KC-4818

293T-ITGB6-ITGAV Cell Line

60761

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Background of 293T-ITGB6-ITGAV Cell Line

ITGB6 gene encodes a protein that is a member of the integrin superfamily. Members of this family are adhesion receptors that function in signaling from the extracellular matrix to the cell. Integrins are heterodimeric integral membrane proteins composed of an alpha chain and a beta chain. The encoded protein forms a dimer with an alpha v chain and this heterodimer can bind to ligands like fibronectin and transforming growth factor beta 1. Alternate splicing results in multiple transcript variants.
The product of ITGAV gene belongs to the integrin alpha chain family. Integrins are heterodimeric integral membrane proteins composed of an alpha subunit and a beta subunit that function in cell surface adhesion and signaling. The encoded preproprotein is proteolytically processed to generate light and heavy chains that comprise the alpha V subunit. This subunit associates with beta 1, beta 3, beta 5, beta 6 and beta 8 subunits. The heterodimer consisting of alpha V and beta 3 subunits is also known as the vitronectin receptor. This integrin may regulate angiogenesis and cancer progression. Alternative splicing results in multiple transcript variants. Note that the integrin alpha 5 and integrin alpha V subunits are encoded by distinct genes.

Specifications

Catalog Number	KC-4818
Cell Line Name	293T-ITGB6-ITGAV Cell Line
NCBI/UniProt Accession Number	NM_000888.5/NM_002210.5
Clone Number	2-7#
Host Cell Line	293T
Description	Stable 293T cell line expressing exogenous ITGB6 and ITGAV gene
Quantity	Two vials of frozen cells (≥2-10⁶/vial)
Stability	Stable in culture over a minimum of 10 passages
Application	Drug screening and biological assays
Freezing Medium	70% DMEM+20% FBS+10% DMSO
Propagation Medium	DMEM+10% FBS +1μg/mL Puromycin
Selection Marker	Puromycin
Morphology	Epithelial
Subculture	Split saturated culture 1:4-1:8 every 2-3 days; seed out at about 1-3 × 10⁵ cells/mL
Incubation	37 °C with 5% CO₂
Storage	Liquid nitrogen immediately upon receiving
Doubling Time	Approximately 30 hours
Mycoplasma Status	Negative

Cell Line Generation

293T-ITGB6-ITGAV cell line was generated using lentivirus expressing ITGB6 and ITGAV sequence.

Characterization

Figure 1. Characterization of ITGB6 over-expression in the 293T-ITGB6-ITGAV stable clone using FACS.(Primary antibody 1: Human IgG1 kappa,isotype control, Cat#KB-1273, Kyinno. Primary antibody 2: Sigvotatug, Cat#KB-1403, Kyinno.)

Figure 2. Characterization of ITGB6 and ITGAV over-expression in the 293T-ITGB6-ITGAV stable clone using PCR.

Cell Resuscitation

Prewarm culture medium (DMEM supplemented with 10% FBS, 1μg/mL Puromycin)in a 37°C water bath.
Thaw the frozen vial in a 37°C water bath for 1-2 minutes.
Transfer the vial into biosafety cabinet, and wipe the surface with 70% ethanol.
Unscrew the top of the vial and transfer the cell suspension gently into a sterile centrifuge tube containing 9.0mL complete culture medium.
Spin at ~ 125 × g for 5-7 minutes at room temperature, and discard the supernatant without disturbing the pellet.
Resuspend cell pellet with the appropriate volume of complete medium and transfer the cell suspension into a T25 culture flask.
Incubate the flask at 37°C, 5% CO₂ incubator.
Split saturated culture 1:4-1:8 every 2-3 days; seed out at about 1-3 × 10⁵ cells/mL.

Cell Freezing

Prepare the freezing medium (70% DMEM + 20% FBS + 10% DMSO) fresh immediately before use.
Keep the freezing medium on ice and label cryovials.
Transfer cells to a sterile, conical centrifuge tube, and count the cells.
Centrifuge the cells at 250×g for 5 minutes at room temperature and carefully aspirate off the medium.
Resuspend the cells at a density of at least 3×10⁶ cells/mL in chilled freezing medium.
Aliquot 1 mL of the cell suspension into each cryovial.
Freeze cells in the CoolCell freezing container overnight in a -80°C freezer.
Transfer vials to liquid nitrogen for long-term storage.

References

Zhang Y, Chen Z, Shen Z, Qian D, Wang G, Wang X, Xi S, Wang X. ITGB6 promotes pancreatic fibrosis and aggravates the malignant process of pancreatic cancer via JAK2/STAT3 signaling pathway. Naunyn Schmiedebergs Arch Pharmacol. 2024 Aug;397(8):6093-6106. doi: 10.1007/s00210-024-03003-z. Epub 2024 Feb 28. PMID: 38418753.
Xu M, Chen X, Yin H, Yin L, Liu F, Fu Y, Yao J, Deng X. Cloning and characterization of the human integrin β6 gene promoter. PLoS One. 2015 Mar 27;10(3):e0121439. doi: 10.1371/journal.pone.0121439. PMID: 25816241; PMCID: PMC4376883.
Inamine T, Nakamura M, Kawauchi A, Shirakawa Y, Hashiguchi H, Aiba Y, Taketomi A, Shirabe K, Nakamuta M, Hayashi S, Saoshiro T, Komori A, Yatsuhashi H, Kondo S, Omagari K, Maehara Y, Ishibashi H, Tsukamoto K; PBC Study Group in NHOSLJ. A polymorphism in the integrin αV subunit gene affects the progression of primary biliary cirrhosis in Japanese patients. J Gastroenterol. 2011 May;46(5):676-86. doi: 10.1007/s00535-010-0351-0. Epub 2010 Dec 1. PMID: 21116829.
Reuning U. Integrin αvβ3 promotes vitronectin gene expression in human ovarian cancer cells by implicating rel transcription factors. J Cell Biochem. 2011 Jul;112(7):1909-19. doi: 10.1002/jcb.23111. PMID: 21433063.