Development of a miniaturized Ti-plasmid and helper plasmid system for Agrobacterium-mediated plant transformation
Tumor-inducing (Ti) plasmid is the requisite for Agrobacterium-mediated plant transformation. Over decades, continuous efforts have been made to improve the efficiency of Agrobacterium-mediated plant transformation and most of them focused on the binary vector system. A binary vector system comprise...
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| Format: | Article |
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Malaysian Society of Molecular Biology and Biotechnology
2023
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| _version_ | 1796983924719616000 |
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| author | Yuh, Leng Teo Shu, Ting Chang Wai, Keat Toh Xin, Yen Tor Chai, Ling Ho Pek, Chin Loh Hann, Ling Wong |
| author_facet | Yuh, Leng Teo Shu, Ting Chang Wai, Keat Toh Xin, Yen Tor Chai, Ling Ho Pek, Chin Loh Hann, Ling Wong |
| author_sort | Yuh, Leng Teo |
| collection | UPM |
| description | Tumor-inducing (Ti) plasmid is the requisite for Agrobacterium-mediated plant transformation. Over decades, continuous efforts have been made to improve the efficiency of Agrobacterium-mediated plant transformation and most of them focused on the binary vector system. A binary vector system comprises of a binary vector of which transferred DNA (T-DNA) resided on and a Ti plasmid to carry those essential virulence genes. In this study, we constructed a miniaturized helper Ti plasmid, designated as pYL102, with the aim to enhance the overall Agrobacterium-mediated transformation rate. The size of pYL102 was reduced to ~60% of the original plasmid pCAMBIA5105. Subsequently, pYL102 was coupled with the broad host range (BHR) bacterial expression vector, pYL101C, of which the key regulatory virulence gene, virG-N54D, was cloned in and expressed under the control of a strong constitutive PINTc promoter. To test the functionality of the constructed vector system, A. tumefaciens C58C1 carrying pYL102, pYL101C::virG-N54D and the transformation vector pGWB2::e35S-sfGFP was used to transform Nicotiana benthamiana leaves by agroinfiltration. Green fluorescence was observed in spots infiltrated with Agrobacterium carrying the test plasmids. The fluorescence intensity from the test agroinfiltrated leaves was significantly higher than those of the mock-infiltrated leaves (p<0.01), indicating the vector system can be used for plant transformation. |
| first_indexed | 2024-03-06T11:13:18Z |
| format | Article |
| id | upm.eprints-100596 |
| institution | Universiti Putra Malaysia |
| last_indexed | 2024-03-06T11:13:18Z |
| publishDate | 2023 |
| publisher | Malaysian Society of Molecular Biology and Biotechnology |
| record_format | dspace |
| spelling | upm.eprints-1005962023-10-09T18:37:19Z http://psasir.upm.edu.my/id/eprint/100596/ Development of a miniaturized Ti-plasmid and helper plasmid system for Agrobacterium-mediated plant transformation Yuh, Leng Teo Shu, Ting Chang Wai, Keat Toh Xin, Yen Tor Chai, Ling Ho Pek, Chin Loh Hann, Ling Wong Tumor-inducing (Ti) plasmid is the requisite for Agrobacterium-mediated plant transformation. Over decades, continuous efforts have been made to improve the efficiency of Agrobacterium-mediated plant transformation and most of them focused on the binary vector system. A binary vector system comprises of a binary vector of which transferred DNA (T-DNA) resided on and a Ti plasmid to carry those essential virulence genes. In this study, we constructed a miniaturized helper Ti plasmid, designated as pYL102, with the aim to enhance the overall Agrobacterium-mediated transformation rate. The size of pYL102 was reduced to ~60% of the original plasmid pCAMBIA5105. Subsequently, pYL102 was coupled with the broad host range (BHR) bacterial expression vector, pYL101C, of which the key regulatory virulence gene, virG-N54D, was cloned in and expressed under the control of a strong constitutive PINTc promoter. To test the functionality of the constructed vector system, A. tumefaciens C58C1 carrying pYL102, pYL101C::virG-N54D and the transformation vector pGWB2::e35S-sfGFP was used to transform Nicotiana benthamiana leaves by agroinfiltration. Green fluorescence was observed in spots infiltrated with Agrobacterium carrying the test plasmids. The fluorescence intensity from the test agroinfiltrated leaves was significantly higher than those of the mock-infiltrated leaves (p<0.01), indicating the vector system can be used for plant transformation. Malaysian Society of Molecular Biology and Biotechnology 2023-01-19 Article PeerReviewed Yuh, Leng Teo and Shu, Ting Chang and Wai, Keat Toh and Xin, Yen Tor and Chai, Ling Ho and Pek, Chin Loh and Hann, Ling Wong (2023) Development of a miniaturized Ti-plasmid and helper plasmid system for Agrobacterium-mediated plant transformation. Asia Pacific Journal of Molecular Biology and Biotechnology, 30 (3). 23 - 32. ISSN 0128-7451; ESSN:2672-7277 https://www.msmbb.my/index.php/archive-issues/18-apjmbb/461-archive-issue-30 10.35118/apjmbb.2022.030.3.03 |
| spellingShingle | Yuh, Leng Teo Shu, Ting Chang Wai, Keat Toh Xin, Yen Tor Chai, Ling Ho Pek, Chin Loh Hann, Ling Wong Development of a miniaturized Ti-plasmid and helper plasmid system for Agrobacterium-mediated plant transformation |
| title | Development of a miniaturized Ti-plasmid and helper plasmid system for Agrobacterium-mediated plant transformation |
| title_full | Development of a miniaturized Ti-plasmid and helper plasmid system for Agrobacterium-mediated plant transformation |
| title_fullStr | Development of a miniaturized Ti-plasmid and helper plasmid system for Agrobacterium-mediated plant transformation |
| title_full_unstemmed | Development of a miniaturized Ti-plasmid and helper plasmid system for Agrobacterium-mediated plant transformation |
| title_short | Development of a miniaturized Ti-plasmid and helper plasmid system for Agrobacterium-mediated plant transformation |
| title_sort | development of a miniaturized ti plasmid and helper plasmid system for agrobacterium mediated plant transformation |
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