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Characterization of DNA aptamers generated against the soft-shelled turtle iridovirus with antiviral effects.

Li P, Zhou L, Yu Y, Yang M, Ni S, Wei S, Qin Q - BMC Vet. Res. (2015)

Bottom Line: Soft-shelled turtle iridovirus (STIV) causes severe systemic disease in farmed soft-shelled turtles (Trionyx sinensis).Electrophoretic mobility shift assays and fluorescent localization showed that the selected aptamers had high binding affinity for STIV.Aptamer QA-36 had the highest calculated binding affinity (K d ) of 53.8 nM.

View Article: PubMed Central - PubMed

Affiliation: Key Laboratory of Tropical Marine Bio-resources and Ecology, South China Sea Institute of Oceanology, Chinese Academy of Sciences, 164 West Xingang Road, Guangzhou, 510301, China.

ABSTRACT

Background: Soft-shelled turtle iridovirus (STIV) causes severe systemic disease in farmed soft-shelled turtles (Trionyx sinensis). More efficient methods of controlling and detecting STIV infections are urgently needed. 

Methods: In this study, we generated eight single-stranded DNA (ssDNA) aptamers against STIV using systematic evolution of ligands by exponential enrichment (SELEX).

Results: The aptamers formed representative stem-loop secondary structures. Electrophoretic mobility shift assays and fluorescent localization showed that the selected aptamers had high binding affinity for STIV. Aptamer QA-36 had the highest calculated binding affinity (K d ) of 53.8 nM. Flow cytometry and fluorescence microscopy of cell-aptamer interactions demonstrated that QA-12 was able to recognize both STIV-infected cells and tissues with a high level of specificity. Moreover, the selected aptamers inhibited STIV infection in vitro and in vivo, with aptamer QA-36 demonstrating the greatest protective effect against STIV and inhibiting STIV infection in a dose-dependent manner.

Discussion: We generated DNA aptamers that bound STIV with a high level of specificity, providing an alternative means for investigating STIV pathogenesis, drug development, and medical therapies for STIV infection.

Conclusions: These DNA aptamers may thus be suitable antiviral candidates for the control of STIV infections.

No MeSH data available.


Related in: MedlinePlus

Affinity of STIV-selected ssDNA aptamer interactions by ELISA. STIV was incubated with increasing concentrations of 5′-biotinylated aptamer. After addition of streptavidin-HRP, the amount of STIV-aptamer complex was calculated and graphed as a function of aptamer concentration. The graph was fit to the equation Y = Bmax*X/(Kd + X) using SigmaPlot software. Results for each aptamer was presented as the mean ± SD of three independent experiments
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Fig3: Affinity of STIV-selected ssDNA aptamer interactions by ELISA. STIV was incubated with increasing concentrations of 5′-biotinylated aptamer. After addition of streptavidin-HRP, the amount of STIV-aptamer complex was calculated and graphed as a function of aptamer concentration. The graph was fit to the equation Y = Bmax*X/(Kd + X) using SigmaPlot software. Results for each aptamer was presented as the mean ± SD of three independent experiments

Mentions: The binding affinities of the aptamers to STIV were assessed by ELISA using 5′-biotinylated aptamers. All the four selected aptamers showed high affinity for STIV, with calculated binding affinities (Kd) of 77.5 nM for QA9, 80.7 nM for QA12, 53.8 nM for QA36, and 71.2 nM for QA92 (Fig. 3). Of the four aptamer candidates, the Kd value of QA36 indicated the highest affinity.Fig. 3


Characterization of DNA aptamers generated against the soft-shelled turtle iridovirus with antiviral effects.

Li P, Zhou L, Yu Y, Yang M, Ni S, Wei S, Qin Q - BMC Vet. Res. (2015)

Affinity of STIV-selected ssDNA aptamer interactions by ELISA. STIV was incubated with increasing concentrations of 5′-biotinylated aptamer. After addition of streptavidin-HRP, the amount of STIV-aptamer complex was calculated and graphed as a function of aptamer concentration. The graph was fit to the equation Y = Bmax*X/(Kd + X) using SigmaPlot software. Results for each aptamer was presented as the mean ± SD of three independent experiments
© Copyright Policy - OpenAccess
Related In: Results  -  Collection

License 1 - License 2
Show All Figures
getmorefigures.php?uid=PMC4588899&req=5

Fig3: Affinity of STIV-selected ssDNA aptamer interactions by ELISA. STIV was incubated with increasing concentrations of 5′-biotinylated aptamer. After addition of streptavidin-HRP, the amount of STIV-aptamer complex was calculated and graphed as a function of aptamer concentration. The graph was fit to the equation Y = Bmax*X/(Kd + X) using SigmaPlot software. Results for each aptamer was presented as the mean ± SD of three independent experiments
Mentions: The binding affinities of the aptamers to STIV were assessed by ELISA using 5′-biotinylated aptamers. All the four selected aptamers showed high affinity for STIV, with calculated binding affinities (Kd) of 77.5 nM for QA9, 80.7 nM for QA12, 53.8 nM for QA36, and 71.2 nM for QA92 (Fig. 3). Of the four aptamer candidates, the Kd value of QA36 indicated the highest affinity.Fig. 3

Bottom Line: Soft-shelled turtle iridovirus (STIV) causes severe systemic disease in farmed soft-shelled turtles (Trionyx sinensis).Electrophoretic mobility shift assays and fluorescent localization showed that the selected aptamers had high binding affinity for STIV.Aptamer QA-36 had the highest calculated binding affinity (K d ) of 53.8 nM.

View Article: PubMed Central - PubMed

Affiliation: Key Laboratory of Tropical Marine Bio-resources and Ecology, South China Sea Institute of Oceanology, Chinese Academy of Sciences, 164 West Xingang Road, Guangzhou, 510301, China.

ABSTRACT

Background: Soft-shelled turtle iridovirus (STIV) causes severe systemic disease in farmed soft-shelled turtles (Trionyx sinensis). More efficient methods of controlling and detecting STIV infections are urgently needed. 

Methods: In this study, we generated eight single-stranded DNA (ssDNA) aptamers against STIV using systematic evolution of ligands by exponential enrichment (SELEX).

Results: The aptamers formed representative stem-loop secondary structures. Electrophoretic mobility shift assays and fluorescent localization showed that the selected aptamers had high binding affinity for STIV. Aptamer QA-36 had the highest calculated binding affinity (K d ) of 53.8 nM. Flow cytometry and fluorescence microscopy of cell-aptamer interactions demonstrated that QA-12 was able to recognize both STIV-infected cells and tissues with a high level of specificity. Moreover, the selected aptamers inhibited STIV infection in vitro and in vivo, with aptamer QA-36 demonstrating the greatest protective effect against STIV and inhibiting STIV infection in a dose-dependent manner.

Discussion: We generated DNA aptamers that bound STIV with a high level of specificity, providing an alternative means for investigating STIV pathogenesis, drug development, and medical therapies for STIV infection.

Conclusions: These DNA aptamers may thus be suitable antiviral candidates for the control of STIV infections.

No MeSH data available.


Related in: MedlinePlus