Transformation and Characterization of Δ12-Fatty Acid Acetylenase and Δ12-Oleate Desaturase Potentially Involved in the Polyacetylene Biosynthetic Pathway from <i>Bidens pilosa</i>
<i>Bidens pilosa</i> is commonly used as an herbal tea component or traditional medicine for treating several diseases, including diabetes. Polyacetylenes have two or more carbon–carbon triple bonds or alkynyl functional groups and are mainly derived from fatty acid and polyketide precur...
| Main Authors: | , , , , , , |
|---|---|
| Format: | Article |
| Language: | English |
| Published: |
MDPI AG
2020-11-01
|
| Series: | Plants |
| Subjects: | |
| Online Access: | https://www.mdpi.com/2223-7747/9/11/1483 |
| _version_ | 1797548858285228032 |
|---|---|
| author | Po-Yen Chen Mi-Jou Hsieh Yung-Ting Tsai Hsiao-Hang Chung Lie-Fen Shyur Cheng-Han Hsieh Kin-Ying To |
| author_facet | Po-Yen Chen Mi-Jou Hsieh Yung-Ting Tsai Hsiao-Hang Chung Lie-Fen Shyur Cheng-Han Hsieh Kin-Ying To |
| author_sort | Po-Yen Chen |
| collection | DOAJ |
| description | <i>Bidens pilosa</i> is commonly used as an herbal tea component or traditional medicine for treating several diseases, including diabetes. Polyacetylenes have two or more carbon–carbon triple bonds or alkynyl functional groups and are mainly derived from fatty acid and polyketide precursors. Here, we report the cloning of full-length cDNAs that encode Δ12-fatty acid acetylenase (designated BPFAA) and Δ12-oleate desaturase (designated BPOD) from <i>B. pilosa</i>, which we predicted to play a role in the polyacetylene biosynthetic pathway. Subsequently, expression vectors carrying BPFAA or BPOD were constructed and transformed into <i>B. pilosa</i> via the <i>Agrobacterium</i>-mediated method. Genomic PCR analysis confirmed the presence of transgenes and selection marker genes in the obtained transgenic lines. The copy numbers of transgenes in transgenic lines were determined by Southern blot analysis. Furthermore, 4–5 <i>FAA</i> genes and 2–3 <i>OD</i> genes were detected in wild-type (WT) plants. Quantitative real time-PCR revealed that some transgenic lines had higher expression levels than WT. Western blot analysis revealed OD protein expression in the selected transformants. High-performance liquid chromatography profiling was used to analyze the seven index polyacetylenic compounds, and fluctuation patterns were found. |
| first_indexed | 2024-03-10T15:06:46Z |
| format | Article |
| id | doaj.art-19d29c1779a349099d7db24ef12a0485 |
| institution | Directory Open Access Journal |
| issn | 2223-7747 |
| language | English |
| last_indexed | 2024-03-10T15:06:46Z |
| publishDate | 2020-11-01 |
| publisher | MDPI AG |
| record_format | Article |
| series | Plants |
| spelling | doaj.art-19d29c1779a349099d7db24ef12a04852023-11-20T19:41:43ZengMDPI AGPlants2223-77472020-11-01911148310.3390/plants9111483Transformation and Characterization of Δ12-Fatty Acid Acetylenase and Δ12-Oleate Desaturase Potentially Involved in the Polyacetylene Biosynthetic Pathway from <i>Bidens pilosa</i>Po-Yen Chen0Mi-Jou Hsieh1Yung-Ting Tsai2Hsiao-Hang Chung3Lie-Fen Shyur4Cheng-Han Hsieh5Kin-Ying To6Agricultural Biotechnology Research Center, Academia Sinica, Taipei 115, TaiwanAgricultural Biotechnology Research Center, Academia Sinica, Taipei 115, TaiwanAgricultural Biotechnology Research Center, Academia Sinica, Taipei 115, TaiwanDepartment of Horticulture, National Ilan University, Yilan 260, TaiwanAgricultural Biotechnology Research Center, Academia Sinica, Taipei 115, TaiwanAgricultural Biotechnology Research Center, Academia Sinica, Taipei 115, TaiwanAgricultural Biotechnology Research Center, Academia Sinica, Taipei 115, Taiwan<i>Bidens pilosa</i> is commonly used as an herbal tea component or traditional medicine for treating several diseases, including diabetes. Polyacetylenes have two or more carbon–carbon triple bonds or alkynyl functional groups and are mainly derived from fatty acid and polyketide precursors. Here, we report the cloning of full-length cDNAs that encode Δ12-fatty acid acetylenase (designated BPFAA) and Δ12-oleate desaturase (designated BPOD) from <i>B. pilosa</i>, which we predicted to play a role in the polyacetylene biosynthetic pathway. Subsequently, expression vectors carrying BPFAA or BPOD were constructed and transformed into <i>B. pilosa</i> via the <i>Agrobacterium</i>-mediated method. Genomic PCR analysis confirmed the presence of transgenes and selection marker genes in the obtained transgenic lines. The copy numbers of transgenes in transgenic lines were determined by Southern blot analysis. Furthermore, 4–5 <i>FAA</i> genes and 2–3 <i>OD</i> genes were detected in wild-type (WT) plants. Quantitative real time-PCR revealed that some transgenic lines had higher expression levels than WT. Western blot analysis revealed OD protein expression in the selected transformants. High-performance liquid chromatography profiling was used to analyze the seven index polyacetylenic compounds, and fluctuation patterns were found.https://www.mdpi.com/2223-7747/9/11/1483polyacetylenes<i>Bidens pilosa</i>acetylenasedesaturase<i>Agrobacterium</i>-mediated transformation<i>FAD2</i> |
| spellingShingle | Po-Yen Chen Mi-Jou Hsieh Yung-Ting Tsai Hsiao-Hang Chung Lie-Fen Shyur Cheng-Han Hsieh Kin-Ying To Transformation and Characterization of Δ12-Fatty Acid Acetylenase and Δ12-Oleate Desaturase Potentially Involved in the Polyacetylene Biosynthetic Pathway from <i>Bidens pilosa</i> Plants polyacetylenes <i>Bidens pilosa</i> acetylenase desaturase <i>Agrobacterium</i>-mediated transformation <i>FAD2</i> |
| title | Transformation and Characterization of Δ12-Fatty Acid Acetylenase and Δ12-Oleate Desaturase Potentially Involved in the Polyacetylene Biosynthetic Pathway from <i>Bidens pilosa</i> |
| title_full | Transformation and Characterization of Δ12-Fatty Acid Acetylenase and Δ12-Oleate Desaturase Potentially Involved in the Polyacetylene Biosynthetic Pathway from <i>Bidens pilosa</i> |
| title_fullStr | Transformation and Characterization of Δ12-Fatty Acid Acetylenase and Δ12-Oleate Desaturase Potentially Involved in the Polyacetylene Biosynthetic Pathway from <i>Bidens pilosa</i> |
| title_full_unstemmed | Transformation and Characterization of Δ12-Fatty Acid Acetylenase and Δ12-Oleate Desaturase Potentially Involved in the Polyacetylene Biosynthetic Pathway from <i>Bidens pilosa</i> |
| title_short | Transformation and Characterization of Δ12-Fatty Acid Acetylenase and Δ12-Oleate Desaturase Potentially Involved in the Polyacetylene Biosynthetic Pathway from <i>Bidens pilosa</i> |
| title_sort | transformation and characterization of δ12 fatty acid acetylenase and δ12 oleate desaturase potentially involved in the polyacetylene biosynthetic pathway from i bidens pilosa i |
| topic | polyacetylenes <i>Bidens pilosa</i> acetylenase desaturase <i>Agrobacterium</i>-mediated transformation <i>FAD2</i> |
| url | https://www.mdpi.com/2223-7747/9/11/1483 |
| work_keys_str_mv | AT poyenchen transformationandcharacterizationofd12fattyacidacetylenaseandd12oleatedesaturasepotentiallyinvolvedinthepolyacetylenebiosyntheticpathwayfromibidenspilosai AT mijouhsieh transformationandcharacterizationofd12fattyacidacetylenaseandd12oleatedesaturasepotentiallyinvolvedinthepolyacetylenebiosyntheticpathwayfromibidenspilosai AT yungtingtsai transformationandcharacterizationofd12fattyacidacetylenaseandd12oleatedesaturasepotentiallyinvolvedinthepolyacetylenebiosyntheticpathwayfromibidenspilosai AT hsiaohangchung transformationandcharacterizationofd12fattyacidacetylenaseandd12oleatedesaturasepotentiallyinvolvedinthepolyacetylenebiosyntheticpathwayfromibidenspilosai AT liefenshyur transformationandcharacterizationofd12fattyacidacetylenaseandd12oleatedesaturasepotentiallyinvolvedinthepolyacetylenebiosyntheticpathwayfromibidenspilosai AT chenghanhsieh transformationandcharacterizationofd12fattyacidacetylenaseandd12oleatedesaturasepotentiallyinvolvedinthepolyacetylenebiosyntheticpathwayfromibidenspilosai AT kinyingto transformationandcharacterizationofd12fattyacidacetylenaseandd12oleatedesaturasepotentiallyinvolvedinthepolyacetylenebiosyntheticpathwayfromibidenspilosai |