Antibiotic-Free RNA-OUT Marker for Mammalian Expression and DNA Vaccine Vectors

NTC Contacts:
Phone: (402) 323-6289 (natx)
Fax: (402) 323-6292
Email: support@natx.com

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NTC PRODUCTS AND SERVICES

MAMMALIAN EXPRESSION VECTORS
X• Antibiotic-Free Vectors
X• Rig-I Activating Vectors
X• Linear Vectors
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X• Gene Silencing (transient)
X• Gene Silencing (retroviral, stable)
X• Anti-Silencing Elements
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X• Autolytic E. coli Cell Lines
X• dcm- Strains

VACCINES & ADJUVANTS
X• DNA Vaccines
X• Rig-I Activating DNA Vaccines
X• Thermostable Protein Expression

PRECISION CLONING
X• Services and Pricing

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X• pVEX™-E. coli Expression Vectors
X• Fluorescent Proteins
X• Products, Vectors, Services, Prices
X• CofA (SAMPLE)

PLASMIDS
X• Small Scale Plasmid DNA
X• Small Scale C of A
X• Large Scale Plasmid DNA
X• Large Scale C of A
X• Plasmid DNA Order Form
X• E. coli Cell paste
X• NEW! Plasmid Detox Service
X• Additional Testing

PROCESS DEVELOPMENT

DNA FERMENTATION PROCESS
X• HyperGRO™ Publications
X• E. coli Cell Paste

HYPERTAQ™ DNA POLYMERASE

CONTRACT QC
X• E. coli Genomic DNA Testing
X• Residual RNA qPCR
X• Residual RNA gel
X• Supercoil Assay
X• Lymulus Endotoxin Testing

In keeping with regulatory agency guidance recommending removal of antibiotic resistance genes from vectors used in humans, NTC now introduces an antibiotic free selection system (Fig.1) based on sucrose. In it, the KanR gene is replaced with a 150bp RNA-OUT antisense RNA, which represses expression of a counter selectable marker (SacB) gene located in the host chromosome. This innovation has the additional advantages of: shrinking the plasmid backbone, making it more potent, and decreasing the cost of production, while retaining high yield replication in E. coli.

Fig. 1: RNA-OUT selectable marker

 DNA Vaccine Vecotr, Antibiotic freeRNA-OUT selectable marker

NTC offers a retrofitting service, directly replacing the Kan gene in your current vectors with the RNA-OUT cassette.
XXFeatures:


• Replaces KanR resistance gene with antisense RNA-OUT, 150bp RNA marker
• Kit includes strain for selection with RNA-OUT
• Enhanced expression with optimized, chimeric promoter-intron (SV40-CMV-HTLV-1 R-U5-synthetic intron)
• Complies with regulatory guidance
• Increased vaccine potency and safety by replacement of KanR gene
• Improved DNA backbone for maximum plasmid production in E. coli

XXVector Specifications:


NTC8485-EGFP.  RNA-OUT replaces kanamycin resistance gene.

DNA Vaccine Vector, NTC8485-EGFP.  Marker is removed to clone a gene of interest.

Vector Features: trpA prokaryotic terminator: 3739-28; Primosomal assembly site (PAS-BH) extended origin: 32-316; pUC replication origin: 317-1331; Sucrose selection marker (RNA-OUT): 1348-1492; SV40 enhancer: 1493-1719; CMV enhancer: 1726-2283; CMV promoter: 2284-2403; Untranslated leader (exon 1): 2404-2590; HTLV-1 R-U5: 2475-2700; Synthetic Rabbit β-globin-based 3’ intron: 2709-2815; Intron: 2591-2815; Exon 2 (SR-protein binding sites-Kozak):2816-2864; EGFP: 2865-3584; SalI-BglII cloning cassette: 2859-3594; Eukaryotic terminator: 3595-3729.