In vitro selection and characterization of single stranded DNA aptamers for luteolin
Thesis (PhD. (Biosciences))
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| Format: | Thesis |
| Language: | English |
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Universiti Teknologi Malaysia
2023
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| Online Access: | http://openscience.utm.my/handle/123456789/626 |
| _version_ | 1825623448347475968 |
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| author | Sabah, Jinan Tuma |
| author_facet | Sabah, Jinan Tuma |
| author_sort | Sabah, Jinan Tuma |
| collection | OpenScience |
| description | Thesis (PhD. (Biosciences)) |
| first_indexed | 2024-03-05T17:34:25Z |
| format | Thesis |
| id | oai:openscience.utm.my:123456789/626 |
| institution | Universiti Teknologi Malaysia - OpenScience |
| language | English |
| last_indexed | 2024-03-05T17:34:25Z |
| publishDate | 2023 |
| publisher | Universiti Teknologi Malaysia |
| record_format | dspace |
| spelling | oai:openscience.utm.my:123456789/6262023-08-21T15:01:15Z In vitro selection and characterization of single stranded DNA aptamers for luteolin Sabah, Jinan Tuma Flavonoids—Analysis DNA Thesis (PhD. (Biosciences)) Luteolin (LUT), also known as 3',4',5,7-Tetrahydroxy-flavone, a flavonoid belongs to the flavone family. It has a broad range of biochemical and pharmacological properties. Aptamers are single stranded deoxyribonucleic acid (ssDNA) or ribonucleic acid (RNA) oligonucleotides that can bind to their targets with high affinity and specificity based on their specific three-dimensional structures. In this study, DNA aptamers that specifically recognize LUT as their target were selected using in vitro selection method called Systematic Evolution of Ligands by Exponential enrichment (SELEX). 76-mer library containing 1015 randomized ssDNA molecules were screened for 8th SELEX cycle with approximately 80 % of the loaded ssDNA was eluted. The recovered ssDNA pool from the 8th SELEX round was amplified with unlabeled primers and cloned into PSTBlue -1 vector. The clones were sent for sequencing. Twenty two of such LUT binding aptamer variants were further analyzed by placing into one of the seven groups depending on their N40 random sequence regions similarity. One representative from each group was selected randomly for further analysis, finalizing with three aptamers designed as LUT#28, LUT#20 and LUT#3 showed high affinity to LUT with Kd values 107.054, 214.623 and 109.781 nM by using the nonlinear regression analysis used SIGMAPLOT 11.0 software, respectively. Analysis prediction of the secondary structure of these aptamers showed highly distinct feature with typical stem and loop. Further specificity experimental analysis with these three aptamers showed that LUT#3 had higher specificity with insignificant binding toward kaempferol and quercetin despite its structural and functional similarity. Conversely, LUT#28 and LUT#20 show high affinity but no specificity. Further study showed that LUT#3 can detect free luteolin in the range of 0.2 to 1 mM in solution. It was suggested that LUT#3 aptamer was the most suitable for luteolin recognition tool at laboratory scale based on the condition tested. The high affinity candidate shows promise for later applications in an aptamer-based system to rapidly luteolin identification Faculty of Biosciences and Medical Engineering 2023-08-21T06:30:37Z 2023-08-21T06:30:37Z 2018 Thesis Dataset NA NA http://openscience.utm.my/handle/123456789/626 en NA; NA application/pdf application/pdf application/pdf application/pdf application/pdf application/pdf Universiti Teknologi Malaysia |
| spellingShingle | Flavonoids—Analysis DNA Sabah, Jinan Tuma In vitro selection and characterization of single stranded DNA aptamers for luteolin |
| title | In vitro selection and characterization of single stranded DNA aptamers for luteolin |
| title_full | In vitro selection and characterization of single stranded DNA aptamers for luteolin |
| title_fullStr | In vitro selection and characterization of single stranded DNA aptamers for luteolin |
| title_full_unstemmed | In vitro selection and characterization of single stranded DNA aptamers for luteolin |
| title_short | In vitro selection and characterization of single stranded DNA aptamers for luteolin |
| title_sort | in vitro selection and characterization of single stranded dna aptamers for luteolin |
| topic | Flavonoids—Analysis DNA |
| url | http://openscience.utm.my/handle/123456789/626 |
| work_keys_str_mv | AT sabahjinantuma invitroselectionandcharacterizationofsinglestrandeddnaaptamersforluteolin |