规格
Cloning Method:	Gateway®
Constitutive or Inducible System:	Inducible
Delivery Method:	Lentiviral
Product Line:	BLOCK-iT™,Gateway®
Product Size:	20 reactions
Promoter:	H1⁄TO
RNAi Type:	shRNA
Selection Agent (Eukaryotic):	Blasticidin,Zeocin™
Vector Type:	pLenti

储存

The BLOCK-iT™ Inducible H1 Lentiviral RNAi System includes the BLOCK-iT™ Inducible H1 RNAi Entry Vector Kit, pLenti4/BLOCK-iT™-DEST vector, pLenti4-GW/H1/TO-lamin shRNA control vector, pLenti6/TR vector, One Shot® Stbl3™ Chemically Competent E.Coli, LR Clonase® enzyme mix, ViraPower™ Zeo Lentiviral Support Kit, Blasticidin, and 293FT Cell Line. The BLOCK-iT™ Lentiviral RNAi Zeo Gateway® Vector Kit contains the pLenti4/BLOCK-iT™-DEST vector, pLenti4-GW/H1/TO-lamin shRNA control vector, and One Shot® Stbl3™ Chemically Competent E.Coli. Store the pLenti4/BLOCK-iT™-DEST, pLenti6/TR vector, Blasticidin, ViraPower™ Lentiviral packaging mix, and the control vector at -20°C. Store the One Shot Stbl3™ Competent Cells 293 FT cell, and LR Clonase® enzyme mix at -80°C. Store Lipofectamine™ 2000 at +4°C. All reagents are guaranteed stable for 6 months when properly stored.

描述

Take control with inducible RNAi-in any cell type. The pLenti4/BLOCK-iT™-DEST expression vector provided in the BLOCK-iT™ Inducible H1 Lentiviral RNAi System can be used to efficiently regulate RNAi in virtually any mammalian cell. Using the BLOCK-iT™ Inducible H1 Lentiviral RNAi System, long-term inducible shRNA expression is possible. A novel cloning process places a double-immediately following an H1/TO pol III promoter to generate an H1/TO RNAi cassette in an inducible BLOCK-iT™ H1/TO entry construct. This can be quickly and efficiently recombined from the pENTR™/H1/TO entry vector into the pLenti4/BLOCK-iT™-DEST vector via a standard Gateway® LR recombination reaction. Transfect this construct along with the ViraPower™ Lentiviral packaging mix into 293FT cells with Lipofectamine™ 2000 Reagent to produce viral particles capable of transducing any mammalian cell. If the cell expresses the tetracycline repressor (TR) protein, inducible RNAi is possible.

The pLenti4/BLOCK-iT™-DEST expression vector enables lentiviral delivery and genomic integration of DNA coding for shRNA. Once expressed, the shRNA is processed by cellular machinery and initiates target-specific RNAi. The pLenti4/BLOCK-iT™-DEST vector offers:

• Gateway® Technology for efficient recombination of the RNAi cassette from the BLOCK-iT™ inducible pENTR™/H1/TO vector
• All required components for efficient lentiviral packaging, delivery, and integration of the shRNA
• Zeocin™ selection marker for fast selection of clonal cell lines containing the RNAi cassette

Regulated shRNA Expression Following Lentiviral DeliveryFollowing transduction of the inducible BLOCK-iT™ Lentiviral RNAi vector into mammalian cells, the promoters and shRNA become stably integrated together as an inducible RNAi cassette. In cell lines that express the TR protein, the shRNA will be efficiently regulated and only expressed upon induction with tetracycline*. Following induction, the integrated shRNA is expressed and meditates a cellular RNAi response.

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