Shape matters: size-exclusion HPLC for the study of nucleic acid structural polymorphism.
Bottom Line: The knowledge of the structure(s) formed by oligonucleotides is thus critical to correctly interpret the results, and gain insight into the biological role of these particular sequences.Case studies are provided to clearly illustrate the all-terrain capabilities of SE-HPLC for oligonucleotide secondary structure analysis.Finally, this manuscript features a number of important observations contributing to a better understanding of nucleic acid structural polymorphism.
Affiliation: ARNA Laboratory, University of Bordeaux, Bordeaux 33000, France INSERM, U869, IECB, Pessac 33600, France.Show MeSH
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Mentions: Next, duplex-forming oligonucleotides, varying in length (10–26 nt) and base composition, are analyzed (Figure 3A). Auto-complementary sequences were chosen as they can be structured in the homo-duplex and/or the hairpin-duplex form, depending on the annealing conditions (nucleic acid concentration, cation concentration, cooling rate). Additionally to the widely studied ds26 and dslac (lac operon) sequences, four sequences are based on the so-called Drew-Dickerson model (ds10, ds14 and the mirror-image sequences ds12-GC and ds12-CG) (35–37). Finally, in order to obtain a reference peak for hairpin structures, the synthetic construct ds15-h, which is not fully auto-complementary, is also injected (Table 1). Indeed, the three central nucleotides forming the loop in the hairpin mismatch in the case of the homo-duplex, rendering the latter significantly less stable than the former (38).
Affiliation: ARNA Laboratory, University of Bordeaux, Bordeaux 33000, France INSERM, U869, IECB, Pessac 33600, France.