Extracted DNA is inherently susceptible to degradation once removed from its protective cellular environment. Effective preservation of this genetic material is important for its continued utility in various fields, including scientific research, medical diagnostics, and forensic analysis.
Factors Affecting DNA Stability
Several environmental and biological elements can compromise the stability of extracted DNA. High temperatures accelerate the chemical reactions that lead to DNA degradation. Exposure to light, particularly ultraviolet (UV) radiation, can cause direct damage by forming abnormal bonds between DNA bases, which disrupt the DNA structure. UV light can also indirectly damage DNA by generating reactive oxygen species.
Enzymes known as nucleases are biological contaminants that actively break down DNA. These enzymes can be present during extraction or introduced through improper handling. Extreme pH conditions, both highly acidic and highly alkaline, also destabilize DNA molecules. Acidic conditions can lead to the loss of purine bases, while alkaline conditions can cause denaturation by breaking hydrogen bonds between DNA strands. Physical stresses, such as repeated cycles of freezing and thawing, can also physically damage DNA, leading to fragmentation.
Standard Storage Methods
Common laboratory methods for storing extracted DNA offer solutions for short to medium-term preservation. Refrigeration at temperatures between 2-8°C is suitable for DNA storage over days to weeks. For this method, DNA is typically dissolved in a buffer like Tris-EDTA (TE) buffer, which helps maintain a stable pH.
For medium-term storage, extending from months to several years, freezing at -20°C or -80°C is widely used. Lower temperatures significantly slow down the chemical and enzymatic processes that cause DNA degradation. DNA is often stored in nuclease-free water or TE buffer when frozen. Another practical approach involves storing DNA in a dried state, either as a dry pellet or on specialized paper matrices like FTA cards. Drying the DNA eliminates the need for continuous refrigeration and reduces nuclease activity, making it convenient for transport or storage at room temperature for slightly extended periods.
Advanced Long-Term Preservation
For ultra-long-term or archival storage, particularly in biobanking settings, more specialized techniques are employed. Cryopreservation in liquid nitrogen at -196°C is the most effective method for indefinite DNA storage, as it effectively halts all biological and most chemical activity. To prevent ice crystal formation during freezing, which can damage DNA, cryoprotectants are often added to the samples before plunging them into liquid nitrogen.
Advanced room-temperature dry storage methods involve stabilizing DNA on specialized matrices or in unique encapsulation systems. These techniques allow for very long-term preservation without the energy demands of refrigeration or freezing, offering a practical solution for large biobanks. Some emerging technologies involve encapsulating DNA within protective polymers or silica matrices, which can shield it from environmental factors like heat and water.
Maintaining Stored DNA Integrity
Preserving the quality of stored DNA extends beyond the initial storage choice, requiring ongoing attention to handling and verification. Clear, durable labeling of each sample is important, along with detailed inventory records noting concentration, extraction date, and source. Accurate record-keeping ensures proper identification and traceability of samples over time. Aliquoting, or dividing the DNA sample into smaller portions, is a practice that minimizes the number of freeze-thaw cycles a single sample undergoes. This reduces physical stress and potential degradation each time a portion is needed for an experiment.
To prevent contamination by nucleases or other degrading agents, strictly adhering to sterile techniques and using nuclease-free reagents, such as nuclease-free water, is important during all handling steps. Regular quality control checks are also important to assess DNA quality before and during storage. Techniques such as spectrophotometry can determine DNA concentration and purity, while gel electrophoresis provides visual confirmation of DNA integrity and can detect fragmentation or RNA contamination. When retrieving samples, gentle thawing at room temperature or on ice is recommended to minimize damage that can occur with rapid temperature changes.