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Species: Homo sapiens
Genetic Insert: PSAM4 GlyR IRES eGFP
Vector Backbone Description: Vector Backbone:pCAG; Vector Types:Mammalian Expression; Bacterial Resistance:Ampicillin
Defining Citation: PMID:30872534
Proper citation: RRID:Addgene_119739 Copy
Species: Synthetic
Genetic Insert: CaMPARI2
Vector Backbone Description: Backbone Marker:Stratagene; Backbone Size:4267; Vector Backbone:pAAV; Vector Types:Mammalian Expression, AAV; Bacterial Resistance:Ampicillin
Defining Citation: PMID:30361563
Proper citation: RRID:Addgene_119723 Copy
Species: Mus musculus
Genetic Insert: Type I procollagen α2 chain
Vector Backbone Description: Backbone Size:5400; Vector Backbone:pcDNA3.1; Vector Types:Mammalian Expression; Bacterial Resistance:Ampicillin
Defining Citation: PMID:30287488
Comments: These plasmids were constructed after the G610C mouse model of OI in which the Gly610 residue (starting from the N-terminal end of the triple helix) was substituted with Cys.
These mouse type I procollagen plasmids were designed for immortalized and primary murine osteoblasts and fibroblasts. Their expression in other cells might cause significant disruption of assembly, folding and trafficking of type I procollagen, resulting in artifacts. Even in osteoblasts and fibroblasts a fraction of transfected chains, particularly proα2, might be trafficked/degraded as monomers instead of being integrated into procollagen heterotrimers. In our experience, the best evidence of normal behavior of the transfected chains is the appearance of fluorescent extracellular collagen fibers ~ 12 h after transfection. Fluorescent molecules should be incorporated into fibers even when the transfected chains contain the N-propeptide cleavage site, because the cleavage is highly conformation sensitive and always incomplete.
We recommend following these guidelines when using the plasmids
1) Only mouse cells that have high expression of endogenous type I procollagen are suitable for studies of physiologically-relevant processes with these constructs.
2) At least 100 μM ascorbic acid at and after transfection is required to ensure proper procollagen folding (for some cells, preincubation with ascorbic acid before transfection might be needed).
3) Optimization of transfection efficiency is needed for all cell types.
4) Experiments beyond 24h after transfection are not recommended, to avoid excessive accumulation of aggregates and cell malfunction caused by increased procollagen synthesis.
5) Lack of extracellular fluorescent fibers 12-24 h after transfection indicates improper procollagen synthesis/trafficking or cellular malfunction.
6) Excessive expression of transfected chains might cause rapid accumulation of large procollagen aggregates in the ER, resulting in cellular malfunction and death.
Contact information: Shakib Omari ([email protected]), Sergey Leikin ([email protected]).
Proper citation: RRID:Addgene_119827 Copy
Species: Photinus pyralis
Genetic Insert: Luciferase
Vector Backbone Description: Backbone Size:8116; Vector Backbone:pLentipuro3/TO/V5-DEST; Vector Types:Mammalian Expression, Lentiviral, Luciferase; Bacterial Resistance:Ampicillin
Defining Citation: PMID:30683938
Proper citation: RRID:Addgene_119816 Copy
Species: Other
Genetic Insert: ecTadA-ecTadA*(7.10)-nSaCas9 (KKH)
Vector Backbone Description: Vector Backbone:pCMV with a BR322 origin; Vector Types:Mammalian Expression; Bacterial Resistance:Ampicillin
Defining Citation: PMID:31110355
Proper citation: RRID:Addgene_119815 Copy
Species: Pseudomonas putida F1
Genetic Insert: CymR
Vector Backbone Description: Backbone Marker:Biedendieck et al., 2007; Vector Backbone:pRBBm34; Vector Types:Bacterial Expression, Synthetic Biology; Bacterial Resistance:Ampicillin
Defining Citation: PMID:30392122
Proper citation: RRID:Addgene_119872 Copy
Species: Mus musculus
Genetic Insert: beta Actin
Vector Backbone Description: Backbone Size:2905; Vector Backbone:PX551; Vector Types:Mouse Targeting, AAV, CRISPR; Bacterial Resistance:Ampicillin
Defining Citation: PMID:29056297
Proper citation: RRID:Addgene_119870 Copy
Species: Homo sapiens
Genetic Insert: Signal transducer and activator of transcription 1
Vector Backbone Description: Backbone Marker:invitrogen; Backbone Size:6000; Vector Backbone:pcDNA3-GFP; Vector Types:Mammalian Expression; Bacterial Resistance:Ampicillin
Comments: Please note hSTAT1 in this plasmid contains the mutations A188T and Q718R compared to GenBank BC002704. The impact of these mutations on plasmid functionality is unknown.
Proper citation: RRID:Addgene_11987 Copy
Species: Homo sapiens
Genetic Insert: MMP7
Vector Backbone Description: Backbone Marker:invitrogen; Backbone Size:6000; Vector Backbone:pcDNA3-GFP; Vector Types:Mammalian Expression; Bacterial Resistance:Ampicillin
Proper citation: RRID:Addgene_11989 Copy
Vector Backbone Description: Backbone Size:7298; Vector Backbone:pCIG3; Vector Types:Mammalian Expression, Lentiviral; Bacterial Resistance:Ampicillin
Defining Citation: PMID:31594807
Comments: Please note that though there are some discrepancies between the backbone sequence from the depositing lab and Addgene's quality control sequence, this plasmid should function as described.
Proper citation: RRID:Addgene_119863 Copy
Species: Synthetic
Genetic Insert: cuOx144 array
Vector Backbone Description: Backbone Size:4904; Vector Backbone:pDEST-Bxb1attB-loxP-PGK-puro-loxP; Vector Types:Mouse Targeting; Bacterial Resistance:Ampicillin
Defining Citation: PMID:31124784
Comments: Please note there are a few ambiguous bases in Addgene's NGS result. This is due to NGS having difficulty assembling repetitive sequences.
The repetitive sequences in this plasmid can be unstable while propagated in bacteria. It is recommended that the repetitive insert be verified by restriction digest and repeat counts be viewed as an approximation.
Please visit https://www.biorxiv.org/content/10.1101/409672v1 for bioRxiv preprint.
Proper citation: RRID:Addgene_119903 Copy
Species: Danio rerio
Genetic Insert: myr-tagged tdTomato and SV40 polyA, followed by FRT site-flanked Kanamycin selection gene
Vector Backbone Description: Backbone Size:3121; Vector Backbone:pGEM-T; Vector Types:Bacterial Expression; Bacterial Resistance:Ampicillin
Defining Citation: PMID:32084358
Proper citation: RRID:Addgene_119867 Copy
Species: Danio rerio
Genetic Insert: NLS-Cre, 2A, mCherry and SV40 polyA, followed by FRT site-flanked Kanamycin selection gene
Vector Backbone Description: Backbone Size:3008; Vector Backbone:pGEM-T; Vector Types:Bacterial Expression; Bacterial Resistance:Ampicillin
Defining Citation: PMID:32084358
Proper citation: RRID:Addgene_119868 Copy
Species: Mycobacteriophage Bxb1
Genetic Insert: Mammalian codon optimized Bxb1 integrase
Vector Backbone Description: Backbone Size:4905; Vector Backbone:pCAGGs; Vector Types:Mammalian Expression; Bacterial Resistance:Ampicillin
Defining Citation: PMID:31124784
Comments: Please visit https://www.biorxiv.org/content/10.1101/409672v1 for bioRxiv preprint.
Proper citation: RRID:Addgene_119901 Copy
Species: Synthetic
Genetic Insert: HA-tdMS2cp-tagRFP-Tx2-nls
Vector Backbone Description: Backbone Size:3526; Vector Backbone:ePiggyBac-UbC; Vector Types:Mammalian Expression; Bacterial Resistance:Ampicillin
Defining Citation: PMID:31124784
Comments: Please visit https://www.biorxiv.org/content/10.1101/409672v1 for bioRxiv preprint.
Proper citation: RRID:Addgene_119908 Copy
Species: Synthetic
Genetic Insert: TetRFlag-nls-tdTom
Vector Backbone Description: Backbone Size:5670; Vector Backbone:ePiggyBac-CAG; Vector Types:Mammalian Expression; Bacterial Resistance:Ampicillin
Defining Citation: PMID:31124784
Comments: Plasmid contains ICP22 NLS with a single aa change that does not affect function. Sequence: RRPAVRSPPLGTRKRK.
Please visit https://www.biorxiv.org/content/10.1101/409672v1 for bioRxiv preprint.
Proper citation: RRID:Addgene_119909 Copy
Species: Synthetic
Genetic Insert: iGlul
Vector Backbone Description: Backbone Marker:EMD Biosciences; Backbone Size:5933; Vector Backbone:pET41a; Vector Types:Bacterial Expression; Bacterial Resistance:Kanamycin
Defining Citation: PMID:29735711
Proper citation: RRID:Addgene_119832 Copy
Species: Synthetic
Genetic Insert: iGluh
Vector Backbone Description: Backbone Marker:EMD Biosciences; Backbone Size:5933; Vector Backbone:pET41a; Vector Types:Bacterial Expression; Bacterial Resistance:Kanamycin
Defining Citation: PMID:29735711
Proper citation: RRID:Addgene_119830 Copy
Species: Homo sapiens
Genetic Insert: GAG FMLV fused to spCAS9
Vector Backbone Description: Vector Backbone:pCDNA3-like; Vector Types:Mammalian Expression; Bacterial Resistance:Ampicillin
Defining Citation: PMID:30604748
Comments: Please visit https://www.biorxiv.org/content/early/2017/10/12/202010 for BioRxiv preprint. Note, there are some additional amino acids on the 3' end of the insert--depositor confirmed this is the version used in the paper and the plasmid should function as described.
Proper citation: RRID:Addgene_119942 Copy
Species: Homo sapiens
Genetic Insert: PAX8
Vector Backbone Description: Backbone Marker:promega; Backbone Size:4818; Vector Backbone:pGL3-Basic; Vector Types:Mammalian Expression; Bacterial Resistance:Ampicillin
Proper citation: RRID:Addgene_11994 Copy
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