mRNA Co-transcription and Synthesis Kit, BP-07
Co-transcriptional kit for Cap1 mRNA synthesis.
Product overview
Co-transcriptional capping is a method for introducing cap structures during in vitro transcription, primarily used to enhance mRNA stability and translation efficiency. The mRNA Co-transcription and Synthesis Kit utilizes T7 RNA polymerase to transcribe DNA sequences downstream of a T7 promoter, using linear double-stranded DNA containing the T7 promoter sequence and an AG initiation sequence as the template, NTPs as substrates, and cap analogs. This process efficiently synthesizes single-stranded RNA with a 5'-m7G Cap1 structure without requiring additional enzymatic capping steps. It can also be used to prepare biotin- or dye-labeled RNA using modified nucleotides as substrates. This kit synthesizes both long and short transcripts via co-transcriptional methods, providing nucleotides including ATP, CTP, GTP, UTP, and the modified N1-Me-pUTP, as well as the cap analog Cap GAG. An input of 1 μg of template DNA can yield 150-200 μg of Cap1 RNA. The RNA synthesized by transcription using this kit is suitable for various downstream applications, such as RNA structure and function studies, RNase protection assays, probe hybridization, RNAi, microinjection, and in vitro translation.
Features and applications
- Co-transcriptional capping enables mRNA transcription and capping in a single step;
- A 1 μg template input can yield 150–200 μg of Cap1 RNA;
- The kit provides Cap GAG, achieving a capping efficiency of ≥90% (up to >95%);
- The kit also provides N1-Me-pUTP, which can further reduce immunogenicity.
- The synthesized mRNA can be used directly for in vitro translation and cell transfection experiments.
- Downstream applications such as RNA structure and function studies, RNase protection assays, probe hybridization, RNAi, microinjection, and in vitro translation.
- Preparation of biotin- or dye-labeled RNA using modified nucleotides as substrates.
Components
| Components | Cat. No. | Volume |
|---|---|---|
| ATP Solution(100 mM) | BP-AS-01 | 100 μL |
| GTP Solution(100 mM) | BP-AS-02 | 100 μL |
| CTP Solution(100 mM) | BP-AS-03 | 100 μL |
| UTP Solution(100 mM) | BP-AS-04 | 100 μL |
| N1-Me-pUTP Solution(100 mM) | BP-AS-07 | 100 μL |
| T7 RNA Polymerase | BP-E01 | 100 μL |
| RNase Inhibitor(40 U/μL) | BP-E02 | 100 μL |
| Inorganic Pyrophosphatase(0.1 U/μL) | BP-E03 | 100 μL |
| DNase I (2 U/μL) | BP-E04 | 100 μL |
| Cap GAG(100 mM) | BP-AS-38 | 100 μL |
| 10×Co-transcription Buffer | BP-AS-31 | 100 μL |
| Precipitation Reagent | BP-AS-10 | 1 mL |
| RNase-free ddH2O | BP-AS-11 | 1 mL |
Performance data
1 mRNA Co-transcription: Template Compatibility and Substrate Flexibility
Description: To evaluate the transcription performance of the Biori mRNA Co-transcription and Synthesis Kit using four DNA templates of varying sizes. Transcription reactions were performed with each template under both natural (UTP) and modified (N1-Me-pUTP) nucleotide conditions. The resulting RNA products were analyzed for yield and integrity.
Conclusion: The Biori mRNA Co-transcription and Synthesis Kit efficiently transcribed all four DNA templates, yielding RNA products with high integrity and robust yields under both natural (UTP) and modified (N1-Me-pUTP) nucleotide conditions, demonstrating its broad template compatibility and substrate flexibility.
2 Capping efficiency analysis of Cap1-mRNA prepared with the mRNA Co-transcription and Synthesis Kit
Description: Cap1-mRNA synthesized using the Biori mRNA Co-transcription and Synthesis Kit was processed with the Pre-treatment Kit for mRNA Capping Rate Detection (Cat: BP-05-10) and subsequently analyzed by LC-MS to determine the capping efficiency.
Conclusion: The Cap1-mRNA prepared using the Biori mRNA Co-transcription and Synthesis Kit achieved a capping efficiency of up to 97.3%, as determined by LC-MS analysis, demonstrating the kit's capability to produce highly capped mRNA transcripts.
3 eGFP expression of Cap1-mRNA in 293T cells
Description: Cap1-mRNA prepared using the Biori mRNA Co-transcription and Synthesis Kit was transfected into 293T cells to evaluate its translational expression efficiency. eGFP fluorescence was monitored to assess the expression level.
Conclusion: The Cap1-mRNA prepared using the Biori mRNA Co-transcription and Synthesis Kit was efficiently expressed in 293T cells, as demonstrated by robust eGFP fluorescence, confirming the kit's capability to produce translationally competent mRNA.
Shipping and storage
| Condition | Recommendation |
|---|---|
| Storage | Store at -20±5°C. |
| Shipping | Ship frozen on dry ice or with validated frozen cold-chain packaging that maintains the specified frozen storage condition. |
| Handling | Avoid repeated freeze-thaw cycles; mix thoroughly after thawing where applicable. |
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