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Genetic Insert: ECFP(loxP)MCS
Vector Backbone Description: Backbone Size:4100; Vector Backbone:pCS2+; Vector Types:Mammalian Expression, Cre/Lox, Flp/FRT; Bacterial Resistance:Ampicillin
Defining Citation: PMID:11376165
Comments: Cre and Flp reporter: from blue protein fluorescence to gene of interested inserted into multiple cloning site (MCS)
Proper citation: RRID:Addgene_31377 Copy
Genetic Insert: Traffic Light Reporter 2.1 (V2468 zfn target) EF Puro
Vector Backbone Description: Backbone Size:5000; Vector Backbone:pCVL; Vector Types:Lentiviral; Bacterial Resistance:Ampicillin
Defining Citation: PMID:21743461
Proper citation: RRID:Addgene_31483 Copy
Genetic Insert: Traffic Light Reporter 1.1 (Sce Target) EF Puro
Vector Backbone Description: Backbone Size:5000; Vector Backbone:pCVL; Vector Types:Lentiviral; Bacterial Resistance:Ampicillin
Defining Citation: PMID:21743461
Proper citation: RRID:Addgene_31482 Copy
Species: Caenorhabditis elegans
Genetic Insert: C. elegans glutamate-gated chloride channel alpha, crystallized construct
Vector Backbone Description: Backbone Marker:Invitrogen; Backbone Size:4800; Vector Backbone:pFastbac1; Vector Types:Insect Expression; Bacterial Resistance:Ampicillin
Defining Citation: PMID:21572436
Proper citation: RRID:Addgene_31488 Copy
Species: Synthetic
Genetic Insert: pHRed
Vector Backbone Description: Backbone Marker:Invitrogen; Backbone Size:2760; Vector Backbone:pRsetB; Vector Types:Bacterial Expression; Bacterial Resistance:Ampicillin
Defining Citation: PMID:21631110
Proper citation: RRID:Addgene_31472 Copy
Species: Homo sapiens
Genetic Insert: preimplantation protein 3
Vector Backbone Description: Backbone Size:3460; Vector Backbone:pQStrep2; Vector Types:Bacterial Expression; Bacterial Resistance:Ampicillin
Defining Citation: PMID:15998469
Comments: Please note that there is a small gap between Addgene's quality control sequence and the reference sequence provided by the depositor. The gap is downstream of the ORF and should not affect function.
Proper citation: RRID:Addgene_31592 Copy
Genetic Insert: SFFV d14 GFP donor EF1s I-SceI
Vector Backbone Description: Backbone Size:5092; Vector Backbone:pCVL; Vector Types:Lentiviral; Bacterial Resistance:Ampicillin
Defining Citation: PMID:21743461
Proper citation: RRID:Addgene_31476 Copy
Genetic Insert: SFFV d14GFP
Vector Backbone Description: Backbone Size:5516; Vector Backbone:pCVL; Vector Types:Lentiviral; Bacterial Resistance:Ampicillin
Defining Citation: PMID:21743461
Proper citation: RRID:Addgene_31475 Copy
Species: Homo sapiens
Genetic Insert: RB shRNA
Vector Backbone Description: Backbone Size:13700; Vector Backbone:pSLIK-EGFP; Vector Types:Mammalian Expression, Lentiviral, RNAi; Bacterial Resistance:Ampicillin
Comments: The whole sequence for the shRNA including the loop should be from 3872-3931 in the vector: AGCGAAACGATTATCCATTCAAATAGTGAAGCCACAGATGTATTTGAAT
The shRNA is downstream of the EGFP
GGATAATCGTT
pSLIK vector from Ian Frasier
Proper citation: RRID:Addgene_31500 Copy
Species: Homo sapiens
Genetic Insert: Actin related protein 2/3 complex, subunit 3
Vector Backbone Description: Backbone Size:3460; Vector Backbone:pQStrep2; Vector Types:Bacterial Expression; Bacterial Resistance:Ampicillin
Defining Citation: PMID:15998469
Comments: The vector pQTEV is derived from Qiagen pQE-2 and contains an inactive chloramphenicol resistance gene sequence.
Please note that Addgene's quality control sequence shows a small deletion in the vector region downstream of insert; the deletion does not affect any features in this construct.
Proper citation: RRID:Addgene_31580 Copy
Genetic Insert: lactose repressor protein
Vector Backbone Description: Backbone Size:4540; Vector Backbone:pEMBL9'; Vector Types:Bacterial Expression; Bacterial Resistance:Ampicillin
Defining Citation: PMID:11266612
Comments: Parent plasmid was pEMBL9' with insert of extended LacI sequence from pIQ (Hare & Sadler, Gene 3 (1978) 269-278.
For expression: E. coli BLIM cells (or any E. coli WITHOUT lactose repressor in the genome or accompanying plasmid)
Proper citation: RRID:Addgene_31490 Copy
Species: T7 phage
Genetic Insert: RNA polymerase
Vector Backbone Description: Backbone Size:5400; Vector Backbone:pBAD33; Vector Types:Bacterial Expression; Bacterial Resistance:Chloramphenicol
Defining Citation: PMID:10610690
Comments: For expression: E. coli CV2
N823D mutation should not affect plasmid function.
Proper citation: RRID:Addgene_31491 Copy
Species: Drosophila melanogaster
Genetic Insert: Rab5
Vector Backbone Description: Backbone Marker:Drosophila Gateway Collection; Vector Backbone:pAGW; Vector Types:Insect Expression; Bacterial Resistance:Ampicillin
Defining Citation: PMID:21169635
Proper citation: RRID:Addgene_31733 Copy
Species: Drosophila melanogaster
Genetic Insert: Rab8
Vector Backbone Description: Backbone Marker:Drosophila Gateway Collection; Vector Backbone:pAGW; Vector Types:Insect Expression; Bacterial Resistance:Ampicillin
Defining Citation: PMID:21169635
Proper citation: RRID:Addgene_31735 Copy
Species: Drosophila melanogaster
Genetic Insert: Rab6
Vector Backbone Description: Backbone Marker:Drosophila Gateway Collection; Vector Backbone:pAGW; Vector Types:Insect Expression; Bacterial Resistance:Ampicillin
Defining Citation: PMID:21169635
Proper citation: RRID:Addgene_31734 Copy
Species: Drosophila melanogaster
Genetic Insert: Rab10
Vector Backbone Description: Backbone Marker:Drosophila Gateway Collection; Vector Backbone:pAGW; Vector Types:Insect Expression; Bacterial Resistance:Ampicillin
Defining Citation: PMID:21169635
Proper citation: RRID:Addgene_31737 Copy
Species: Homo sapiens
Genetic Insert: ALK5
Vector Backbone Description: Backbone Size:4716; Vector Backbone:pRK5; Vector Types:Mammalian Expression; Bacterial Resistance:Ampicillin
Comments: This construct is beginning from a Signal Peptide of human TbRI/ALK5, followed by a HA tag, the extracellular cDNA sequence of human TbRI/ALK, and ended by a Flag tag. This vector to produces a secreted TbRI/ALK5 extracellular form, with two individual tags on each side.
Proper citation: RRID:Addgene_31720 Copy
Vector Backbone Description: Backbone Marker:Khavari Lab; Backbone Size:11100; Vector Backbone:LZRS-RfA; Vector Types:Retroviral; Bacterial Resistance:Chloramphenicol and Ampicillin
Defining Citation: PMID:15342384
Proper citation: RRID:Addgene_31601 Copy
Genetic Insert: BoNT/A LC
Vector Backbone Description: Backbone Size:0; Vector Backbone:pBN3; Vector Types:Bacterial Expression; Bacterial Resistance:Ampicillin
Defining Citation: PMID:18940613
Comments: The author states that although the construct is from a toxic protein, it only contains one third of the biologically active protein,i.e., it is inactive and harmless per se, and it does not require BL4 level for its expression/manipulation.
Toxic BoNT/A is made of two 'chains' (i.e. polypeptides) linked to each other by one disulfide bond. The light chain (LC) is 50KDa long, whereas the heavy chain (HC) is 100 KDa. The HC is necessary for the toxin to target and enter motorneurons. This construct is the LC of BoNTA, and it completely lacks the HC, so it is unable to intoxicate neurons.
Protein sequence of BoNT/A fragment present in this plasmid (corresponds to YP_001386738.1)
MSGLEVSFEELRTFGGHDAKFIDSLQENEFRLYYYNKFKDIASTLNKAKSIVGTTASLQYMKNVFKEKYLLSEDTSGKFSVDKLKFDKLYKMLTEIYTEDNFVKFFKVLNRKTYLNFDKAVFKINIVPKVNYTIYDGFNLRNTNLAANFNGQNTEINNMNFTKLKNFT
Nucleotide sequence of BoNT/A fragment present in this plasmid based on Addgene's Sanger sequencing results:
ATGAGTGGGTTAGAAGTAAGCTTTGAGGAACTTAGAACATTTGGGGGACATGATGCAAAGTTTATAGATAGTTTACAGGAAAACGAATTTCGTCTATATTATTATAATAAGTTTAAAGATATAGCAAGTACACTTAATAAAGCTAAATCAATAGTAGGTACTACTGCTTCATTACAGTATATGAAAAATGTTTTTAAAGAGAAATATCTCCTATCTGAAGATACATCTGGAAAATTTTCGGTAGATAAATTAAAATTTGATAAGTTATACAAAATGTTAACAGAGATTTACACAGAGGATAATTTTGTTAAGTTTTTTAAAGTACTTAACAGAAAAACATATTTGAATTTTGATAAAGCCGTATTTAAGATAAATATAGTACCTAAGGTAAATTACACAATATATGATGGATTTAATTTAAGAAATACAAATTTAGCAGCAAACTTTAATGGTCAAAATACAGAAATTAATAATATGAATTTTACTAAACTAAAAAATTTTACT
Proper citation: RRID:Addgene_31602 Copy
Species: Homo sapiens
Genetic Insert: MCU
Vector Backbone Description: Backbone Marker:Invitrogen; Backbone Size:2787; Vector Backbone:pDONR223; Vector Types:pDONR Gateway; Bacterial Resistance:Spectinomycin
Defining Citation: PMID:21685886
Comments:
*A3T aa residue substitution in MCU, which is not known to affect protein function
Proper citation: RRID:Addgene_31726 Copy
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