Design, Chemical Modification and Coupling Process Development of siRNA
RNAi refers to the phenomenon of specific inhibition of target genes through the formation of double-stranded RNA by antisense RNA and plus strand RNA. By artificially introducing double-stranded RNA with the same sequence as the endogenous target gene, mRNA degradation of the endogenous target gene is induced to achieve the purpose of blocking gene expression. SiRNA is an important intermediate effector molecule by which RNAi action occurs. SiRNA technology can conveniently suppress specific genes, which can be endogenous or exogenous. Endogenous genes, such as oncogenes and disease-related genes, and exogenous genes, such as viral genes, can be specifically inhibited, which can be used in clinical gene therapy and the development of new drugs.
Creative Biolabs has passed the ISO international quality management system certification and obtained the cGMP oligonucleotide API manufacturing license, which will create the world's leading cGMP oligonucleotide API base.
Our technical service for siRNA synthesis:
- Chemical synthesis. mRNA sequences are inferred from gene databases and gene prediction software of Creative Biolabs to select target sequences as templates. The sense sequence and antisense strand are synthesized by chemical method of 2'O-tri-isopropyl silyloxymethyl and then the two chains are annealed to form a double chain body. This method is a classical method for RNA synthesis. Although it is expensive, the obtained siRNA sequence is the most accurate.
- In vitro transcription. Since G is the first nucleotide required for the initiation of T7RNA polymerase, GN17CN2-like sequence is selected from each target mRNA sequence as the DNA template sequence of siRNA. N2 is the 3' suspended nucleotide of siRNA. UU-3 'can be selected on the sense sequence. This method is simple to use, low cost and stable inhibition. It has been widely used in RNAi experiments, which is most suitable for siRNA design sequence screening. However, it is still a time-consuming method for large-scale experiments.
Advantages of purification methods service:
The synthesis method of RNA is the same as DNA, but the chemical synthesis of RNA is more difficult, due to the low coupling efficiency of RNA base, easy decomposition of products and the need for special protection. Creative Biolabs is equipped with various specifications of automatic synthesis equipment and advanced purification equipment. Creative Biolabs has a wealth of RNA synthesis experience and has improved a set of RNA synthesis and purification technology, providing high quality RNA synthesis products. There are three levels of RNA synthesis products: standard purification, PAGE purification and HPLC purification. You can choose the appropriate level of products according to the experimental needs.
- Standard purification: Standard purification with 80% product purity is suitable for the preparation of short-stranded RNA, such as RNA Linker, short-stranded RNA probe, siRNA, miRNA mimic, miRNA inhibitor, miRNA agomir, miRNA antagomir. PAGE purification should be performed when the RNA strand length exceeded 30mer.
- PAGE purification: PAGE purification with 90% product purity is suitable for the synthesis of various long RNA, such as antisense nucleic acid, long RNA probe, miRNA precusor. Short RNA can also be purified by this method.
- HPLC purification: HPLC purification with more than 95% product purity is extremely high purity, but it is only suitable for the purification of short-stranded RNA.
References
- Rizvi NF, et al. RNA as a small molecule druggable target. Bioorg Med Chem Lett. 2017, 27(23):5083-5088.
- Sztuba-Solinska J, et al. Unveiling the druggable RNA targets and small molecule therapeutics. Bioorg Med Chem. 2019, 27(10):2149-2165.
*For Research Use Only. Not for use in diagnostic procedures.