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Vector Backbone Description: Backbone Size:4731; Vector Backbone:pET; Vector Types:Bacterial Expression; Bacterial Resistance:Kanamycin
Comments: This plasmid is an empty vector to be used with a LIC cloning protocol.
It has a TEV cleavable His6-MBP fusion tag on the N-terminus and a Kan resistance.
To clone into this vector, add LIC fusion tags to the 5' end of your PCR primers.
Forward - 5'TACTTCCAATCCAATGCA3'
Reverse - 5'TTATCCACTTCCAATGTTATTA3'
Linearize the plasmid with SspI and gel purify.
When digesting the DNA with T4 polymerase for LIC, use dCTP for insert and dGTP for vector. The 13-series vectors were designed to enable rapid cloning for a co-expression system. Vectors are based on Novagen's Duet system. Each gene is expressed on its own vector, which has been optimized so that each gene expresses at approximately equal levels. The 13-series vectors are all compatible with 2-series transfer vectors, so if cotransformation fails, there is a readily available backup in polycistronic expression.
13S CDF origin SpecR13K ColA origin KanR2-series transfer ColE1 origin AmpR13S vectors must be cotransformed with a 2-series vector for optimal expression (that is, the presence of an empty 2A-T vector enhances expression of a gene in 13S). For triple expressions, the proteins all seem to express at approximately equal levels. Genes in the CDF vector consistently express at very slightly lower levels, so if you're trying to pull down stoichiometric complexes, it's probably best to put His6-fusion protein in this vector. More information on this vector can be found through http://qb3.berkeley.edu/qb3/macrolab/
Proper citation: RRID:Addgene_48317 Copy
Species: Synthetic
Genetic Insert: HYPER-RED-C199S
Vector Backbone Description: Backbone Size:3975; Vector Backbone:pC1; Vector Types:Mammalian Expression; Bacterial Resistance:Kanamycin
Defining Citation: PMID:25330925
Proper citation: RRID:Addgene_48252 Copy
Species: Escherichia coli CFT073
Genetic Insert: ClbP
Vector Backbone Description: Backbone Marker:Novagen; Backbone Size:5370; Vector Backbone:pET29b-(+); Vector Types:Bacterial Expression; Bacterial Resistance:Kanamycin
Defining Citation: PMID:23406518
Proper citation: RRID:Addgene_48244 Copy
Species: Escherichia coli CFT073
Genetic Insert: ClbP-pep
Vector Backbone Description: Backbone Marker:Novagen; Backbone Size:5370; Vector Backbone:pET29b-(+); Vector Types:Bacterial Expression; Bacterial Resistance:Kanamycin
Defining Citation: PMID:23406518
Proper citation: RRID:Addgene_48243 Copy
Species: Synthetic
Genetic Insert: ECFP
Vector Backbone Description: Backbone Marker:Life Technologies/Invitrogen; Backbone Size:2651; Vector Backbone:pDONRP2R-P3; Vector Types:Gateway Cloning; Bacterial Resistance:Kanamycin
Defining Citation: PMID:23957834
Proper citation: RRID:Addgene_48353 Copy
Vector Backbone Description: Backbone Size:5775; Vector Backbone:pXOON; Vector Types:Mammalian Expression, Xenopus Oocyte Expression; Bacterial Resistance:Kanamycin
Proper citation: RRID:Addgene_48699 Copy
Species: Caenorhabditis elegans
Genetic Insert: snf-5
Vector Backbone Description: Backbone Size:5052; Vector Backbone:pXOOM; Vector Types:Mammalian Expression, Xenopus Oocyte Expression; Bacterial Resistance:Kanamycin
Defining Citation: PMID:23580723
Proper citation: RRID:Addgene_48697 Copy
Species: Synthetic
Genetic Insert: AmCyan
Vector Backbone Description: Backbone Marker:clontech; Backbone Size:4000; Vector Backbone:pmR-mCherry; Vector Types:Mammalian Expression; Bacterial Resistance:Kanamycin
Comments: the nAmCyan and the mCherry genes are separated by a P2A sequence that results in 2 separate proteins. It also includes BsmBI restriction sites flanking the nAmCyan gene which result in BsiWI and BssHII overhangs for cloning in-frame with P2A-mCherry.
Proper citation: RRID:Addgene_48686 Copy
Genetic Insert: HIV Integrase
Vector Backbone Description: Vector Backbone:pET29a; Vector Types:Bacterial Expression; Bacterial Resistance:Kanamycin
Defining Citation: PMID:23691126
Comments: The depositing lab recommends bacteria strain Rosetta (DE3)pLysS for protein expression.
Please contact [email protected] for additional information about this plasmid.
Proper citation: RRID:Addgene_48683 Copy
Genetic Insert: PFV Integrase
Vector Backbone Description: Vector Backbone:pET29a; Vector Types:Bacterial Expression; Bacterial Resistance:Kanamycin
Defining Citation: PMID:23691126
Comments: The depositing lab recommends bacteria strain Rosetta (DE3)pLysS for protein expression.
Please contact [email protected] for additional information about this plasmid.
Proper citation: RRID:Addgene_48682 Copy
Species: Synthetic
Genetic Insert: sgRNA targeting GTCCCCTCCACCCCACAGTG, compatible with S. thermophilus #1 Cas9, hU6 promoter
Vector Backbone Description: Backbone Marker:Invitrogen; Vector Backbone:pCR-BluntII-TOPO; Vector Types:CRISPR; Bacterial Resistance:Kanamycin
Defining Citation: PMID:24076762
Proper citation: RRID:Addgene_48672 Copy
Species: Synthetic
Genetic Insert: TD prototspacer A/PAM with reporter EYFP
Vector Backbone Description: Vector Backbone:pSC101-kan; Vector Types:CRISPR; Bacterial Resistance:Kanamycin
Defining Citation: PMID:24076762
Proper citation: RRID:Addgene_48666 Copy
Species: Synthetic
Genetic Insert: NM prototspacer A/PAM with reporter EYFP
Vector Backbone Description: Vector Backbone:pSC101-kan; Vector Types:CRISPR; Bacterial Resistance:Kanamycin
Defining Citation: PMID:24076762
Proper citation: RRID:Addgene_48664 Copy
Species: Synthetic
Genetic Insert: TD prototspacer B/PAM with reporter EYFP
Vector Backbone Description: Vector Backbone:pSC101-kan; Vector Types:CRISPR; Bacterial Resistance:Kanamycin
Defining Citation: PMID:24076762
Proper citation: RRID:Addgene_48663 Copy
Species: Synthetic
Genetic Insert: SP/NM prototspacer B/PAM with reporter EYFP
Vector Backbone Description: Vector Backbone:pSC101-kan; Vector Types:CRISPR; Bacterial Resistance:Kanamycin
Defining Citation: PMID:24076762
Proper citation: RRID:Addgene_48661 Copy
Species: Homo sapiens
Genetic Insert: AQP9
Vector Backbone Description: Backbone Marker:Clontech; Backbone Size:4731; Vector Backbone:pEGFP-C1; Vector Types:Mammalian Expression; Bacterial Resistance:Kanamycin
Defining Citation: PMID:17346701
Proper citation: RRID:Addgene_48808 Copy
Genetic Insert: sfGFP and luxI
Vector Backbone Description: Vector Backbone:pZ vector; Vector Types:Bacterial Expression, Synthetic Biology; Bacterial Resistance:Kanamycin
Defining Citation: PMID:22178928
Comments: * The presence of glucose will reduce the amount of expression from the Lux promoter driving GFP and other genes. Omitting the glucose could potentially make the cell overburdened.
Proper citation: RRID:Addgene_48887 Copy
Genetic Insert: Sh ble
Vector Backbone Description: Backbone Marker:Life Technologies (Invitrogen); Backbone Size:2500; Vector Backbone:pDONR221; Vector Types:Gateway attB1 & attB2; Bacterial Resistance:Kanamycin
Defining Citation: PMID:19432966
Comments: Please note that Addgene's sequencing results found a few differences when compared to the full plasmid sequence provided by the depositing laboratory. According to the depositing laboratory, these differences are not a concern for the function of the plasmid.
Proper citation: RRID:Addgene_48995 Copy
Genetic Insert: aminoglycoside 3' phosphotransferase
Vector Backbone Description: Backbone Marker:Life Technologies (Invitrogen); Backbone Size:2500; Vector Backbone:pDONR221; Vector Types:Gateway attB1 & attB2; Bacterial Resistance:Kanamycin
Defining Citation: PMID:19432966
Comments: Please note that Addgene's sequencing results found a few differences when compared to the full plasmid sequence provided by the depositing laboratory. According to the depositing laboratory, these differences are not a concern for the function of the plasmid.
Proper citation: RRID:Addgene_48993 Copy
Species: Other
Genetic Insert: hygromycin phosphotransferase
Vector Backbone Description: Backbone Marker:Dankort Lab; Vector Backbone:pBEG R2+xL3 (Addgene plasmid 48952); Vector Types:Gateway Cloning; Bacterial Resistance:Kanamycin
Defining Citation: PMID:24146852
Proper citation: RRID:Addgene_48985 Copy
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