Kasturi Selvam , Mohamad Ahmad Najib , Muhammad Fazli Khalid , Muhammad Hafiznur Yunus , Habibah A. Wahab , Azian Harun , Ummu Afeera Zainulabid , Khairul Mohd Fadzli Mustaffa , Ismail Aziah
{"title":"针对假丝酵母伯克霍尔德氏菌 BipD 抗原的 ssDNA 短肽的分离与表征","authors":"Kasturi Selvam , Mohamad Ahmad Najib , Muhammad Fazli Khalid , Muhammad Hafiznur Yunus , Habibah A. Wahab , Azian Harun , Ummu Afeera Zainulabid , Khairul Mohd Fadzli Mustaffa , Ismail Aziah","doi":"10.1016/j.ab.2024.115655","DOIUrl":null,"url":null,"abstract":"<div><h3>Background</h3><p>Melioidosis is difficult to diagnose due to its wide range of clinical symptoms. The culture method is time-consuming and less sensitive, emphasizing the importance of rapid and accurate diagnostic tests for melioidosis. <em>Burkholderia</em> invasion protein D (BipD) of <em>Burkholderia pseudomallei</em> is a potential diagnostic biomarker. This study aimed to isolate and characterize single-stranded DNA aptamers that specifically target BipD.</p></div><div><h3>Methods</h3><p>The recombinant BipD protein was produced, followed by isolation of BipD-specific aptamers using Systematic Evolution of Ligands by EXponential enrichment. The binding affinity and specificity of the selected aptamers were evaluated using Enzyme-Linked Oligonucleotide Assay.</p></div><div><h3>Results</h3><p>The fifth SELEX cycle showed a notable enrichment of recombinant BipD protein-specific aptamers. Sequencing analysis identified two clusters with a total of seventeen distinct aptamers. AptBipD1, AptBipD13, and AptBipD50 were chosen based on their frequency. Among them, AptBipD1 exhibited the highest binding affinity with a <em>K</em><sub>d</sub> value of 1.0 μM for the recombinant BipD protein. Furthermore, AptBipD1 showed significant specificity for <em>B. pseudomallei</em> compared to other tested bacteria.</p></div><div><h3>Conclusion</h3><p>AptBipD1 is a promising candidate for further development of reliable, affordable, and efficient point-of-care diagnostic tests for melioidosis.</p></div>","PeriodicalId":7830,"journal":{"name":"Analytical biochemistry","volume":"695 ","pages":"Article 115655"},"PeriodicalIF":2.6000,"publicationDate":"2024-08-28","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":"0","resultStr":"{\"title\":\"Isolation and characterization of ssDNA aptamers against BipD antigen of Burkholderia pseudomallei\",\"authors\":\"Kasturi Selvam , Mohamad Ahmad Najib , Muhammad Fazli Khalid , Muhammad Hafiznur Yunus , Habibah A. Wahab , Azian Harun , Ummu Afeera Zainulabid , Khairul Mohd Fadzli Mustaffa , Ismail Aziah\",\"doi\":\"10.1016/j.ab.2024.115655\",\"DOIUrl\":null,\"url\":null,\"abstract\":\"<div><h3>Background</h3><p>Melioidosis is difficult to diagnose due to its wide range of clinical symptoms. The culture method is time-consuming and less sensitive, emphasizing the importance of rapid and accurate diagnostic tests for melioidosis. <em>Burkholderia</em> invasion protein D (BipD) of <em>Burkholderia pseudomallei</em> is a potential diagnostic biomarker. This study aimed to isolate and characterize single-stranded DNA aptamers that specifically target BipD.</p></div><div><h3>Methods</h3><p>The recombinant BipD protein was produced, followed by isolation of BipD-specific aptamers using Systematic Evolution of Ligands by EXponential enrichment. The binding affinity and specificity of the selected aptamers were evaluated using Enzyme-Linked Oligonucleotide Assay.</p></div><div><h3>Results</h3><p>The fifth SELEX cycle showed a notable enrichment of recombinant BipD protein-specific aptamers. Sequencing analysis identified two clusters with a total of seventeen distinct aptamers. AptBipD1, AptBipD13, and AptBipD50 were chosen based on their frequency. Among them, AptBipD1 exhibited the highest binding affinity with a <em>K</em><sub>d</sub> value of 1.0 μM for the recombinant BipD protein. Furthermore, AptBipD1 showed significant specificity for <em>B. pseudomallei</em> compared to other tested bacteria.</p></div><div><h3>Conclusion</h3><p>AptBipD1 is a promising candidate for further development of reliable, affordable, and efficient point-of-care diagnostic tests for melioidosis.</p></div>\",\"PeriodicalId\":7830,\"journal\":{\"name\":\"Analytical biochemistry\",\"volume\":\"695 \",\"pages\":\"Article 115655\"},\"PeriodicalIF\":2.6000,\"publicationDate\":\"2024-08-28\",\"publicationTypes\":\"Journal Article\",\"fieldsOfStudy\":null,\"isOpenAccess\":false,\"openAccessPdf\":\"\",\"citationCount\":\"0\",\"resultStr\":null,\"platform\":\"Semanticscholar\",\"paperid\":null,\"PeriodicalName\":\"Analytical biochemistry\",\"FirstCategoryId\":\"99\",\"ListUrlMain\":\"https://www.sciencedirect.com/science/article/pii/S0003269724001994\",\"RegionNum\":4,\"RegionCategory\":\"生物学\",\"ArticlePicture\":[],\"TitleCN\":null,\"AbstractTextCN\":null,\"PMCID\":null,\"EPubDate\":\"\",\"PubModel\":\"\",\"JCR\":\"Q2\",\"JCRName\":\"BIOCHEMICAL RESEARCH METHODS\",\"Score\":null,\"Total\":0}","platform":"Semanticscholar","paperid":null,"PeriodicalName":"Analytical biochemistry","FirstCategoryId":"99","ListUrlMain":"https://www.sciencedirect.com/science/article/pii/S0003269724001994","RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":null,"EPubDate":"","PubModel":"","JCR":"Q2","JCRName":"BIOCHEMICAL RESEARCH METHODS","Score":null,"Total":0}
Isolation and characterization of ssDNA aptamers against BipD antigen of Burkholderia pseudomallei
Background
Melioidosis is difficult to diagnose due to its wide range of clinical symptoms. The culture method is time-consuming and less sensitive, emphasizing the importance of rapid and accurate diagnostic tests for melioidosis. Burkholderia invasion protein D (BipD) of Burkholderia pseudomallei is a potential diagnostic biomarker. This study aimed to isolate and characterize single-stranded DNA aptamers that specifically target BipD.
Methods
The recombinant BipD protein was produced, followed by isolation of BipD-specific aptamers using Systematic Evolution of Ligands by EXponential enrichment. The binding affinity and specificity of the selected aptamers were evaluated using Enzyme-Linked Oligonucleotide Assay.
Results
The fifth SELEX cycle showed a notable enrichment of recombinant BipD protein-specific aptamers. Sequencing analysis identified two clusters with a total of seventeen distinct aptamers. AptBipD1, AptBipD13, and AptBipD50 were chosen based on their frequency. Among them, AptBipD1 exhibited the highest binding affinity with a Kd value of 1.0 μM for the recombinant BipD protein. Furthermore, AptBipD1 showed significant specificity for B. pseudomallei compared to other tested bacteria.
Conclusion
AptBipD1 is a promising candidate for further development of reliable, affordable, and efficient point-of-care diagnostic tests for melioidosis.
期刊介绍：
The journal''s title Analytical Biochemistry: Methods in the Biological Sciences declares its broad scope: methods for the basic biological sciences that include biochemistry, molecular genetics, cell biology, proteomics, immunology, bioinformatics and wherever the frontiers of research take the field.
The emphasis is on methods from the strictly analytical to the more preparative that would include novel approaches to protein purification as well as improvements in cell and organ culture. The actual techniques are equally inclusive ranging from aptamers to zymology.
The journal has been particularly active in:
-Analytical techniques for biological molecules-
Aptamer selection and utilization-
Biosensors-
Chromatography-
Cloning, sequencing and mutagenesis-
Electrochemical methods-
Electrophoresis-
Enzyme characterization methods-
Immunological approaches-
Mass spectrometry of proteins and nucleic acids-
Metabolomics-
Nano level techniques-
Optical spectroscopy in all its forms.
The journal is reluctant to include most drug and strictly clinical studies as there are more suitable publication platforms for these types of papers.
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