RNA Lasso® Probes
Current detection methods that require nucleic acid hybridization have a trade-off between sensitivity and specificity:
Specific detection of HCV genotypes with SomaGenics' Lasso probes. Note absence of cross-reaction.
- Long probes (≥60 nucleotides) provide good efficacy of hybridization but have poor sequence specificity.
- Short probes (<25 nucleotides) provide higher sequence-specificity but lower efficacy of hybridization than long probes.
The compromise between efficacy and specificity is in large part responsible for the significant signal variability that plagues the use of microarrays and other hybridization-based assays. While the diagnostics industry has focused on improvements in probe immobilization and data analysis, there is a need for improvements in probe design that addresses the specificity-sensitivity tradeoff. SomaGenics developed its RNA Lasso® detection platform to fulfill this need.
RNA Lasso probes, which consist of a
short target-complementary sequence flanked by two modified hairpin
ribozyme domains, have a unique combination of high sequence
specificity and strenth of binding.
The hairpin ribozyme domains, which have a fixed sequence, serve several purposes: they self-cleave when embedded in longer RNA, allowing the Lasso to be produced by in vitro transcription; they allow circularization of the RNA molecule; and their secondary structure improves the performance of the Lasso probe.
Advantages of Lasso RNA probes
Lassos are an ideal combination of efficiency, sensitivity and specificity
- Match or exceed the hybridization kinetics of long polynucleotide probes
- Match or exceed the sequence-specificity of short oligonucleotide probes
- Match or exceed the sensitivity (signal-to-noise ratio) of ordinary probes
- Form strong complexes with long RNA targets that are more stable than ordinary RNA-RNA and RNA-DNA duplexes
- Stable under PAGE with 8M urea
- Can efficiently hybridize with structured ssRNA and dsDNA targets
- Circular structure enhances the sequence-specificity and efficiency of hybridization
- Allow efficient DNA and RNA target capture and enrichment, for quantification by RT-qPCR and resequencing
- Dallas, A., Balatskaya, S.V., Kuo, T-C., Vlassov, A.V., Kaspar, R.L., Kisich, K., Kazakov, S.A., Johnston, B.H. (2008) Hairpin ribozyme-antisense RNA derivatives act as molecular Lassos. Nucleic Acids Res. 36(21):6752-66; Epub Oct 25, 2008. PMID 18953032
- S. A. Kazakov, S. V. Balatskaya, and B. H. Johnston (2006) Ligation of the hairpin ribozyme in cis induced by freezing and dehydration. RNA 12: 446-456. PMID 16495237
- S. A Kazakov and B. H. Johnston (2008) Development of gene profile-responsive antisense agents. In Gene Profiles in Drug Design, B. A Lidbury and S. Mahalingam, eds., CRC Press, Boca Rotan, FL.