mRNA Synthesis

Hanhai New Enzyme possesses the necessary restriction endonucleases for the preparation of linearized plasmid templates, enzyme raw materials for in vitro transcription, modified nucleosides, cap analogues, purification, and quality control materials. It can provide customized synthesis of mRNA with high yield, purity, and cost-effectiveness in different specifications.

Hanhai New Enzyme provides full process services from sequence design to mRNA synthesis, supporting the synthesis of various forms of RNA such as linear mRNA, saRNA, CircRNA, etc. It also has a professional quality control platform, which can better safeguard the research and development of RNA related products for customers.

Technical process
Self-Amplifying RNA

Self amplifying RNA (saRNA) is an mRNA that encodes a replicase and can self replicate within cells using itself as a template. SaRNA contains the basic elements of mRNA (cap, 5'UTR, 3'UTR, and variable length polyadenylation tail), with a large open reading frame (ORF) at the 5 'end encoding four non structural proteins (nsP1-4) and a subgenome promoter to facilitate RNA self replication. The self replication property of saRNA can achieve lower dosages of RNA drugs, prolong protein expression time in vivo, and reduce side effects.

Hanhai New Enzyme can provide customized synthesis services for saRNA, selecting the optimal expression elements, sequence design, and base modification to ensure efficient in vivo and in vitro expression of saRNA, and better assist in the research and development of saRNA products.


Case 1
Case 2
Product name
saRNA1
saRNA2
Hat adding method
Cotranscription
Cotranscription
Modified nucleoside
UTP
UTP
mRNA length
8000nt
9020nt
A260/280
2.1
2.1
Appearance
No impurities/precipitation
No impurities/precipitation
Analysis of mRNA Integrity by mRNA Liquid Capillary Electrophoresis Nucleic Acid Analyzer
The delivery efficiency of saRNA EGFP under low-dose conditions is higher than that of mRNA, showing a significant dose advantage
The expression persistence of self replicating saRNA has a significant advantage in persistence
D1
D2
D3
D4
4.0E+06
3.0E+06
2.0E+06
1.0E+06
0.0E+06
10ng
25ng
50ng
100ng
500ng
1ug
100ng
500ng
1ug
saRNA-Fluc
mRNA-Fluc
mRNA-EGFP (AG,UTP)
saRNA-EGFP (AG,UTP)
Time (d)
Translation level of saRNA Fluc cells
Long term expression detection in mice
CircRNA

Circular RNA (circRNA) is a novel non coding RNA produced by reverse splicing of pre mRNA precursors. Unlike traditional linear RNA, circRNA has a closed circular structure formed by covalent bonds at the 5 'and 3' ends, which is not affected by RNA nucleases, resulting in more stable expression, less degradation, and lower immunogenicity.

Hanhai New Enzyme can provide customized synthesis services for CircRNAs, supporting the synthesis of circular RNAs of different fragment lengths, ensuring efficient expression of CircRNAs in vivo and in vitro, and better assisting in RNA therapies such as Car-T and protein replacement in vivo, as well as the research and development of RNA related products such as gene regulation and drug delivery.

Submit Requirements
Sale confirmation
Sequence design (optional)
Gene synthesis (optional)
Plasmid amplification (optional)
RNA raw material synthesis
Quality testing
Service Process
DELIVERY STANDARDS
Service category
Project
Duration (working days)
Deliver
Sequence design (not undertaken separately)
Sequence design
5
Sequence file
Templates preparation
Plasmid synthesis
20
Plasmid sequencing files, if requested by the customer, provide synthesized raw plasmids (1-2 μ g)
Plasmid amplification
5
Plasmid sequencing files, if requested by the customer, can be provided to the customer if there are remaining plasmids produced, and no additional preparation will be made if there are no remaining plasmids
mRNA raw material synthesis
Synthesis of mRNA Raw Solution - Non nucleotide Modified mRNA (Research Grade)
5
mRNA stock solution and COA
mRNA raw material synthesis - nucleotide modified mRNA (scientific grade)
5
mRNA Raw Solution Synthesis - Non nucleotide Modified mRNA (Industrial Grade)
5
mRNA raw material synthesis - nucleotide modified mRNA (industrial grade)
5
Service category
Project
Duration (working days)
Test method
Can provide quality inspection services (not separately undertaken)
Double-stranded RNA
1
ELISA
Total protein residue
1
NanoOrange
DNA template residue
1
Fluorescence quantitative PCR
Endotoxin
1
TAL
RNase residue
1Fluorescence probe method
DNase residue
1Fluorescence probe method
Host DNA residue
1Fluorescence quantitative PCR
NTP and hat like residue
1
HPLC
Customized synthesis of CircRNA
Buffer gradient
Buffer gradient 1 44.0%
Buffer gradient 2 48.7%
Buffer gradient 3 44.3%
Reaction temperature
41℃ 39.5%
43℃ 50.3%
45℃ 59.0%
47℃ 64.2%
53℃49.9%
RNaseR digestion treatment
Analysis time (minutes)
Analysis time (minutes)
Analysis time (minutes)
After buffer, T7 mutant, reaction system, and reaction temperature optimization, the proportion of ring formation was significantly increased< RNaseR digestion treatment resulted in a circular RNA proportion of over 90%.
Tools for RNA Finished Products

In mRNA drug research, it is often necessary to use ready-made mRNA raw materials as cell or animal experimental controls, or to explore purification processes, screen fillers, optimize mRNA expression levels, and evaluate carrier delivery efficiency.

Hanhai New Enzyme provides a variety of IVT mRNA raw materials in stock, including COVID-19, EGPF encoding, Fluc or mCherry reporter genes, DNA endonuclease gene Cas9, erythropoietin gene EPO precursor, etc. Linear mRNA and saRNA forms can be selected to provide more assistance for mRNA research.

Product features:
Stability
It can be stored for one month at 2-8 ℃ and can be repeatedly frozen and thawed more than 10 times
Excellent performance
High integrity, with a cap rate of over 97%
Product validated through cell expression
High intracellular expression efficiency
Complete categories and stable supply
Spot supply, short lead time
CASE
Can produce high-purity mRNA of different lengths

Case 1
Case2
Case 3
Case 4
Product name
mRNA1
mRNA2
mRNA3
saRNA
Hat adding method
Co transcription method with cap
Co transcription method with cap
Co transcription method with cap
Co transcription method with cap
Modified nucleoside
N1-Me-pUTP
N1-Me-pUTP
N1-Me-pUTP
N1-Me-pUTP
mRNA length
500nt
1000nt
2000nt
4000nt
A260/280
1.85
1.84
1.881.92
Appearance
No impurities/precipitation
No impurities/precipitation
No impurities/precipitationNo impurities/precipitation
Capillary electrophoresis nucleic acid analysis

mRNA integrity

Multiple testing items for mRNA raw material can be provided
Testing items
Case 1
Case 2
Case 3
Case 4
mRNA integrity
90.6%
91.9%
93.2%
91.4%
Residual endotoxins
<10EU/mg
<10EU/mg
<10EU/mg
<10EU/mg
RNase residue
<1.56x10^-9 U/μL
<1.56x10^-9 U/μL
<1.56x10^-9 U/μL
<1.56x10^-9 U/μL
Protein residue
<0.5μg/mg
0.521μg/mg
0.53μg/mg
0.5μg/mg
Residual plasmid template
3.07x10^-4 μg/mg
5.18x10^-6 μg/mg
1.05x10^-4 μg/mg
7.21x10^-5 μg/mg
dSRNA residue
0.0789%
0.1749%
0.0389%
0.3266%
Cap rate
98.71%
100%
98.77%
100%
Poly (A) distribution
Qualified
Qualified
Qualified
Qualified
Sequence correctness
Qualified
Qualified
Qualified
Qualified
Service ordering and consultation
Website Link
Learn more
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Email:hhxm-info@hzymes.com
Large scale production base: Building 6, Wuhan Precision Medical Industry Base, No. 9 Gaokeyuan Third Road, Wuhan City
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