On the journey of molecular exploration, one of the key considerations is the stability of genetic building blocks, oligonucleotides. These short sequences of nucleotides play a major role in genetic research, diagnostics, and therapeutic developments.
However, oligonucleotides require careful handling and storage to maintain their integrity and functionality like any valuable tool. This blog will discuss how to store oligonucleotides for optimal stability and ensure your oligos remain at their best when you need them the most.
Temperature and Medium
Storage temperature and the chosen medium significantly influence the real-time stability of oligonucleotides. Among these factors, temperature takes the most important place. Extensive studies reveal that oligos stored at -20°C, whether in dry form or resuspended in TE buffer or nuclease-free water, maintain stability for up to two years (24 months)
At a refrigerated temperature of 4°C, oligonucleotides exhibit comparable stability across different storage mediums-dry, TE buffer, or nuclease-free, remaining stable beyond one year (>60 weeks). Therefore, the recommended best practice is freezing oligonucleotides at -20 °C in TE buffer, nuclease-free water, or dried form for storage periods of up to 24 months, with minimized exposure to UV light.
Medium at Elevated Temperatures
When evaluating stability under stress conditions such as 37°C, the choice of storage medium becomes essential. Oligonucleotides stored in nuclease-free water demonstrate the least stability, followed by dried forms. In contrast, those preserved in TE buffer (IDTE, pH 8.0) exhibit superior stability under elevated temperatures.
Even when oligonucleotides are dried, a minute amount of moisture can potentially lead to DNA damage over time. Hence, freezing in TE buffer, nuclease-free water, or dry form is optimal for long-term storage.
TE Buffer for Maximum Stability
Suppose we discuss DNA oligo stability across different storage mediums- dried, nuclease-free water, and TE buffer (pH 7.5/8.0). Oligonucleotides stored in TE buffer have the highest stability over extended periods, especially at higher storage temperatures. Tris (tris(hydroxymethyl)aminomethane) maintains a stable pH, while EDTA (ethylenediaminetetraacetic acid) acts as a chelating agent, preventing DNA degradation by nucleases.
Resuspending and storing DNA oligonucleotides in TE buffer rather than nuclease-free water is recommended for enhanced stability. IDT provides IDTE (1X TE Solution) in various volumes and pH options to facilitate this process.
Shipping Conditions
Lyophilized (dried) oligos subjected to 37°C for up to 25 weeks maintain a minimal loss of activity (ΔCq <1.5), ensuring functionality. This inherent stability extends even to oligos stored in water, the least stable medium, demonstrating functionality for multiple weeks at elevated temperatures.
Shipping oligonucleotides in a dry state offers flexibility for researchers to choose their preferred storage medium without compromising stability.
Freeze-Thaw
In many cases, repeated freeze-thaw cycles are required during experimental procedures. Extensive testing reveals that oligonucleotide stability remains unaffected after 30 cycles, regardless of the resuspension medium- nuclease-free water or IDTE buffer. This strength is true for both standard and modified oligonucleotides.
While oligos maintain stability through multiple cycles, environmental exposure could introduce nucleases. To prevent contamination, it is advisable to aliquot oligos before storage.
Modified Oligos
Oligonucleotides with modifications exhibit similar stability characteristics to unmodified counterparts. Thus, they should adhere to the same storage guidelines. Freezing emerges as the optimal method for prolonged storage, with IDTE buffer aiding stability at temperatures above freezing. Short-term exposure to UV light or ambient lab conditions does not significantly impact the functionality of fluorophore-modified oligos. However, extended storage in darkness is recommended to preserve fluorophore stability.
Take Care of RNA Stability
The inherent instability of RNA, owing to its chemical structure, presents unique challenges. RNA oligonucleotides are particularly susceptible to degradation by prevalent RNases in laboratory environments. Optimal storage involves the use of RNAse-free water and buffers. IDTE (1X TE Solution) is recommended for short-term storage, while long-term preservation mandates storage as an ethanol precipitate at -80°C.
Following these guidelines can help protect oligonucleotide stability and the functionality of these molecular tools.
Read our blog about Oligonucleotide Synthesis Process