Tissue-SELEX for screening nucleic acid aptamers targeting atherosclerotic tissue
10.20039/j.cnki.1007-3949.2025.11.003
- VernacularTitle:Tissue-SELEX技术筛选靶向动脉粥样硬化组织的核酸适配体
- Author:
Jianmin CHEN
1
;
Zhimian SHI
;
Yanhong LI
;
Haiqing ZHAO
;
Wenwang LANG
;
Zhongcheng MO
;
Jiangbo WANG
Author Information
1. 广西壮族自治区南溪山医院(广西壮族自治区第二人民医院)药学部,广西桂林市 541002
- Publication Type:Journal Article
- Keywords:
atherosclerosis;
systematic evolution of ligands by exponential enrichment;
nucleic acid aptamers
- From:
Chinese Journal of Arteriosclerosis
2025;33(11):937-943
- CountryChina
- Language:Chinese
-
Abstract:
Aim Systematic evolution of ligands by exponential enrichment(SELEX)techniquewas employed to screen and identify nucleic acid aptamers that specifically bind to mouse atherosclerotic pathological tissues,aiming to pro-vide a research foundation for the development of molecular targets and diagnostic reagents for early atherosclerosis.Methods A single-stranded DNA(ssDNA)library with a capacity of 1015～1016 was constructed,which was then subjec-ted to binding-elution(negative selection)with normal mouse vascular tissue slices.The eluted library was subsequently bound to atherosclerotic tissue slices for binding-elution(positive selection).PCR was used to amplify the positive and negative screening products,and agarose gel electrophoresis was used to verify the amplified products.The ssDNA library after multiple rounds of selection was sequenced using T-A cloning and sequencing to obtain the primary structure of the nu-cleic acid aptamers,and the secondary structure was predicted using the Mfold online software.The selected nucleic acid aptamers were labeled with a FAM fluorescent group at the 5'-end and were bound to both positive and negative selection tissue slices,with fluorescence intensity observed under a fluorescence microscope.Image Pro Plus 6.0 was used to cal-culate the relative average fluorescence intensity to evaluate the binding specificity of nucleic acid aptamers.Results After 8 rounds of selection,agarose gel electrophoresis imaging showed PCR amplification products in the positive selection lanes,while no PCR amplification products were observed in the negative selection lanes,indicating the successful acquisi-tion of a nucleic acid aptamer library that specifically binds to atherosclerotic tissues.Five nucleic acid aptamers were i-dentified by T-A cloning and sequencing,and their predicted secondary structures all had stem-loop structures.Immuno-fluorescence staining verified that five nucleic acid aptamers had different degrees of binding with As blood vessels,and the quantitative results of the relative average fluorescence intensity showed that nucleic acid aptamer No.11 had the highest relative average fluorescence intensity value,which can be used as a candidate nucleic acid aptamer for subsequent re-search.Conclusion Specific nucleic acid aptamers that bind to atherosclerotic vesselswere successfully obtained,providing a research foundation for further screening of early molecular targets of Asand developing in vivo early diagnostic reagents.
