TCR CRISPR/Cas9 Lentivirus (Non-Integrating)

Catalog #

78062

$995 *

Size: 500 µl x 2

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Bulk/Custom

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Description

The T-Cell Receptor (TCR) is found on the surface of T-cells and is responsible for recognizing antigens bound to MHC (Major Histocompatibility Complex) molecules. Activation of the TCR results in activation of downstream NFAT signaling. The TCR consists of a heterodimer of two different protein chains, of which the alpha (α) and beta (β) chains are the predominant chains.

The TCR CRISPR Lentiviruses are replication incompetent, HIV-based VSV-G pseudo-typed lentiviral particles that are ready to be transduced 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 4 sgRNA (single guide RNA) targeting human TRAC (T-Cell Receptor Alpha Constant) and human TRBC1 (T-Cell Receptor Beta Constant 1) regions of the α and β chains.

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 PD-1 CRISPR/Cas9 Lentivirus.

Figure 1. Schematic of the Lenti-vector used to generate the TCR CRISPR/Cas9 Lentivirus.

Gene Target	Primer ID:	sgRNA Sequence
TRAC	TCR-1	AGAGTCTCTCAGCTGGTACA
TRAC	TCR-2	TGTGCTAGACATGAGGTCTAA
TRBC1	TCR-3	GGAGAATGACGAGTGGACCC
TRBC1	TCR-4	GCAGTATCTGGAGTCATTGA

Figure 2. List of sgRNA Sequences in the TCR CRISPR/Cas9 Lentivirus.

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Synonyms

SpCas9 lentivirus, CRISPR lentivirus, Cas9 lentivirus, T-Cell Receptor CRISPR

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Product Info

Storage and Usage

Citations

Species

Replication incompetent HIV

Supplied As

Two vials (500 μl x 2) of lentivirus at a titer ≥1 x 10⁶ TU/ml. The titer will vary with each lot; the exact value is provided with each shipment.

Formulation

The lentiviruses were produced from HEK293T cells in 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.

Instructions for Use

See assay protocol for detailed instructions.

Applications

1. Transient knock-down of TCR in a target cells pool.
2. Generation of stable TCR knock-out cells using transient puromycin selection (48h maximum) followed by limited dilution.

Shipping Temperature

-80°C

Notes

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

Biosafety
None of the HIV genes (gag, pol, rev) will be expressed in the transduced cells. Although the pseudotyped lentiviruses are replication-incompetent, they do require the use of a Biosafety Level 2 facility. BPS recommends following all federal, state, local, and institutional regulations and using all appropriate safety precautions.

Scientific Category

CRISPR

Regulatory Status

Research Use Only

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TCR CRISPR/Cas9 Lentivirus (Non-Integrating)

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