments: Cas9 insert contains K604R, D839A and N863A mutations relative to reference sequence WP_010922251.1. Depositor states that these mutations should not affect function. This plasmid contains a human codon optimized form of cas9.

Proper citation: RRID:Addgene_63798 Copy   

•   Source: Addgene

http://www.addgene.org/63799

Species: Homo sapiens
Genetic Insert: ST1-dCas9-VPR
Vector Backbone Description: Vector Backbone:pcDNA3.3 TOPO; Vector Types:Mammalian Expression, CRISPR, Synthetic Biology; Bacterial Resistance:Ampicillin
Defining Citation: PMID:25730490
Comments: Cas9 insert contains K604R, D839A and N863A mutations relative to reference sequence WP_010922251.1. Depositor states that these mutations should not affect function.

Proper citation: RRID:Addgene_63799 Copy   

•   Source: Addgene

http://www.addgene.org/63850

Species: Synthetic
Genetic Insert: Theo45 theophylline riboswitch
Vector Backbone Description: Vector Backbone:pFTV1; Vector Types:Synthetic Biology; Bacterial Resistance:Chloramphenicol
Defining Citation: PMID:26621913

Proper citation: RRID:Addgene_63850 Copy   

•   Source: Addgene

http://www.addgene.org/63889

Vector Backbone Description: Backbone Size:3000; Vector Backbone:pBluescript; Vector Types:Bacterial Expression, phage display; Bacterial Resistance:Ampicillin
Defining Citation: PMID:1896445
Comments: pComb8 is nearly identical to the original pComb3 vector but contains the phage pVIII fusion protein for multi-valent expression along the sides of the phage via fusion with pVIII. It was designed for phage display of Fabs which are cloned into two separate cloning cassettes one chain at a time using SacI/XbaI restriction sites (for the light chain) and XhoI/SpeI restriction sites (for the heavy chain). The heavy chain is expressed fused to the pVIII. Both cassettes have the pelB leader sequence, the light chain and heavy chain-geneVIII fusion product assemble in the periplasm to form the Fab fragment. Soluble Fab can be expressed by removing the gene for the pVIII phage fusion protein by SpeI/NheI digest. Sufficient soluble protein is also found in the periplasmic space as a result of proteolysis.

Proper citation: RRID:Addgene_63889 Copy   

•   Source: Addgene

http://www.addgene.org/63885

Species: Synthetic
Genetic Insert: ER-localized low-affinity GCaMP3(10.19)
Vector Backbone Description: Backbone Marker:OTTC/NIDA; Vector Backbone:pOTTC374 - pAAV EF1a DIO iRFP; Vector Types:AAV; Bacterial Resistance:Ampicillin
Defining Citation: PMID:26451944

Proper citation: RRID:Addgene_63885 Copy   

•   Source: Addgene

http://www.addgene.org/63888

Vector Backbone Description: Backbone Size:3000; Vector Backbone:pBluescript; Vector Types:Bacterial Expression, phage display; Bacterial Resistance:Ampicillin
Defining Citation: PMID:1896445
Comments: The original pComb3 vector was designed for phage display of Fabs which are cloned into two separate cloning cassettes one chain at a time using SacI/XbaI restriction sites (for the light chain) and XhoI/SpeI restriction sites (for the heavy chain). The heavy chain is expressed fused to the pVIII. Both cassettes have the pelB leader sequence, the light chain and heavy chain-geneIII fusion product assemble in the periplasm to form the Fab fragment. This vector system has also been used for the display of a wide variety of other proteins like zinc fingers, peptides, and cDNA fragments. Phage can be produced which express Fab or other proteins or peptides of interest fused to the phage pIII protein for expression on the head of the phage. Soluble Fab can be expressed by removing the gene for the pIII phage fusion protein by SpeI/NheI digest. Sufficient soluble protein is also found in the periplasmic space as a result of proteolysis.

Proper citation: RRID:Addgene_63888 Copy   

•   Source: Addgene

http://www.addgene.org/63881

Species: Mus musculus
Genetic Insert: Tet1 promoter fragment A
Vector Backbone Description: Backbone Size:4807; Vector Backbone:pGL3-Basic; Vector Types:Mammalian Expression, Luciferase; Bacterial Resistance:Ampicillin
Defining Citation: PMID:25582196

Proper citation: RRID:Addgene_63881 Copy   

•   Source: Addgene

http://www.addgene.org/63935

Vector Backbone Description: Vector Backbone:pST50Trc; Vector Types:Bacterial Expression; Bacterial Resistance:Ampicillin
Defining Citation: PMID:15766881
Comments: Replace DHFR with GOI using BamHI/NgoMIV; Clone out cassette using XbaI/BamHI

Proper citation: RRID:Addgene_63935 Copy   

•   Source: Addgene

http://www.addgene.org/63893

Species: Mus musculus
Genetic Insert: Nanog homeobox
Vector Backbone Description: Backbone Marker:Promega; Backbone Size:5256; Vector Backbone:pGL3-Control; Vector Types:Mammalian Expression; Bacterial Resistance:Ampicillin
Defining Citation: PMID:22020437
Comments: Addgene's quality control sequence with EBV-rev primer shows a single bp mismatch with the available reference sequence.

Proper citation: RRID:Addgene_63893 Copy   

•   Source: Addgene

http://www.addgene.org/63890

Species: Synthetic
Genetic Insert: SS Stuffer
Vector Backbone Description: Backbone Size:3300; Vector Backbone:pComb3H; Vector Types:Bacterial Expression, phage display; Bacterial Resistance:Ampicillin
Defining Citation: PMID:10986398
Comments: pComb3X is the newest of the pComb vectors. Improvements over pComb3 include increased stability and introduction of an asymmetric SfiI cassette for directional cloning of full Fab, scFv, peptide and other protein for phage display. 6xHis and HA tags allow for purification and detection. An amber stop codon was introduced to turn-off expression of the pIII fusion protein by switching to a non-supressor strain of E. coli allowing production of soluble protein without subcloning. Alternatively, the gene for phage protein pIII can be removed by SpeI/NheI digest. pComb3XSS is recommended for preparation of vector for library cloning. The “SS” refers to the double stuffer, a 1200bp stuffer in the Fab light chain cloning region bounded by SacI and XbaI restriction sites and a 300bp stuffer in Fab heavy chain cloning region bound by XhoI and SpeI restriction sites. Also, the 1600bp double stuffer (both stuffers plus the leader sequence between the Fab light chain and heavy chain cloning regions) can be removed by SfiI digest so that non-Fab genes of interest can be cloned. Also available on Addgene: pComb3XTT and pComb3XLambda are only needed at templates for the construction of chimeric Fab libraries as described in Phage Display: A Laboratory Manual. pComb3XTT can also be used as an Fab expression control. Select References: Barbas, C. F., III; Burton, D. R.; Scott, J.K., Silverman, G.J. Eds. (2001) Phage Display: A Laboratory Manual; Cold Spring Harbor Laboratory Press: Cold Spring Harbor, New York Rader, C; Popkov, M.; Neves, J.A.; Barbas III, C.F. (2002) Integrin _v_3 Targeted Therapy of Kaposi’s Sarcoma with an In Vitro Evolved Antibody. FASEB, 16(14):2000-2. Berry, J.D.; Rutherford, J.; Silverman, G.J.; Kaul, R.; Elia, M.; Gobuty, S.; Fuller, R.; Plummer, F.A.; & Barbas III, C.F. (2003) Development of Functional Human Monoclonal Single-Chain Variable Fragment Antibody Against HIV-1 From Human Cervical B cells. Hybridoma and Hybridomics, 22(2):, 97-108. Jendreyko N, Popkov M, Beerli RR, Chung J, McGavern DB, Rader C, Barbas CF 3rd. (2003) Intra-diabodies: Bispecific, tetravalent antibodies for the simultaneous functional knockout of two cell surface receptors. J Biol Chem., 278(48):, 47812-9. Steinberger, P.; Sutton, J.K.; Rader, C.; Elia, M.; and Barbas III, C. F. (2000) Generation and Characterization of a Recombinant Human CCR5-specific Antibody: A Phage Display Approach for Rabbit Antibody Humanization. J. Biol. Chem., 275,, 36073-36078. Goncalves, J.; Kilva, F.; Freitas-Vieira, A.; Santa-Marta, M.; Malho, R.; Yang, X.; Gabuzda, D.; and Barbas III, C.F. (2002) Functional Neutralization of HIV-1 Vif Protein by Intracellular Immunization Inhibits Reverse Transcription and Viral Replication. J. Biol. Chem., 277(35):32036-45. Popkov, M.; Mage, R.G.; Alexander, C.B.; Thundivalappil, S.; Barbas III, C.F.; Rader, C. (2003) Rabbit immune repertoires as sources for therapeutic monoclonal antibodies: The impact of Kappa Allotype-correlated variation in cysteine content on antibody libraries selected by phage display. J. Mol. Biol., 325:325-335. Chung, J.; Rader, C.; Popkov, M.; Hur, Y.-M.; Kim, H.-K.; Lee, Y.-J.; & Barbas III, C. F. (2004) Integrin _IIb_3 specific synthetic human monoclonal antibodies and HCDR3 peptides that potently inhibit platelet aggregation,FASEB 18(2):361-3. Popkov, M.; Jendreyko, N.; Gonzalez-Sapienza, G.; Mage, R.G.; Rader, C.; Barbas III, C.F. (2004) Human/mouse cross-reactive anti-VEGF receptor 2 recombinant antibodies selected from animmune b9 allotype rabbit antibody library, J. Immunol. Methods, 288(1-2):149-164. Popkov, M.; Rader, C.; Barbas III, C.F. (2004) Isolation of human prostate cancer reactive antibodies using phage display technology, J. Immunol. Methods, 291:137-151. Tanaka, F.; Fuller, R.; Barbas III, C.F. (2005) Development of Small Designer Aldolase Enzymes: Catalytic Activity, Folding, and Substrate Specificity. Biochemistry, 44:7583-7592. This plasmid has been found to be prone to multimerization. Multimerization often does not impact plasmid function, but may reduce transformation efficiencies. See our article on Plasmid Dimers and Multimers for more information (https://blog.addgene.org/plasmids-101-dimers-and-multimers)

Proper citation: RRID:Addgene_63890 Copy   

•   Source: Addgene

http://www.addgene.org/64049

Species: Mus musculus
Genetic Insert: GLUT4
Vector Backbone Description: Backbone Size:9124; Vector Backbone:pLenti-hiko; Vector Types:Mammalian Expression, Lentiviral; Bacterial Resistance:Ampicillin
Defining Citation: PMID:25575350
Comments: The c-Myc sequence was inserted into the region encoding the 1st exofacial loop of mouse GLUT4 in the background of pmCherry-N1. The fragment of Myc-GLUT4-mCherry was then sub-cloned into pLenti-hiko vector via NheI + HpaI digestion. Discrepancies between the Addgene QC sequence and the full sequence are not of functional concern. Please cite the following article: Lim, C.-Y. et al. Tropomodulin3 is a novel Akt2 effector regulating insulin-stimulated GLUT4 exocytosis through cortical actin remodeling. Nat Commun 6, 5951 (2015).

Proper citation: RRID:Addgene_64049 Copy   

•   Source: Addgene

http://www.addgene.org/64048

Species: Saccharomyces cerevisiae
Genetic Insert: Lifeact
Vector Backbone Description: Backbone Size:9124; Vector Backbone:pLenti-hiko; Vector Types:Mammalian Expression, Lentiviral; Bacterial Resistance:Ampicillin
Defining Citation: PMID:25575350
Comments: The Lifeact sequence was initially reported in Riedl, J. et al. Lifeact: a versatile marker to visualize F-actin. Nat Methods 5, 605–607 (2008). Please cite the following article as follows: Lim, C.-Y. et al. Tropomodulin3 is a novel Akt2 effector regulating insulin-stimulated GLUT4 exocytosis through cortical actin remodeling. Nat Commun 6, 5951 (2015).

Proper citation: RRID:Addgene_64048 Copy   

•   Source: Addgene

http://www.addgene.org/64053

Genetic Insert: see comments
Vector Backbone Description: Vector Backbone:N/A; Vector Types:; Bacterial Resistance:Ampicillin
Defining Citation: PMID:24452804
Comments: Escherichia coli MG1655 mutS_fs dnaG_Q576A exoX- xonA- xseA- 1255700::tolQRA Δ(ybhB-bioAB)::[λcI857 N(cro-ea59)::tetR-bla] tolC-

Proper citation: RRID:Addgene_64053 Copy   

•   Source: Addgene

http://www.addgene.org/64060

Genetic Insert: tubulin
Vector Backbone Description: Vector Backbone:L304; Vector Types:Mammalian Expression, Lentiviral; Bacterial Resistance:Ampicillin
Defining Citation: PMID:23126280
Comments: Addgene sequencing results identified a duplication of the EGFP-Tubulin insert. It is not known how this impacts plasmid function.

Proper citation: RRID:Addgene_64060 Copy   

•   Source: Addgene

http://www.addgene.org/64072

Vector Backbone Description: Backbone Marker:Gene Transfer Vector Core University of Iowa; Backbone Size:5679; Vector Backbone:pacAd5; Vector Types:Adenoviral; Bacterial Resistance:Ampicillin
Defining Citation: PMID:25337876

Proper citation: RRID:Addgene_64072 Copy   

•   Source: Addgene

http://www.addgene.org/64070

Species: Mus musculus
Genetic Insert: BICD2N (1-400)
Vector Backbone Description: Backbone Size:5000; Vector Backbone:pOmniBac; Vector Types:Insect Expression; Bacterial Resistance:Gentamicin
Defining Citation: PMID:24986880

Proper citation: RRID:Addgene_64070 Copy   

•   Source: Addgene

http://www.addgene.org/64008

Vector Backbone Description: Vector Backbone:pET3aTR; Vector Types:Bacterial Expression; Bacterial Resistance:Ampicillin
Defining Citation: PMID:15766881

Proper citation: RRID:Addgene_64008 Copy   

•   Source: Addgene

http://www.addgene.org/64086

Species: Vesicular Stomatitis Virus
Genetic Insert: VSV matrix protein
Vector Backbone Description: Backbone Size:4795; Vector Backbone:pCAGEN; Vector Types:Mammalian Expression; Bacterial Resistance:Ampicillin

Proper citation: RRID:Addgene_64086 Copy   

•   Source: Addgene