PHKB Knockout cell line (HEK293)

Catalog Number: KO07978

Specifications KO Solutions KO Cell Line Workflow Handling procedures Downloads Related products

Specifications

Product Information
Product Name	PHKB Knockout cell line (HEK293)
specification	1*10^6
Storage and transportation	Dry ice preservation/T25 live cell transportation.
Cell morphology	Epithelioid, adherent cell
Passage ratio	1:3~1:6
species	Human
Gene	PHKB
Gene ID	5257
Build method	Electric rotation method / virus method
Mycoplasma testing	Negative
Cultivation system	90%DMEM+10% FBS
Parental Cell Line	HEK293
Quality Control	Genotype: PHKB Knockout cell line (HEK293) >95% viability before freezing. All cells were tested and found to be free of bacterial, viruses,mycoplasma and other toxins.

Gene Information
Gene Official Full Name	phosphorylase kinase regulatory subunit betaprovided by HGNC
Gene Description	Phosphorylase kinase is a polymer of 16 subunits, four each of alpha, beta, gamma and delta. The alpha subunit includes the skeletal muscle and hepatic isoforms, encoded by two different genes. The beta subunit is the same in both the muscle and hepatic isoforms, encoded by this gene, which is a member of the phosphorylase b kinase regulatory subunit family. The gamma subunit also includes the skeletal muscle and hepatic isoforms, encoded by two different genes. The delta subunit is a calmodulin and can be encoded by three different genes. The gamma subunits contain the active site of the enzyme, whereas the alpha and beta subunits have regulatory functions controlled by phosphorylation. The delta subunit mediates the dependence of the enzyme on calcium concentration. Mutations in this gene cause glycogen storage disease type 9B, also known as phosphorylase kinase deficiency of liver and muscle. Alternatively spliced transcript variants encoding different isoforms have been identified in this gene. Two pseudogenes have been found on chromosomes 14 and 20, respectively.[provided by RefSeq, Feb 2010]
Expression	Ubiquitous expression in thyroid (RPKM 18.4), adrenal (RPKM 12.6) and 25 other tissues See more

KO Solutions

We develop gene knockout solutions tailored to customer requirements and the condition of the target gene.

Editing Tools

Cas9 Protein

Cas9 mRNA sgRNA

Cas9 Plasmid

Cas9 Virus

A – Exon KO

gRNAs are designed in the introns flanking the exon, targeting non-multiple-of-3 base deletions in the exon, resulting in frameshift mutations.

B - Frameshift KO

gRNAs are designed within the exon, creating non-multiple-of-3 base deletions to induce frameshift mutations.

C - Complete KO

The entire gene coding sequence is deleted, achieving large-scale knockout effects.

KO Cell Line Workflow

KO Strategy Design

CRISPR Plasmid/Lentiviral Vector Construction

Lentiviral Packaging

Cell Transfection/Lentiviral Infection

Drug Selection

Cell Cryopreservation

Quality Control

Sequencing Validation

Monoclonal Cell Line Generation

Pool Efficiency Validation

Handling procedures

Cell Thawing

Pre-warm complete culture medium in a 37°C water bath.
Thaw the cryovial in a 37°C water bath for 1–2 minutes.
Transfer the cryovial to a biosafety cabinet (BSC) and wipe the surface with 70% ethanol.
Loosen the cap and gently transfer the cell suspension into a sterile centrifuge tube containing 9 mL of complete medium.
Centrifuge at 125 × g for 5–7 min at room temperature (RT), then discard the supernatant.
Resuspend the cell pellet in 5 mL of complete medium and transfer the suspension into a T25 flask.
Incubate the cells at 37°C in a 5% CO₂ incubator.
Recommended subculturing ratio: 1:2 to 1:3, reaching confluency in 2–3 days.

Cell Passaging

When cell confluence reaches 80–90%, proceed with passaging.
Pre-warm complete medium, PBS, and trypsin (0.25% Trypsin-EDTA, Gibco 25200-056) in a 37°C water bath. Once near 37°C, spray the bottles with 75% ethanol and place them in the BSC.
Retrieve the culture flask from the incubator, spray with 75% ethanol, and transfer it to the BSC.
To avoid dislodging cells, gently rinse the monolayer with PBS along the upper wall of the flask. Discard the PBS after washing (use 2 mL for T25 flasks).
Add an appropriate volume of trypsin (1.5 mL for T75, 0.5 mL for T25) and gently tilt the flask to ensure full coverage. Adjust the volume as needed. After 1–2 min, when most cells detach, neutralize digestion by adding an equal volume of complete medium. Gently pipette with a 5 mL serological pipette to ensure complete detachment.
Transfer the cell suspension to a 15 mL centrifuge tube and spin at 300 × g for 5 min. Discard the supernatant.
Resuspend the pellet in 5 mL of complete medium, adjust the seeding ratio as required, and replenish the flask with fresh medium (13–15 mL for T75, 5 mL for T25). Add 1% penicillin-streptomycin (dual antibiotic).
Tighten the cap, gently swirl to mix, and place the flask in a 37°C, 5% CO₂ incubator.

Cell Cryopreservation

Prepare cryopreservation medium in advance and pre-chill.
Ensure cells meet freezing criteria: healthy morphology, late-log phase growth, and absence of contamination or senescence (verify under a microscope).
Digest and centrifuge the cells (refer to the passaging protocol).
Resuspend the cell pellet in cryopreservation medium (1 mL per vial), mix thoroughly, and aliquot into cryovials.
Place the vials in a freezing container and store at −80°C overnight.
For long-term storage, transfer the cryovials to a liquid nitrogen tank.

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Datasheet

Please note that all services are for research use only. Not intended for any clinical use.

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