Custom circRNA Synthesis

IVT RNA

Circular mRNA (circRNA) has been widely used in the study of gene or protein functions. Meanwhile, it is popular in the development of prophylactic and therapeutic biologics, similar to mRNA.

Yaohai Bio-Pharma has established a set of circRNA synthesis technologies to provide high-quality RNA, such as sequence design and optimization, in vitro transcription (IVT), circularization/cyclization, purification, and lipid nanoparticle (LNP) encapsulation. All products are released under the stringent quality control (QC) standards.

CircRNA Applications

Compared with linear mRNA, circular RNA (circRNA) is more stable in structure and is a major hot spot in current nucleic acid drug research. It has a wide range of applications in different fields, whether as coding circRNA or non-coding circRNA, including prophylactic vaccines for infectious diseases, therapeutic oncology vaccines, therapeutic oncology drugs, protein replacement therapy, regenerative medicine, cell and gene therapy (CGT), molecular sponge role and RNA aptamer.

Service Details

Process	Optional Service	Service Details	Delivery Period (day)
circRNA Sequence Design and Optimization	Design and Optimization of Coding Sequences	CDS Codon Optimization	1
circRNA Sequence Design and Optimization	Design and Optimization of Non-Coding Sequences	Intron and Exon Sequence, Homologous Arm and Spacer Design and Optimization	1-2
Circular Plasmid Preparation	Gene Synthesis	Gene Synthesis (Third-party)	7-10
	Plasmid Amplification	Plasmid Amplification	2
	Plasmid Amplification	Plasmid Extraction	2
Linearized Plasmid Preparation	Plasmid Linearization and Purification	Plasmid Linearization	1
Linearized Plasmid Preparation	Plasmid Linearization and Purification	Linearization DNA Purification	1
In Vitro Transcription and Cyclization	In Vitro Transcription and Cyclization	In Vitro Transcription and Cyclization Reactions	1-2
	In Vitro Transcription and Cyclization	RNase R Digestion	1-2
	Optimization-optional	Reaction Composition, Time Optimization	2-5
circRNA Purification	Conventional Purification Method	Lithium Chloride Precipitation	1
	Conventional Purification Method	Magnetic Bead Purification	1
	Highly Pure circRNA Purification	Preparative High Performance Liquid Chromatography (HPLC)	1-2
	Buffer Exchange	Ultrafiltration	1
circRNA Lyophilization	Sample Filling	Filling	2-3
	Lyophilization	Pre-freezing
		Primary Drying (Sublimation)
		Secondary Drying (Desorption)
circRNA-LNP Encapsulation	LNP Encapsulation	Material and Liquid Pretreatment	2
	LNP Encapsulation	Microfluidic Device Mixing	2
	Buffer Exchange	Tangential Flow Filtration	1
	Sterilize	Filtration	1
Quality Control of Plasmid DNA	Concentration/purity	Ultraviolet (UV) Spectrophotometry	1-2
	Plasmid conformation	Agarose Gel Electrophoresis (AGE)
	Plasmid conformation	Capillary Electrophoresis (CE)-Optional
	Plasmid identify	Restriction Enzyme Identification/AGE
Quality Control of circRNA	Concentration/purity	Ultraviolet (UV) Spectrophotometry	-
	Purity	Agarose Gel Electrophoresis (AGE)/E-Gel	0.5
	Purity	HPLC-optional	1
Quality Control of circRNA-LNP	Encapsulation Efficiency	RiboGreen Method	1
	Particle Size	Dynamic Light Scattering (DLS)	1
	Polydispersity Index	Dynamic Light Scattering (DLS)	1
	Zeta Potential	Dynamic Light Scattering (DLS)	1
Cell-based Potency Assay	Cell Transfection	Cell plating, cell transfection	4
Cell-based Potency Assay	Target Protein Detecting	Fluorescence observation, Western Blot/ELISA	1-3

Case Study

We transfected enhanced green fluorescent protein circRNA (eGFP circRNA), mCherry circRNA, luciferase circRNA into 293T cells, and detected target protein through fluorescence, luminescent or color signal, western blot (WB), enzyme-linked immunosorbent assay (ELISA).

Cell-based Potency Assays of circRNA Samples by Yaohai Bio-Pharma

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Cell-based Potency Assays