Custom miRNA Sponges Synthesis

MicroRNAs (miRNAs) play crucial roles in regulating various biological processes, from development to disease. Understanding their functions and mechanisms of action is essential for unraveling complex gene regulatory networks. One powerful tool for studying miRNA function is the use of miRNA sponges, synthetic RNA molecules designed to sequester specific miRNAs and inhibit their activity.

What are miRNA Sponges?

miRNA sponges, also known as competitive endogenous RNA (ceRNA) molecules, are synthetic transcripts containing multiple binding sites for a specific miRNA. These sponges function as competitive inhibitors by sequestering miRNAs and preventing them from binding to their target mRNAs, thus derepressing the expression of miRNA-regulated genes. By titrating specific miRNAs, miRNA sponges enable researchers to investigate the biological functions of miRNAs and their roles in various physiological and pathological processes.

(A) In the absence of sponge treatment, the target mRNA (grey) of a specific miRNA seed family (red complex) is inhibited; (B) Upon introduction of sponge transgene, sponge mRNA (green) is expressed at high levels, sequestering miRNA complexes and rescuing the expression of endogenous targets.

miRNA sponges contain multiple high-affinity binding sites for miRNA. When miRNA sponges are highly expressed in cells, they can inhibit the activity of miRNA families that share the same seed sequence (2-7 nucleotides in the miRNA that determine the specificity of target gene recognition). miRNA sponges can be highly expressed in target cells through transient transfection. Transfection methods may include lipofection or electroporation, and different promoters can be selected, such as CMV, U6 promoters, and tissue-specific promoters. Different types of vectors, such as inducible and constitutive expression vectors, can also be chosen, along with different reporter genes to monitor different cellular phenotypes, such as GFP, luciferase, etc. To enhance infection efficiency, methods like lentivirus, adenovirus, or adeno-associated virus can be utilized.

BOC Sciences' Custom miRNA Sponges Service

BOC Sciences is proud to offer a custom miRNA sponge synthesis service, providing researchers with tailor-made miRNA sponge constructs to facilitate their miRNA studies. Our expert team of scientists utilizes state-of-the-art techniques and cutting-edge technology to design and synthesize high-quality miRNA sponge vectors tailored to the specific needs of our clients. There are various ways regarding the sequence design of miRNA sponges, including bulged sponges, direct antisense design, TuD (tough decoy) RNA design, etc. BOC can choose the most appropriate cloning strategy to construct MBS-containing sponges according to the needs, which can be used for the inhibition of one, two, or more than two miRNAs, respectively.

Types of miRNA Sponge from BOC Sciences

• Target Mimics: Designed to mimic the targets of specific miRNAs, thereby sequestering and inhibiting their activity.
• Decoys: Act as decoy targets for miRNAs, diverting them from their endogenous targets.
• miRNA Target (miRT) Sequences: Contain sequences complementary to specific miRNAs, effectively titrating them and preventing them from exerting their regulatory functions.
• miRNA Erasers: Designed to erase the effects of specific miRNAs by sequestering them and preventing their interaction with target mRNAs.
• Lentivirus-mediated Antagomirs: Utilize lentiviral vectors to deliver antagomirs, synthetic RNA molecules designed to inhibit specific miRNAs.

Notes for BOC Sciences' Custom miRNA Sponge Service

• Binding Sites: Typically contain four to ten binding sites for the target miRNA, separated by short nucleotide sequences.
• Site Structure: Binding sites may be perfectly complementary or contain bulged mismatches, with the latter being more effective due to increased stability of interaction with the miRNA.
• Spacer Sequences: Introduce variations in spacer sequences to reduce the risk of recombination during cloning and unintended binding motifs for other regulatory factors.
• Stabilizing Elements: Terminal stem-loops may be included in PolIII-generated sponge RNAs to enhance stability.
• Promoter Choice: Utilize strong promoters, such as the CMV promoter, to maximize sponge expression.
• Delivery Methods: Employ plasmid transfection or viral delivery for transient or stable expression, respectively, with consideration for cell-type specificity and potential disruption of endogenous genes.
• Reporter Inclusion: Incorporate protein-coding genes upstream of miRNA binding sites for quantitative analysis and selection of high-expressing clones.

Applications of miRNA Sponges from BOC Sciences

• Cell Culture Models: Initial use of miRNA sponges involved transient treatment and assays in various cell culture models, demonstrating versatility in terms of cell type, promoter, vector, reporter gene, and targeted miRNA.
• Chromosomal Integrations: Achieved stable sponge activity by expressing the transgene from chromosomal integrations using various viral vectors, including lentiviruses and retroviruses.
• Viral Delivery: Utilization of viral vectors, such as adenovirus, lentivirus, and adeno-associated virus (AAV), enables delivery of sponge constructs to tissues in live animals for in vivo studies.
• Disease Models: Application of sponge constructs in disease models, such as cardiac hypertrophy, autoimmune encephalomyelitis, and cancer metastasis, highlights their therapeutic potential and ability to modulate disease progression.
• Model Organisms: Successful expression of miRNA sponges in model organisms like Drosophila and plants provides valuable insights into miRNA function and regulation in vivo, offering a platform for studying complex biological processes and disease phenotypes.
• Functional Studies: Stable sponge expression facilitates probing of miRNA roles in differentiation pathways, disease progression, and metastasis, providing insights into miRNA-mediated regulatory networks and potential therapeutic targets.

Advantages of miRNA Sponges from BOC Sciences

• Expertise: With over 20 years of experience in the industry, BOC Sciences boasts a team of highly skilled scientists with expertise in molecular biology, RNA synthesis, and bioinformatics. Our scientists work closely with clients to design customized miRNA sponge constructs optimized for their research goals.
• Quality: At BOC Sciences, quality is our top priority. We adhere to strict quality control measures throughout the synthesis process to ensure the purity, stability, and functionality of our miRNA sponge constructs. Each synthesized miRNA sponge undergoes rigorous validation assays to confirm its efficacy and specificity.
• Flexibility: We understand that each research project is unique, which is why we offer flexible synthesis options to accommodate diverse experimental requirements. Whether you need a single miRNA sponge construct or a library of constructs targeting multiple miRNAs, BOC Sciences can tailor our services to meet your specific needs.
• Fast Turnaround Time: Time is of the essence in research, which is why we strive to deliver fast and efficient service without compromising on quality. Our streamlined synthesis process allows for quick turnaround times, ensuring that researchers receive their custom miRNA sponge constructs promptly.

If you have custom synthesis needs for more types of miRNA Sponges, please feel free to contact us.

Reference

1. M, S, Ebert and P, A, Sharp. MicroRNA Sponges: Progress and Possibilities. RNA. 2010, 16(11): 2043–2050.