1AGM College of Pharmacy Varur, Hubballi.
2Hi-Tech College of Pharmacy, Chandrapur.
3,4D. B. Patil College of Pharmacy, Parola.
5College of Pharmacy (poly), Sawarde.
6Regional forensic Laboratory, Pune.
Buccal epithelial cell sampling has become a preferred method for human DNA collection due to its simplicity, non-invasiveness, and minimal biohazard risk. However, delays between collection and processing—common in forensic investigations and population-based genetic research—can threaten DNA integrity. This review synthesizes current evidence on the stability of buccal swab DNA during two-week storage periods under various environmental conditions. DNA degradation mechanisms—including enzymatic, hydrolytic, oxidative, and microbial processes—are examined, alongside empirical findings from forensic and biomedical studies. Results consistently demonstrate that refrigerated (? 4 °C) or dry-stored swabs retain stable DNA yield, purity, and amplifiability for up to 14 days. In contrast, warm and humid storage accelerates degradation, resulting in reduced quantity and partial amplification failure. The review discusses implications for forensic chain-of-custody reliability and genetic research logistics, providing best-practice recommendations for collection, storage, and transport. Overall, buccal swab DNA demonstrates robust short-term stability when handled properly, supporting its continued use in forensic and genetic applications worldwide.
Buccal epithelial cell sampling has emerged as one of the most practical and ethically acceptable methods for human DNA collection. Its widespread use spans forensic casework, paternity testing, ancestry determination, and large-scale genomic and epidemiological studies. The method’s appeal lies in its non-invasive nature, minimal biohazard risk, and ease of collection compared with venipuncture or tissue biopsy. Modern PCR and sequencing technologies have further enhanced its feasibility by requiring minimal DNA input.
Despite these advantages, buccal DNA is sensitive to post-collection environmental factors such as temperature, humidity, and microbial contamination. In real-world scenarios—crime scene investigations, disaster victim identification, or remote field surveys—sample transport and processing may be delayed for several days or weeks. Such delays raise concerns about DNA degradation and its potential impact on analytical reliability.
Understanding how storage duration and environmental conditions affect buccal DNA quality is therefore crucial. This review synthesizes available research examining DNA integrity over storage periods of up to two weeks, explores degradation mechanisms, and evaluates implications for both forensic and genetic applications.
2. Mechanisms Affecting Buccal Swab DNA Integrity
2.1 Overview
DNA degradation in stored buccal swabs results primarily from enzymatic activity, microbial growth, and environmental stress. Once epithelial cells lyse, endogenous nucleases and external conditions jointly contribute to fragmentation and contamination.
2.2 Endonuclease activity
Endonucleases released from lysed cells rapidly cleave DNA phosphodiester bonds. Their activity increases with temperature and moisture but is inhibited by cold or desiccated storage. Refrigeration (≤ 4 °C) or inclusion of chelating agents (e.g., EDTA) effectively suppresses enzymatic degradation.
2.3 Hydrolytic cleavage
Hydrolysis occurs when moisture facilitates spontaneous cleavage of phosphodiester bonds, leading to depurination and strand breaks. The process accelerates under humid and warm conditions, particularly at ≥ 25 °C. This degradation primarily affects longer DNA fragments, reducing efficiency of PCR amplification and sequencing.
2.4 Oxidative damage
Reactive oxygen species (ROS) generated by exposure to UV light or oxygen cause oxidative base modifications such as 8-oxoguanine. Although slower than enzymatic degradation, oxidative damage accumulates over time and can produce sequencing errors or inhibit amplification, especially in mitochondrial assays.
2.5 Microbial contamination
The oral microbiome contributes exogenous DNA and nucleases that degrade host DNA when swabs remain moist. Environmental microbes may further colonize damp samples. Drying and sealed storage suppress microbial proliferation, preserving DNA integrity.
2.6 Temperature and humidity influences
Temperature and humidity exert the strongest control over degradation kinetics. DNA remains stable up to 14 days at ≤ 4 °C, while at room temperature and high humidity, measurable degradation appears after 5–7 days. Desiccation or use of silica gel mitigates such effects even without refrigeration.
2.7 Summary
Enzymatic, hydrolytic, oxidative, and microbial processes collectively determine DNA stability. Dry, cool, and low-humidity storage minimizes these risks, forming the basis for best-practice preservation guidelines.
3. Review of Evidence on Two-Week Storage Stability
3.1 Overview
Empirical studies consistently show that buccal swab DNA is stable for up to 14 days under proper storage. Temperature and humidity are the main determinants of degradation rate.
3.2 Representative studies
|
Study |
Storage Conditions |
Duration |
Main Findings |
Implications |
|
Giles et al., 2009, Eur J Epidemiol |
4 °C, buffer |
0–14 days |
No decline in yield or purity |
Stable for research storage |
|
Nuraini et al., 2023, Med J Indonesia |
4 °C, neonatal swabs |
14 days |
Purity (1.75–1.96) stable; PCR success 100 % |
Suitable for sequencing |
|
Rachmawati et al., 2020, J Teknol Lab |
RT vs 4 °C |
7 days |
Quantity drops at RT; STRs still amplifiable |
Refrigeration advised |
|
Sari et al., 2026, Univ Airlangga Repo |
Dry, 4 °C |
14 days |
No yield or purity change |
Confirms short-term stability |
|
Kebede et al., 2018, Afr J Biotechnol |
RT, humid |
15 days |
30–40 % yield loss after 7–14 days |
Degradation in tropical climate |
3.3 Forensic relevance
STR profiles remain complete after 14 days at 4 °C but may become partial under warm or humid conditions. Properly stored samples meet evidentiary standards for identification, though chain-of-custody documentation of storage conditions is essential.
3.4 Genetic and biomedical relevance
For mailed or field-collected samples, 14-day storage under dry conditions yields DNA suitable for genotyping, sequencing, and array analysis. Only extreme humidity significantly reduces performance.
3.5 Synthesis
Collectively, the studies demonstrate that refrigerated or desiccated storage preserves DNA integrity for two weeks, whereas warm, moist conditions induce degradation.
4. Forensic and Genetic Implications and Recommendations
4.1 Forensic applications
In forensic casework, buccal swabs serve as reference samples for STR profiling. When stored ≤ 4 °C or air-dried, full STR profiles remain obtainable after two weeks. Even minor degradation rarely compromises identification, though allele dropouts can occur in humid storage. Documentation of time and temperature during transport enhances evidentiary reliability.
4.2 Genetic and biomedical applications
Population studies and clinical genetics rely on self-collected swabs often transported by mail. Empirical data show that dried samples remain stable for up to 14 days in transit. DNA from such samples supports PCR, SNP genotyping, and sequencing applications. Only assays requiring ultra-long fragments, such as long-read sequencing or methylation profiling, demand fresher samples.
4.3 Best-practice recommendations
4.4 Policy and research implications
Standardized two-week stability data support operational flexibility in forensic and research settings. Future protocols should incorporate temperature monitoring, desiccant use, and clear labeling. Extended-duration studies under extreme conditions are still needed.
5. CONCLUSIONS AND FUTURE PERSPECTIVES
Buccal swabs provide a robust and reliable source of genomic DNA for both forensic and genetic applications. The collective evidence indicates that DNA integrity remains uncompromised for up to two weeks under refrigerated or dry conditions. Degradation becomes problematic primarily under high humidity or heat.
Forensic practice benefits from the assurance that reference samples stored ≤ 14 days retain evidentiary validity, while genetic research gains logistical flexibility for remote and large-scale collections. Continued research into swab materials, field stabilization technologies, and standardized reporting will further enhance reproducibility.
Overall, the simplicity, affordability, and proven short-term stability of buccal swabs affirm their continued role as a cornerstone of modern forensic and genomic DNA collection.
REFERENCES
Madhu Kalasad*, Aboli Dagamwar, Pratiksha Patil, Damini Patil, Ruchi Bhuran, Mahesh Thange, Storage-Related Variations in buccal swab DNA integrity over two weeks: Forensic and Genetic Considerations, Int. J. of Pharm. Sci., 2025, Vol 3, Issue 11, 266-271 https://doi.org/10.5281/zenodo.17511934
10.5281/zenodo.17511934
