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PDE5A KO cell line available to order. KO validated by Next Generation Sequencing. Free of charge wild type control provided.

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Images

Western blot - Human PDE5A knockout A549 cell line (AB301140), expandable thumbnail
  • Next Generation Sequencing - Human PDE5A knockout A549 cell line (AB301140), expandable thumbnail

Key facts

Cell type
A549
Species or organism
Human
Tissue
Lung
Form
Liquid
Knockout validation
Next Generation Sequencing

Alternative names

5''-cyclic phosphodiesterase, CGB-PDE, CN5A, CN5N, PDE 5, PDE5A1, PDE5A_HUMAN, Phosphodiesterase 5A, Phosphodiesterase 5A cGMP specific, Phosphodiesterase isozyme 5, cGMP binding cGMP specific 3' 5' cyclic nucleotide phosphodiesterase, cGMP binding cGMP specific 35 cyclic nucleotide phosphodiesterase, cGMP binding/cGMP specific phosphodiesterase, cGMP specific 3'5' cyclic phosphodiesterase, cGMP specific phosphodiesterase PDE5A2, cGMP specific phosphodiesterase type 5A, cGMP-binding cGMP-specific phosphodiesterase, cGMP-specific 3''

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PDE5A KO cell line available to order. KO validated by Next Generation Sequencing. Free of charge wild type control provided.

Key facts

Cell type
A549
Form
Liquid
Disease
Carcinoma
Concentration
Loading...

Properties

Gene name
PDE5A
Gene editing type
Knockout
Gene editing method
CRISPR technology
Knockout validation
Next Generation Sequencing

Cell culture

Biosafety level
EU: 1 US: 1
Adherent/suspension
Adherent
Gender
Male

Handling procedures

Initial handling guidelines

Upon arrival, the vial should be stored in liquid nitrogen vapor phase and not at -80°C. Storage at -80°C may result in loss of viability.

1. Thaw the vial in 37°C water bath for approximately 1-2 minutes.
2. Transfer the cell suspension (0.8 mL) to a 15 mL/50 mL conical sterile polypropylene centrifuge tube containing 8.4 mL pre-warmed culture medium, wash vial with an additional 0.8 mL culture medium (total volume 10 mL) to collect remaining cells, and centrifuge at 201 x g (rcf) for 5 minutes at room temperature. 10 mL represents minimum recommended dilution. 20 mL represents maximum recommended dilution.
3. Resuspend the cell pellet in 5 mL pre-warmed culture medium and count using a haemocytometer or alternative cell counting method seed all remaining cells into a T25.
4. Incubate the culture at 37°C incubator with 5% CO2. Check the culture one day after revival and continue to check until 80% confluent. Media change can be given if needed.
5. Once confluent passage into an appropriate flask at a density of 2x104 cells/cm2. Seeding density is given as a guide only and should be scaled to align with individual lab schedules. Cultures should be monitored daily.

Subculture guidelines
  • All seeding densities should be based on cell counts gained by established methods.
  • A guide seeding density of 2x104 cells/cm2 is recommended.
  • Cells should be passaged when they have achieved 80-90% confluence.
  • Do not allow the cell density to exceed 7x104 cells/cm2.
Cryopreservation medium
Cell Freezing Medium-DMSO Serum free media, contains 8.7% DMSO in MEM supplemented with methyl cellulose.

Storage

Shipped at conditions
Dry Ice
Appropriate short-term storage conditions
-196°C
Appropriate long-term storage conditions
-196°C

Notes

Although we aim to provide customers with a homozygous clone, feasibility will be dependent on the biology of the protein. Should only heterozygous edits be achieved, you will be notified of the outcome and be asked to confirm whether the cell line is acceptable. All clones will be accompanied with DNA sequencing data, and the mutation description.

Recommended control: Human wild-type A549 cell line (ab288558). Please note a wild-type cell line is not automatically included with a knockout cell line order, if required please add recommended wild-type cell line at no additional cost using the code WILDTYPE-TMTK1.

We will provide viable cells that proliferate on revival.

This product is subject to limited use licenses from The Broad Institute and ERS Genomics Limited, and is developed with patented technology. For full details of the limited use licenses and relevant patents please refer to our limited use license and patent pages.

Supplementary info

This supplementary information is collated from multiple sources and compiled automatically.
Activity summary

PDE5A also known as PDE5 or phosphodiesterase type 5 plays a critical role in hydrolyzing cyclic guanosine monophosphate (cGMP) to its inactive form. This enzyme is part of the phosphodiesterase family and has a molecular mass of approximately 100 kDa. PDE5A is expressed mainly in the smooth muscle cells of blood vessels and in the corpus cavernosum of the penis. Other expression sites include the lung heart and skeletal muscle.

Biological function summary

PDE5A regulates intracellular levels of cGMP an important secondary messenger in various signaling pathways. It is not part of a large protein complex but directly influences the contractility of smooth muscle tissues. By controlling cGMP levels PDE5A modulates vascular tone and influences processes such as vasodilation. This regulation is essential for maintaining a balance in the contractile state of muscles impacting blood flow and other vital functions.

Pathways

PDE5A contributes significantly to the nitric oxide (NO)-cGMP signaling pathway which is important for vascular relaxation. In this pathway nitric oxide stimulates guanylate cyclase to produce cGMP which then activates specific kinases and ion channels. Related proteins such as guanylate cyclase and cGMP-dependent protein kinases work alongside PDE5A to mediate smooth muscle relaxation and vasodilation particularly in penile erectile tissue.

Associated diseases and disorders

PDE5A shows strong connections to erectile dysfunction and pulmonary hypertension. In the case of erectile dysfunction impaired regulation of cGMP by PDE5A affects the relaxation of smooth muscle leading to inadequate blood flow. Pulmonary hypertension involves elevated blood pressure in the lung arteries which PDE5A influences through its role in controlling vascular tone. The enzyme guanylate cyclase associates closely with PDE5A in these contexts as both influence cGMP pathways involved in these conditions.

Product promise

We are dedicated to supporting your work with high quality reagents and we are here for you every step of the way should you need us.

In the unlikely event of one of our products not working as expected, you are covered by our product promise.

Full details and terms and conditions can be found here:
Terms & Conditions.

2 product images

  • Western blot - Human PDE5A knockout A549 cell line (ab301140), expandable thumbnail

    Western blot - Human PDE5A knockout A549 cell line (ab301140)

    Western blot: Anti-PDE5A antibody [EPR24129-95] (Anti-PDE5A/PDE5 antibody [EPR24129-95] ab259945) staining at 1/1000 dilution, shown in green; Mouse anti-Alpha Tubulin [DM1A] (Anti-alpha Tubulin antibody [DM1A] - Loading Control ab7291) loading control staining at 1/20000 dilution, shown in magenta. In Western blot, Anti-PDE5A/PDE5 antibody [EPR24129-95] ab259945 was shown to bind specifically to PDE5A. A band was observed at 95 kDa in wild-type A549 cell lysates with no signal observed at this size in PDE5A knockout cell line. To generate this image, wild-type and PDE5A knockout A549 cell lysates were analysed. First, samples were run on an SDS-PAGE gel then transferred onto a nitrocellulose membrane. Membranes were blocked in 3 % milk in TBS-0.1 % Tween® 20 (TBS-T) before incubation with primary antibodies overnight at 4 °C. Blots were washed four times in TBS-T, incubated with secondary antibodies for 1 h at room temperature, washed again four times then imaged. Secondary antibodies used were Goat anti-Rabbit IgG H&L 800CW and Goat anti-Mouse IgG H&L 680RD at 1/20000 dilution.

    All lanes: Western blot - Anti-PDE5A/PDE5 antibody [EPR24129-95] (Anti-PDE5A/PDE5 antibody [EPR24129-95] ab259945) at 1/1000 dilution

    Lane 1: Wild-type A549 cell lysate at 20 µg

    Lane 2: Western blot - Human PDE5A knockout A549 cell line (ab301140)

    Lane 2: PDE5A knockout A549 cell lysate at 20 µg

    Lane 3: K562 cell lysate at 20 µg

    Secondary

    Lanes 1 - 3: Goat anti-Rabbit IgG H&L 800CW at 1/20000 dilution

    Lanes 1 - 3: Goat anti-Mouse IgG H&L 680RD at 1/20000 dilution

    Performed under reducing conditions.

    Predicted band size: 100 kDa

    Observed band size: 95 kDa

  • Next Generation Sequencing - Human PDE5A knockout A549 cell line (ab301140), expandable thumbnail

    Next Generation Sequencing - Human PDE5A knockout A549 cell line (ab301140)

    217 bp deletion after Ser 77 (allele 1); 205 bp deletion after Leu 82 (allele 2) of WT protein

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Product protocols

For this product, it's our understanding that no specific protocols are required. You can:

Please note: All products are 'FOR RESEARCH USE ONLY. NOT FOR USE IN DIAGNOSTIC OR THERAPEUTIC PROCEDURES'.

For licensing inquiries, please contact partnerships@abcam.com