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EDi001-A-2

AST23-1KO-3, AST22-1KO-3, AST-23_SCAKO Clone 3, AST-22_SNCAKO Clone 3

Gene-edited iPSC line

In Stock
At European Collection of Authenticated Cell Cultures (ECACC)
A CLIP contains information about a cell line including any specific third party obligations relating to, for example, licensing obligations or the donor consent which affect the use of the cell line.
A batch specific Certificate of Analysis will be available to download once you receive your EBiSC iPSC line.

General#

Cell Line
hPSCreg Name EDi001-A-2
Alternative name(s)
AST23-1KO-3, AST22-1KO-3, AST-23_SCAKO Clone 3, AST-22_SNCAKO Clone 3
Cell line type Human induced pluripotent stem cell (hiPSC)

Provider
Depositor University of Edinburgh (ED)
Owner College of Medicine and Veterinary Medicine
Distributors
EBiSC
European Collection of Authenticated Cell Cultures (ECACC)
Roslin Cells (RC)
Derivation country United States

External Databases
hPSCreg EDi001-A-2
BioSamples SAMEA3323846
Cellosaurus CVCL_LE52
ECACC 66540167
CLO CLO_0102713
Wikidata Q54831973

General Information
* Is the cell line readily obtainable for third parties?
Yes
Research use: allowed
Clinical use: allowed
Commercial use: allowed
Subclone of

Donor Information#

General Donor Information
Sex female

Phenotype and Disease related information (Donor)
Diseases A disease was diagnosed.
Parkinson disease
Parkinson disease in the OLS
This is a PD line, with the control being EDi002-A lines and CRISPR/Cas9-corrected EDi001-A-1, EDi001-A-2, EDi001-A-3 and EDi001-A-4
The donor is a carrier of a disease-associated mutation and affected.
Genetic variants
SNCA (target)
4q22.1
Heterozygous
The donor carries a triplication of the alpha-synuclein gene, resulting in 4 copies of SNCA. The copies of SNCA are situated in a heterozygous triplication configuration. See Figure 1 of Petrucci, 2015 for a graphic representation of the heterozygous triplication.
Family history Strong family history of Parkinson’s disease due to autosomal dominant inheritance of SNCA triplication
Is the medical history available upon request? Y Mov Disord. 2011 Sep;26(11):2134-6. doi: 10.1002/mds.23776

Donor Relations
Other cell lines of this donor
All cell lines of this donor's relatives
Has daughter:

External Databases (Donor)
BioSamples SAMEA3319991

hIPSC Derivation#

General
The source cell information can be found in the parental cell line EDi001-A.

Reprogramming method
Vector type Integrating
Vector Virus (Retrovirus)
Genes
Is the used vector excisable?
No
Absence of reprogramming vector(s)?
Unknown
Reprogramming vectors silenced?
Yes
Methods used
RT-PCR

Vector free reprogramming
Type of used vector free reprogramming factor(s)
None

Other
Selection criteria for clones Nickase pair mediated SNCA Knock-out of 1 SNCA alleles. Therefore 3 alleles remain.
Derived under xeno-free conditions
No
Derived under GMP?
No
Available as clinical grade?
No

Culture Conditions#

Latest released batch
Culture medium mTeSR
Passage method EDTA
Surface coating Matrigel / Geltrex
O2 concentration 20
CO2 concentration 5
Temperature 37
The following are the depositor culture conditions, they do not refer to any specific batch.
Surface coating Laminin
Passage method Enzymatically
Accutase
O2 Concentration 20 %
CO2 Concentration 5 %
Medium Essential 8™
Supplements
Rock inhibitor added while passaging 10 nM

Characterisation#

Analysis of Undifferentiated Cells
Marker Expressed Immunostaining RT-PCR FACS Enzymatic Assay Expression Profiles
SSEA-4
Yes
TRA 1-60
Yes
SSEA-1
No
Differentiation Potency
Endoderm
Ont Id: UBERON_0000925
In vitro spontaneous differentiation
Mesoderm
Ont Id: UBERON_0000926
In vitro spontaneous differentiation
Ectoderm
Ont Id: UBERON_0000924
In vitro spontaneous differentiation

Microbiology / Virus Screening
HIV 1 Negative
HIV 2 Negative
Hepatitis B Negative
Hepatitis C Negative
Mycoplasma Negative

Sterility
Inoculation for microbiological growth No Contaminants Detected
Mycoplasma Not Detected
Viability Viable post-cryopreservation

Genotyping#

Karyotyping (Cell Line)
Has the cell line karyotype been analysed?
No

Other Genotyping (Cell Line)

Genetic Modification#

Disease/phenotype related modifications
Parkinson disease
Parkinson disease in the OLS
Details: This is a PD line, with the control being NAS lines and CRISPR/Cas9-corrected AST22 lines
Genetic modifications
SNCA (target)
Gene knock-out
4q22.1
CRISPR/Cas was employed to create a mutation in Exon2, disrupting the ATG start site and some sequence 5' to the coding start of SNCA. PCR amplicons of the CRISPR site were cloned using the TOPO cloning technique. Sequencing of 8 TOPO clones detected 2 deletions and 1 insertion; however, sequencing of the CRISPR sites shows that only one of these modifications disrupts the ATG site. This indicates 1 allele was putatively knocked out, and 3 alleles remain. Additional comment: there are two additional ATG sites located immediately downstream of the normal ATG start site, one of which is in-frame (9 bases) to the initial ATG. It is possible that inactivation of the first ATG start site might not be completely sufficient to prevent translation from a second downstream ATG.
CRISPR-associated (CRISPR/Cas) System