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To isolate functional nucleic acids that bind to defined targets with high affinity and specificity, which are known as aptamers, the systematic evolution of ligands by exponential enrichment (SELEX) methodology has emerged as the preferred approach. Here, we propose a computational approach, SELEX in silico, that allows the sequence space to be more thoroughly explored regarding binding of a certain target. Our approach consists of two steps: (i) secondary structure-based sequence screening, which aims to collect the sequences that can form a desired RNA motif as an enhanced initial library, followed by (ii) sequence enrichment regarding target binding by molecular dynamics simulation-based virtual screening. Our SELEX in silico method provided a practical computational solution to three key problems in aptamer sequence searching: design of nucleic acid libraries, knowledge of sequence enrichment, and identification of potent aptamers. Six potent theophylline-binding aptamers, which were isolated by SELEX in silico from a sequence space containing 4(13) sequences, were experimentally verified to bind theophylline with high affinity: Kd ranging from 0.16 to 0.52 μM, compared with the dissociation constant of the original aptamer-theophylline, 0.32 μM. These results demonstrate the significant potential of SELEX in silico as a new method for aptamer discovery and optimization.

Inspired by concerns about food safety, the metabolic landscape of biogenic amines (BAs) was elucidated during industrial semidry Chinese rice wine fermentation. The main fermentation process represented the largest contribution to BA formation, which corresponded to 69.1% (54.3 mg/L). Principal component analysis revealed that total acid and ethanol were strongly correlated with BAs, indicating that BA formation favored acidic and stressful conditions. Other than putrescine (PUT), spermidine (SPD), and spermine (SPM), 5 BAs exhibited strong relationships with the precursor amino acids (R > 0.85). PUT was mainly decarboxylated from arginine (89.6%) whereas SPD (100%) and SPM (83.1%) were obtained from ornithine. Interestingly, some SPD could convert back to PUT (24.3%). All 8 BAs showed good relationships with lactic acid bacteria (LAB) (R around 0.75). Moreover, among the five main LAB genera, Lactobacillus had a positive correlation with BA formation.

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The utilization of amine-negative starter based on an understanding of nitrogen metabolism is a useful method for controlling biogenic amine (BA) in Chinese rice wine (CRW) fermentation. The contribution of brewing materials to protein degradation was analyzed; wheat Qu protein had no effect, and yeast autolysis generated 10% amino nitrogen. Milling degree of rice was strongly correlated with BAs formation ( R = 0.99). Subsequently, Lactobacillus plantarum and Staphylococcus xylosus were coinoculated as amine-negative starter at an optimized ratio of 1:2. Coinoculation induced a significant reduction in total BAs (43.7%, 44.5 mg L), putrescine (43.0%, 20.4 mg L), tyramine (42.8%, 14.3 mg L), and histamine (42.6%, 3.5 mg L) content. Notably, BAs degradation ability of Staphylococcus xylosus was stronger than the suppression effect of Lactobacillus plantarum, and higher lactic acid bacteria (LAB) amount has a positive correlation with lower BAs content. Overall, mixed strains exerted a synergistic effect in lowering BAs accumulation via decarboxylation and transamination.

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