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hTERT-HPNE E6/E7/K-RasG12D
hTERT-HPNE E6/E7/K-RasG12D
規格:
價格:
編號:B241993
品牌:Mingzhoubio

標準菌株
定量菌液
DNA
RNA

規格:
凍干粉
斜面
甘油
平板


產品名稱
hTERT-HPNE E6/E7/K-RasG12D
商品貨號
B241993
Organism
Homo sapiens, human
Tissue
pancreas, duct
培養基
Product Format
frozen
Morphology
epithelial-like
Culture Properties
adherent
Biosafety Level
2  [Cells contain Human Papilloma (HPV16) 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.

Age
52 years
Gender
male
Storage Conditions
liquid nitrogen vapor phase
Karyotype
This is a pseudodiploid human cell line of male origin. Clonal aberrations included the derivative chromosomes: t(3;18)(p21;q11.2) [balanced translocation], del(6)(q15), add(8)(q11.2) and der(21)t(17;21)(q21;p11.2). The percentage of cells with the normal male chromosome complement increased at high passage and non-clonal aberrations were seen in approximately 20% of the examined cells at all passages.
Derivation
Method: developed by infection of hTERT-HPNE E6/E7 cells (ATCC CRL-4036) with retroviral vector (pLXSN) carrying a G12D mutant of the isoform b of human K-Ras
Clinical Data
male
Antigen Expression
positive for nestin (flow cytometry) (verified at ATCC)
Genes Expressed
positive for nestin (flow cytometry)(verified at ATCC)
Tumorigenic
NO
Complete Growth Medium
The base medium for this cell line is:
  • 75% DMEM without glucose (Sigma Cat#. D-5030 with additional 2 mM L-glutamine and 1.5 g/L sodium bicarbonate)
  • 25% Medium M3 Base (Incell Corp. Cat# M300F- 500)
To make the complete growth medium, add the following components to the base medium:
  • fetal bovine serum 5% (final conc.)
  • 10 ng/ml human recombinant EGF
  • 5.5 mM D-glucose (1g/L)
  • 750 ng/ml puromycin

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.
  1. Remove and discard culture medium.
  2. Briefly rinse the cell layer with Ca++/Mg++ free Dulbecco's phosphate-buffered saline (D-PBS) or 0.25% (w/v) Trypsin - 0.53 mM EDTA solution to remove all traces of serum which contains trypsin inhibitor.
  3. Add 2.0 to 3.0 mL of 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°C to facilitate dispersal.
  4. Add 2.0 to 3.0 mL of complete growth medium and aspirate cells by gently pipetting.
  5. Transfer cell suspension to a centrifuge tube and spin at approximately 125 X g for 5 to 10 minutes. Discard supernatant.
  6. Resuspend the cell pellet in fresh growth medium. Add appropriate aliquots of the cell suspension to new culture vessels. An inoculum of 4 X 103 to 6 X 103 viable cells/cm2 is recommended.
  7. Incubate cultures at 37°C. Subculture when cell density reaches between 5 X 104 and 6 X 104 cells/cm2.
Subcultivation ratio: 1:8 to 1:12 twice weekly
Medium renewal: every 2 to 3 days
Cryopreservation
Freeze medium: fetal bovine serum (FBS), 90%; DMSO, 10%
Storage temperature: liquid nitrogen vapor phase
Culture Conditions
Temperature: 37°C
Atmosphere: air, 95%; carbon dioxide (CO2), 5%
STR Profile
CSF1PO: 12
D13S317: 12, 13
D16S539: 12, 13
D5S818: 11
D7S820: 9, 10
TH01: 8, 9
TPOX: 8, 11
vWA: 17
Amelogenin: XY
Population Doubling Level (PDL)

Longevity: >15 PDLs post-cryopreservation recovery

Population Doubling Time
approximately 29 hours
Name of Depositor
M Ouellette
Year of Origin
July 2002
References

Lee KM. et al. Immortalization with telomerase of the Nestin-positive cells of the human pancreas. Biochem. Biophys. Res. Commun. 301(4):1038-1044, 2003. Pubmed: 12589817

Lee KM. et al. Notch2-positive progenitors with the intrinsic ability to give rise to pancreatic ductal cells. Lab. Invest. 85 (8): 1003-1012, 2005. Pubmed: 15924149

Campbell PM, et al. K-Ras promotes growth transformation and invasion of immortalized human pancreatic cells by Raf and phosphatidylinositol 3-kinase signaling. Cancer Res. 67(5): 2098-2106, 2007. PubMed: 17332339

Campbell PM, et al. Ras-driven transformation of human nestin-positive pancreatic epithelial cells. Methods Enzymol. 439: 451-465, 2008. PubMed: 18374182

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