KC-6278

AsPC-1-VEGFA-KO Cell Line

61265

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Background of AsPC-1-VEGFA-KO Cell Line

VEGFA (vascular endothelial growth factor A), encoded by the VEGFA gene (Gene ID: 7422), is a key member of the PDGF/VEGF growth factor family. It encodes a heparin-binding disulfide-linked homodimer that specifically induces the proliferation and migration of vascular endothelial cells, playing an indispensable role in both physiological and pathological angiogenesis. Physiologically, it is critical for embryonic blood vessel formation, while pathologically, its upregulation is closely associated with tumor progression, diabetic microvascular complications, and inflammatory responses such as those induced by SARS-CoV-2 infection. Alternative splicing and translation initiation of VEGFA generate multiple isoforms with distinct biological activities, including proangiogenic and antiangiogenic variants. As a well-studied therapeutic target, VEGFA has been extensively investigated in various diseases.

Specifications

Catalog Number	KC-6278
Cell Line Name	AsPC-1-VEGFA-KO Cell Line
NCBI/UniProt Accession Number	7422
Clone Number	1A1
Host Cell Line	AsPC-1
Description	Stable AsPC-1 clone with human VEGFA gene knockout
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% RPMI1640+20% FBS+10% DMSO
Propagation Medium	RPMI1640+10% FBS
Selection Marker	N/A
Morphology	Epithelial
Subculture	Split saturated culture 1:3-1:6 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 46 hours
Mycoplasma Status	Negative

Cell Line Generation

AsPC-1-VEGFA-KO cell line was generated using the CRISPR method.

Characterization

Figure 1: Characterization of AsPC-1-VEGFA-KO cell line stable clone using PCR sequencing.

Figure 2: Characterization of AsPC-1-VEGFA-KO cell line stable clone using RT-PCR sequencing.

Cell Resuscitation

Prewarm culture medium (RPMI1640+10% FBS)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 1:3-1:6 every 2-3 days; seed out at about 1-3 × 10⁵ cells/mL.

Cell Freezing

Prepare the freezing medium (70% RPMI1640 + 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

Dabhi B, Mistry KN, Patel H, Lal S. Vascular endothelial growth factor insertion/deletion gene polymorphism in West Indian patients of type 2 diabetes and diabetic nephropathy. Indian J Biochem Biophys. 2015 Apr;52(2):209-12. PMID: 26118134.