CRBN CRISPR/Cas9 Lentivirus (Non-Integrating)

Catalog #

78518

$995 *

Size: 500 µl x 2

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Bulk/Custom

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*US Pricing only. For international pricing, please contact your local distributor.

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Description

Cereblon (CRBN) forms an E3 ubiquitin ligase complex which is responsible for ubiquitinating proteins that regulate various developmental processes. CRBN also binds to Calcium Activated Potassium Channel subunit alpha-1 (KCNMA1) to regulate ion transport. Moreover, mutations in CRBN may play an underlying role in tumor cells acquiring resistance to immunotherapy.

The CRBN CRISPR/Cas9 Lentiviruses are replication incompetent, HIV-based VSV-G pseudo-typed lentiviral particles that are ready to transduce into almost all types of mammalian cells, including primary and non-dividing cells. The particles contain a CRISPR/Cas9 gene driven by an EF1A promoter, along with 5 sgRNA (single guide RNA) targeting human CRBN.

The non-integrating lentivirus is made with a mutated integrase, resulting in only transient expression of the Cas9 and sgRNA. Although using the non-integrating lentivirus results in lower knockdown efficiency, the Cas9 isn’t permanently expressed, which lowers the risk of off-targeting, and there are no random integrations into the cell’s genome. Knockout cell lines can still be generated following cell sorting or limited dilution, because even though the Cas9 and sgRNA expression is transient, the changes in the genomic DNA from the Cas9 nuclease activity and NHEJ repair are permanent.

Figure 1: Schematic of the lenti-vector used to generate the CRBN CRISPR/Cas9 Lentivirus.

Gene Target:	sgRNA Sequence:
CRBN	ACCAATGTTCATATAAATGG
CRBN	CTGACTGTGTTCTTAGCTCA
CRBN	TTACATACTGTATGTGATGT
CRBN	TTCTAATTGAACTGCAGACA
CRBN	TCAAGAAACAGCTACGTGAA

Table 1: List of sgRNA Sequences in the CRBN CRISPR/Cas9 Lentivirus.

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Product Info

Storage and Usage

Citations

Supplied As

Two vials (500 µl x 2) of lentivirus at a titer ≥1 x 10⁷ TU/ml. The titer will vary with each lot; the exact value will be provided with each shipment.

Formulation

The lentivirus particles were produced from HEK293T cells. They are supplied in cell culture medium containing 90% DMEM + 10% FBS.

Storage/Stability

Lentiviruses are shipped with dry ice. For long-term storage, it is recommended to store the lentiviruses at -80°C for up to 12 months from date of receipt. Avoid repeated freeze/thaw cycles. Titers can drop significantly with each freeze/thaw cycle.

Applications

Transient knockdown of CRBN in target cell pools
Generation of stable CRBN knockout cells following transient puromycin selection (48h maximum) followed by limiting dilution cloning.

Shipping Temperature

-80°C

Notes

Biosafety
The lentiviruses are produced with the SIN (self-inactivation) lentivector which ensures self-inactivation of the lentiviral construct after transduction. None of the HIV genes (gag, pol, rev) will be expressed in the transduced cells, as they are expressed from packaging plasmids lacking the packing signal and are not present in the lentivirus particle. Although the pseudotyped lentiviruses are replication-incompetent, they require the use of a Biosafety Level 2 facility. BPS Bioscience recommends following all local federal, state, and institutional regulations and using all appropriate safety precautions.

Troubleshooting Guide
Visit bpsbioscience.com/lentivirus-faq for detailed troubleshooting instructions. For all further questions, please email [email protected].

License Disclosure
The CRISPR/CAS9 technology is covered under numerous patents, including U.S. Patent Nos. 8,697,359 and 8,771,945, as well as corresponding foreign patents applications, and patent rights.

Warnings

Avoid freeze/thaw cycles

Scientific Category

CRISPR

Regulatory Status

Research Use Only

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CRBN CRISPR/Cas9 Lentivirus (Non-Integrating)

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