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CAR47

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價格:詢價

編號:B179760

品牌:Mingzhoubio

標準菌株

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DNA

RNA

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凍干粉

斜面

甘油

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產品名稱

CAR47

商品貨號

B179760

Organism

Homo sapiens, human

Tissue

adrenal gland

培養基

Cell Type

fibroblast

Product Format

frozen

Morphology

fibroblast-like

Culture Properties

adherent

Biosafety Level

2 [Cells contain SV40 viral DNA sequences]

Biosafety classification is based on U.S. Public Health Service Guidelines, it is the responsibility of the customer to ensure that their facilities comply with biosafety regulations for their own country.

Disease

primary pigmented adrenocortical disease (PPNAD) and Carney complex

Age

adult

Gender

female

Applications

This an immortalized cell line bearing a naturally occurring PRKA1A-inactivating mutation that is associated with multiple tumor formation derived from the adrenal tissue of a patient with Carney complex. PRKAR1A is a tumor suppressor gene. Applications include PKA associated characterization of cAMP-mediated mechanisms of cell cycle, signaling, proliferation and endocrine tumorigenesis.

Storage Conditions

liquid nitrogen vapor phase

Images

Derivation

The CAR47 cell line was derived from the adrenal gland that was removed from a patient diagnosed with PPNAD and Carney complex. After the first three to four passages, all cells in the growing cell lines had a fibroblastoid appearance, whereas both polygonal adrenocortical cells and fibroblasts were apparent at the beginning. At passage 10, the cells were transduced with a recombinant lentivirus expressing the large T antigen of SV40. The cell line was further cultured for several passages in the presence of blasticidin to ensure maintenance of the integrated lentivral cassette. In addition to the large T antigen, the integrated cassette also contains the gene that confers blasticidin resistance. The cells were initially frozen at passage 20.

Clinical Data

adult
female
patient diagnosed with PPNAD, Carney complex, and Cushing syndrome

Comments

The CAR47 cell line was derived from an adrenal gland that was removed from a patient diagnosed with PPNAD and Carney complex. This is an immortalized cell line with a naturally-occurring inactivating mutation in PRKAR1A, the regulatory subunit type 1A (R1alpha) of protein kinase A (PKA), which is associated with tumor formation.

PKA isozyme balance is critical for the control of cAMP signaling and related cell cycle and proliferation changes. Aberrant cAMP signaling has been linked to adrenocortical and other, mostly endocrine, tumors. Inactivating mutations in the PRKAR1A gene are a known cause of Carney complex - an autosomal dominant multiple neoplasia syndrome associated with skin, heart, and other myxomas and a variety of endocrine tumors.

Complete Growth Medium

The base medium for this cell line is ATCC-formulated DMEM Medium Catalog No. 30-2002. To make the complete growth medium, add the following components to the base medium:

10.0 µg/mL Blasticidin

Fetal bovine serum to a final concentration of 20%

Subculturing

Volumes used in this protocol are for 75 cm² flasks; proportionally reduce or increase amount of dissociation medium for culture vessels of other sizes.

Remove and discard culture medium.
Briefly rinse the cell layer with Ca⁺⁺/Mg⁺⁺ free Dulbecco's phosphate-buffered saline (D-PBS) or 0.05% (w/v) Trypsin - 0.02% EDTA solution to remove all traces of serum which contains trypsin inhibitor.
Add 1.0 to 2.0 mL of 0.05% Trypsin-EDTA solution to flask and observe cells under an inverted microscope until cell layer is dispersed (usually within 5 to 15 minutes). Note: To avoid clumping do not agitate the cells by hitting or shaking the flask while waiting for the cells to detach. Cells that are difficult to detach may be placed at 37.0°C to facilitate dispersal.
Add 6.0 to 8.0 mL of complete growth medium and aspirate cells by gently pipetting.
Transfer cell suspension to a centrifuge tube and spin at approximately 125 x g for 5 to 10 minutes. Discard supernatant.
Resuspend the cell pellet in fresh growth medium. Add appropriate aliquots of the cell suspension to new culture vessels.

Subcultivation Ratio: A subcultivation ratio of 1:3 to 1:5 is recommended
Medium Renewal: 2 to 3 times per week
Seeding Density: 1.0 x 10⁴ to 3.0 x 10⁴ cells/cm²

Cryopreservation

Freeze Medium: Complete growth medium, 70%; FBS, 20%; DMSO, 10%
Storage Temperature: Liquid nitrogen vapor phase

Culture Conditions

Atmosphere: air, 95%; carbon dioxide (CO₂), 5%
Temperature: 37°C

STR Profile

Amelogenin: X
CSF1PO: 10
D13S317: 12,14
D16S539: 11
D5S818: 11,12
D7S820: 8,11
TH01: 7,9.3
TPOX: 8,11
vWA: 17,18

Functional Tests

validation of PRKA1A-inactivating mutation

Name of Depositor

M. Nesterova

Year of Origin

2005

References

Nesterova M, et al. An immortalized human cell line bearing a PRKAR1A-inactivating mutation: Effects of overexpression of the wild-type allele and other protein kinase A subunits. J. Clin. Endocrinol. Metab. 93: 565-571, 2008. PubMed: 18056771

Kirschner LS, et al. Genetic heterogeneity and spectrum of mutations of the PRKAR1A gene in patients with the Carney complex. Human Mol. Gen. 9: 3037-3046, 2000. PubMed: 11115848

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