iPSC Reprogramming Factors Delivery by Episomal Vectors

Overview Service Features Published Data FAQs Scientific Resources Related Services

As a world-leading service provider in the field of iPSC generation, Creative Biolabs offers various factor delivery systems for iPSC reprogramming. With years of research, we currently present a non-integrative approach for the factor delivery based on the episomal vector.

Introduction of Episomal Vector Systema

As an alternative to integration-defective viruses, a novel iPSC reprogramming strategy based on episomal vectors has been developed. This strategy can benefit from the simple implementation that can be achieved with a standard molecular biology experience, and avoid the labor-intensive and time-consuming viral particles production. A typical example is the OriP/Epstein–Barr nuclear antigen‑1‑based episomal vectors (OriP/EBNA1) which contains two components: Orip and EBNA1. Once transfected into host cells, the EBNA1-encoded protein can be expressed via a viral promoter which enables recognize the Orip sequence and result in plasmid amplification following DNA amplification of the host cells. The removal of episomal vectors from host cells can be achieved by culturing in the absence of drug selection. By co-transfecting several OriP/EBNA1 vectors that express different factor combinations, we have generated iPSCs from human foreskin fibroblasts (HFFs) successfully. It is noteworthy that the SV40LT antigen is used as one of the reprogramming factors because of the ability to inactivate both the retinoblastoma and the p53. Therefore，we suitably sacrifce reprogramming efficiency for better production safety against the potential tumorigenic risk.

Minicircle Vectors

In order to avoid the potential methylatable prokaryotic backbone sequences and also decrease the size of reprogramming episomes, we have also designed the novel minicircle vectors which allow the expression of reprogramming factors as non-replicating, non-integrating episomes. Minicircle vectors are supercoiled DNA molecules without antibiotic resistance gene and bacterial origin of replication. By cloning the polycistronic cassette including OCT4, SOX2, LIN28, and NANOG (OSLN) into the minicircle vector, the reprogramming can be achieved forwards human stem cells in a short time. After iPSC reprogramming, the human iPSCs would be obtained for the further drug discovery and clinical trials.

Advantages Compared with Plasmid

Higher transfection efficiency.
Longer ectopic expression of the transgenes

Fig.1 Non-viral delivery methods by episomal vectors.

Services at Creative Biolabs

With our well-established iPSC reprogramming factor delivery systems, the experienced scientists in Creative Biolabs are devoted to helping you with iPSC reprogramming and generation. We prefer to selecting the best delivery system according to your specific situation. Creative Biolabs also provides other services regarding iPSC technology, please do not hesitate to contact us for more details.

Here's an overview of how our service works.

Key Processes	Descriptions
Cell Collection and Culturing	We accept target cells from our customers and develop suitable conditions for culture.
Cell Transfection	The reprogramming factors are carefully chosen for their ability to induce pluripotency like OCT4, SOX2, KLF4, c-MYC, NANOG, LIN28. Our methodology involves a non-integrative episomal-based system to transfect the cells. This method eliminates the need for viral vectors, reducing risks associated site mutations, immunogenic responses and biological waste. And our technology also presents a higher reprogramming efficiency.
Colony Picking	Once the cells have been transfected, they are cultured and monitored for the emergence of iPSC colonies. After approximately 14-21 days, iPSC colonies begin to appear.
Cell Expansion	We will expand for further characterization and validation. We ensure high-quality standards and validate every iPSC line by checking for pluripotency, trilineage differentiation potential, genomic stability and monitoring iPSC morphology.
Deliverables	Our clients will receive high-quality, episomal vector reprogrammed iPSC lines, along with all the relevant QC data.

Features of Our Services

Our service is a highly advanced and robust toolkit. This innovative and convenient service boasts several vital features, making it an attractive choice for researchers and businesses across the globe.

Non-integrative System - Our episomal vectors are non-integrative, ensuring your iPSCs don’t carry any unwanted, potentially mutagenic foreign DNA.
Complete Service - From the initial isolation and culture of somatic cells, to the final validation of pluripotency, we provide a start-to-finish service.
Safety and Removability - Episomal vectors can be safely removed after the reprogramming process, leaving no residual traces in the iPSCs.
Fast and Qualified - Our expert team provides a rapid turnaround, typically generating validated iPSC lines in less than two months. We maintain high-quality standards, providing full documentation for all reprogramming steps and validation results.
High Efficiency - Our service utilizes an optimized process achieving high iPSC reprogramming efficiency, accelerating your research or product development pipeline.

Through these outstanding features, we provide a complete solution for iPSC reprogramming that we are confident will accelerate your stem cell research. Our service is a robust and reliable choice.

Published Data

Below are the findings presented in the article related to iPSC reprogramming factors delivery by episomal vectors.

M Kim et al. reveals an effective gene delivery method specifically for Lymphoblastoid cell lines (LCLs). They found that LCLs appear to be refractory toward retroviral and lentiviral transduction. Consequently, lentiviral and retroviral transduction of OCT4, SOX2, KFL4 and c-MYC into LCLs does not elicit iPSC colony formation. And they found that transfection of oriP/EBNA-1-based episomal vectors by electroporation is an efficient gene delivery system into LCLs, enabling iPSC generation from LCLs. These iPSCs expressed pluripotency makers (OCT4, NANOG, SSEA4, SALL4) and could form embryoid bodies.

Fig. 2 Electroporat LCLs with episomal vector encoding reprograming factors.³

FAQs

Q: Is there any risk or potential harm in this process to the cells?
A: Our method of reprogramming using episomal vectors is one of the safest as it does not involve any genetic modifications or the use of viral vectors. However, like any biological process, there can potentially be some level of stress on the cells during reprogramming. We manage this through careful monitoring and optimized protocols to ensure the health and viability of the cells.
Q: What types of cells can be reprogrammed using this service?
A: A wide range of human adult cells can be reprogrammed using our service, primarily blood cells and fibroblasts. However, the efficiency of reprogramming might vary among different cell types. Feel free to discuss your specific needs, and we'll advise on the best approach.
Q: Can you ensure that the induced iPSCs will be free from episomal vectors?
A: Yes, as part of our quality control, we perform multiple rounds of cellular passaging to ensure depletion of episomal vectors from the generated iPSCs, ensuring that you receive high purity, episome-free iPSCs.
Q: Can I trust the efficiency of your method?
A: Absolutely. We utilize a robust reprogramming protocol that has been optimized for maximum efficiency. We have many satisfied customers who have successfully utilized our iPSCs for their advanced research studies.

Scientific Resources

iPSC Reprogramming

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References

González, F. (2011). “Methods for making induced pluripotent stem cells: reprogramming a la carte.” Nature Reviews Genetics 12(4), 231.
Hu, W. (2015). “Derivation, expansion, and motor neuron differentiation of human-induced pluripotent stem cells with non-integrating episomal vectors and a defined xenogeneic-free culture system.” Molecular Neurobiology 53(3), 1-12.
Kim, Myunghyun, et al. "Generation of induced pluripotent stem cells from lymphoblastoid cell lines by electroporation of episomal vectors." International Journal of Stem Cells 16.1 (2023): 36.

For Research Use Only. Not For Clinical Use.