Addgene - pF149 pSOD1G85RAcGFP1 Plasmid Data

Plasmid 26410: pF149 pSOD1G85RAcGFP1

Gene/insert name:
SOD1
Insert size:
470
Species:
H. sapiens (human)
Entrez Gene:
SOD1 (ALS, ALS1, IPOA, SOD, hSod1, homodimer)
Fusion protein or tag:
GFP
Terminal:
C terminal on insert
Mutation:
G85R
Vector backbone:
pAcGFP1-N1
(Search Vector Database)
Backbone manufacturer:
Clontech
Vector type:
Mammalian Expression
Backbone size w/o insert (bp):
4700
Cloning site 5':
EcoRI
Site destroyed during cloning:
No
Cloning site 3':
SalI
Site destroyed during cloning:
No
5' sequencing primer:
CMV-F List of Sequencing Primers
Bacterial resistance(s)
Kanamycin
Growth strain(s)
DH5alpha
Growth temperature (℃):
37
High or low copy:
Unknown
Selectable markers:
Neomycin
Person or lab that originally
cloned the gene/insert:
human cDNA insert from cDNA clone cSOD1 from Professor Yoram Groner as described in Lieman-Hurwitzet al., 1982 and Sherman et al 1983.
Sequence: View sequences (2)
Principal Investigator:
Elizabeth Fisher
Terms and Licenses:
MTA
Clontech Limited Use Label License

Comments:

The complete cDNA was subcloned into the vector pET28. Site directed mutagenesis changed wildtype DNA sequence for codon 85 of the cDNA from GGC to CGC, resulting in the G85R mutation in the protein and then a PCR amplified the complete cDNA with EcoRI and SalI sites on the 5’ and 3’ end respectively. This EcoRI-Sal1 0.47kb fragment was cloned into the respective EcoRI and SalI sites of pAcGFP1N1. Thus SOD1 is 5’ of GFP and SOD1 lies upstream of GFP in the SOD1-GFP fusion protein.

Addgene has sequenced a portion of this plasmid for verification. Click here for the sequencing result.

Article: Modification of superoxide dismutase 1 (SOD1) properties by a GFP tag--implications for research into amyotrophic lateral sclerosis (ALS). Stevens et al (PLoS One. 2010 . 5(3):e9541. PubMed)

Please acknowledge the principal investigator and cite this article if you use this plasmid in a publication. Also, please include the text "Addgene plasmid 26410" in your Materials and Methods section.