Hiding Secret Messages in Living Cells in the Age of Next-Generation Sequencing

Hiding secret messages (SMs) based on DNA technologies plays a critical role in DNA computing. In recent years, scientists have hidden SMs at a specific site in the plasmids of various living cells. Unfortunately, this strategy was proposed without considering the attack of next-generation sequencing (NGS). Nowadays, NGS, which is a revolutionary massively parallel sequencing technique, is developing rapidly. With the help of NGS, sequencing completely unknown genomes is becoming popular in various laboratories throughout the world. The ability to sequence completely unknown genomes is a major threat to existing hiding strategies in living cells. To prevent against the attack of NGS, this paper proposes a scheme to hide SMs in living cells (SHSM). Compared with previous studies, the main contribution of SHSM is changing the specific single hiding site into random multiple hiding sites through the application of seamless clustered regularly interspaced short palindromic repeats/CRISPR-associated9 (CRISPR/Cas9) technique. In SHSM, SMs are hidden in living cells with different puzzling messages at different sites every time to prevent against the attack of NGS. To read the SM, high-fidelity polymerase chain reaction (PCR) and Sanger sequencing are used. A hash function is used to ensure the integrity of the message. These measures mean that an attacker would not be able to determine the location of the SM using NGS. The feasibility of SHSM was validated by a wet-lab experiment, and the security was demonstrated by the system's entropy. The author hopes this study will bring attention to the threat of NGS, and advance the development of hiding SMs in living cells.