TcBuster™-M Transposase mRNA
R&D Systems | Catalog # TCB-001.1-100
Key Product Details
Features
Key Benefits
- Delivers large, multicistronic DNA cargo
- Efficient and stable gene delivery for a wide range of genome engineering applications
- Integrates cargo with reduced preference for transcript regions compared to lentivirus
- Combines with other gene editing reagents, such as CRISPR, in one electroporation step
- Reduces time and costs compared to traditional viral vector-based technologies
Product Summary for TcBuster™-M Transposase mRNA
Gene delivery is a crucial step for a wide variety of genome engineering workflows, which have traditionally utilized viral vectors to achieve expression of exogenous DNA sequences. However, the generation of a virus is time consuming, inconsistent, and has limited cargo capacity. TcBuster-M Transposase mRNA, in combination with a compatible DNA TcBuster transposon (plasmid), form the TcBuster non-viral gene delivery system. The TcBuster non-viral gene delivery system is a solution for stable integration of DNA cargo and addresses many of the limitations of viral vectors. This product requires a TcBuster DNA transposon to be functional. For control transposons, checkout the CD19CAR-DHFR-eGFP Control for cell therapy applications or Insert On eGFP Control for immortalized cell applications. Please contact us for more information on custom transposons at techsupport@bio-techne.com.
Gene delivery is a critical step in engineering cells for a wide variety of applications. For T and NK immunotherapies, delivery of CAR or TCR DNA cargo is crucial for the potency of the end drug product. Genome engineering involving iPSCs can require stable delivery of complex DNA cargo. For biologics, this system can faciliate stable gene transfer and high titers for work in CHO and HEK cell lines. Because of the diverse applications of the TcBuster system, independent experimental optimization and/or process development will be required. Please visit the TcBuster Non-Viral Gene Engineering page for more information on the system.
Scientific Data Images for TcBuster™-M Transposase mRNA
Mechanistic Diagram of the TcBuster Non-Viral Gene Editing System.
The TcBuster system components are introduced into cells via electroporation (1). The TcBuster-M transposase mRNA is then translated into the transposase enzyme (2), which binds to the inverted terminal repeats (ITRs) on the DNA transposon (3). The transposase enzyme excises the genes of interest (GOI) from the transposon (4) and inserts them into the host genome (5). The GOI mRNA is transcribed from the host genome (6), and the protein is stably expressed in the edited cells (7).
Efficient gene editing of T cells modified with different sized cargos on different electroporation platforms.
Primary T cells from 3 donors were expanded for 7 days after genetic modification in GMP Human T Cell Media (Catalog # CCM038-GMP-1L) supplemented with 5% hAB serum and 10 ng/mL each of IL-7 (Catalog # BT-007-GMP) and IL-15 (Catalog # BT-015-GMP) in a 6 well G-Rex®. (A). Representative flow plots of genetically modified T cells with a 5.1 kb insert containing a GFP sequence, introduced on the ThermoFisher Neon™, Lonza 4D- Nucleofector®, or MaxCyte®. (B).Three DNA cargos introduced on the same electroporation platforms achieve high editing efficiency in T cells (>40%). Data points represent the average of 3 donors with technical duplicates ± SD.
TcBuster edited CD19-CAR-T cells specifically kill CD19+ target cells at low E:T ratios.
T cells from 3 donors were expanded for 7 days after genetic modification in GMP Human T Cell Media (Catalog # CCM038-GMP-1L) supplemented with 5% hAB serum and 10 ng/mL each of IL-7 (Catalog # BT-007-GMP) and IL-15 (Catalog # BT-015-GMP) in a 6 well G-Rex®. (A) TcBuster efficiently integrates cargos of different sizes regardless of electroporation platform, resulting in a similar number of total modified T cells produced. (B) T cells from 3 donors modified with the 5.1 kb plasmid were cryopreserved in CS10 following 7 days of culture after electroporation. T cells were thawed and immediately added to either CD19+ or CD19- target cells at different E:T ratios and controlled target cell growth after 24 hours of co-culture.
Genetic characterization of TcBuster genetically modified primary T cells shows a favorable insertional profile and low copy number.
(A) T cells from 3 donors were collected after 9 days of culture for digital PCR analysis of the population’s average adjusted vector copy number (VCN). (B) Integration site analysis (ISA) was performed on 3 different T cell donors that were modified with a different range of plasmid sizes with TcBuster or a lentiviral vector. DNA libraries were Illumina sequenced by GeneWerk (now ProtaGene).
TcBuster edits primary T cells as efficiently or better than lentivirus.
T cells from 3 donors were expanded for 9 days in GMP Human T Cell Media, supplemented with 5% hAB serum and 10 ng/mL each of IL-7 and IL-15, in a 6 well G-Rex®. The GOI is refers to the promoter and genes integrated in the genome. Graph shows the efficiency of genetic modification for both TcBuster and lentivirus, with TcBuster showing comparable or better editing efficiency. Data points represent the average of 3 donors ± SD.Formulation, Preparation, and Storage
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Product Documents for TcBuster™-M Transposase mRNA
Product Specific Notices for TcBuster™-M Transposase mRNA
FOR LABORATORY RESEARCH USE ONLY. NOT FOR USE IN DIAGNOSTIC PROCEDURES.
- The safety and efficacy of this product in diagnostic or other clinical uses has not been established.
- For certain applications, results may vary due to differences in donor derived cell populations.
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