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Keywords = LBA 4404

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11 pages, 1213 KiB  
Article
Agrobacterium tumefaciens-Mediated Genetic Transformation of Eclipta alba
by Diwakar Aggarwal, Vasudha Datta, Hardeep Singh Tuli, Pawan Kumar and Seema Ramniwas
Int. J. Plant Biol. 2024, 15(3), 641-651; https://doi.org/10.3390/ijpb15030047 - 10 Jul 2024
Cited by 1 | Viewed by 1518
Abstract
Eclipta alba (Linn.) Hassk. (Asteraceae) is a high value medicinal plant which possesses diverse medicinal properties. It is an important herb for the treatment of various disorders, and is primarily used as a hepatoprotectant. Its major biochemical constituents include wedelolactone and dimethyl-wedelolactone (coumestans), [...] Read more.
Eclipta alba (Linn.) Hassk. (Asteraceae) is a high value medicinal plant which possesses diverse medicinal properties. It is an important herb for the treatment of various disorders, and is primarily used as a hepatoprotectant. Its major biochemical constituents include wedelolactone and dimethyl-wedelolactone (coumestans), which possess anti-hepatotoxic properties. Due to its numerous medicinal properties, it is in high demand by the pharmaceutical industry and therefore requires urgent biotechnological interventions for its improvement. Therefore, the present study was constructed with the aim of developing an efficient genetic transformation protocol for E. alba, which will help in the mass production of the active compounds found in E. alba. Agrobacterium tumefaciens strain LBA 4404, containing vector pBI121, was used to genetically transform the plant, and the effect of various factors such as infection type, light cycle effect, effect of pH, among others, on the genetic transformation efficiency was analyzed. Regenerated transformed shoots were confirmed using the standard Polymerase Chain Reaction PCR method. Kanamycin-resistant and beta- glucurosidaseGUS-positive shoots indicated the development of transgenic shoots in E. alba. Amplification of nptll and uidA genes confirmed the integration of t-DNA transgenic shoots. In conclusion, various factors affecting the transformation efficiency were analyzed, and a reliable A. tumefaciens-mediated genetic transformation protocol was developed. Full article
(This article belongs to the Section Plant Biochemistry and Genetics)
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14 pages, 3399 KiB  
Article
Optimization of Agrobacterium Mediated Genetic Transformation in Paspalum scrobiculatum L. (Kodo Millet)
by Ritika Bhatt, Prem Prakash Asopa, Rohit Jain, Aditi Kothari-Chajer, Shanker Lal Kothari and Sumita Kachhwaha
Agronomy 2021, 11(6), 1104; https://doi.org/10.3390/agronomy11061104 - 28 May 2021
Cited by 18 | Viewed by 4711
Abstract
An efficient and reproducible protocol for Agrobacterium tumefaciens mediated genetic transformation was developed for kodo millet (Paspalum scrobiculatum L.) by optimizing various parameters. Agrobacterium strains EHA 105 and LBA 4404 harboring plasmids pCNL 56 and pCAMBIA 2300, respectively, provided the highest transformation [...] Read more.
An efficient and reproducible protocol for Agrobacterium tumefaciens mediated genetic transformation was developed for kodo millet (Paspalum scrobiculatum L.) by optimizing various parameters. Agrobacterium strains EHA 105 and LBA 4404 harboring plasmids pCNL 56 and pCAMBIA 2300, respectively, provided the highest transformation efficiency. Addition of acetosyringone (AS) in infection medium (200 µM-EHA 105, 250 µM-LBA 4404) and co-cultivation medium (50 µM) increased the transformation efficiency. Transient and stable expression of gus gene was confirmed with histochemical assay of infected embryos and leaves of transformed plants, respectively. The best GUS response was obtained by pretreatment of callus with an antinecrotic mixture (10 mg/L Cys + 5 mg/L Ag + 2.5 mg/L As) at infection time of 20 min followed by co-cultivation for 3 days (EHA 105) and 5 days (LBA 4404) in dark. Regenerated transgenic plants were obtained after 8 to 10 weeks of selection on callus induction medium (NAA 0.5 mg/L, BAP 1 mg/L) containing 50 mg/L Kan + 250 mg/L Cef and were rooted for 2 weeks on MS medium containing PAA (1 mg/L) and phytagel. The plantlets established in greenhouse showed normal growth. Therefore, the protocol developed in the present study can be used for development of improved varieties of kodo millet. Full article
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13 pages, 269 KiB  
Article
RNA Interference of 1-Aminocyclopropane-1-carboxylic Acid Oxidase (ACO1 and ACO2) Genes Expression Prolongs the Shelf Life of Eksotika (Carica papaya L.) Papaya Fruit
by Rogayah Sekeli, Janna Ong Abdullah, Parameswari Namasivayam, Pauziah Muda, Umi Kalsom Abu Bakar, Wee Chien Yeong and Vilasini Pillai
Molecules 2014, 19(6), 8350-8362; https://doi.org/10.3390/molecules19068350 - 19 Jun 2014
Cited by 18 | Viewed by 8120
Abstract
The purpose of this study was to evaluate the effectiveness of using RNA interference in down regulating the expression of 1-aminocyclopropane-1-carboxylic acid oxidase gene in Eksotika papaya. One-month old embryogenic calli were separately transformed with Agrobacterium strain LBA 4404 harbouring the three different [...] Read more.
The purpose of this study was to evaluate the effectiveness of using RNA interference in down regulating the expression of 1-aminocyclopropane-1-carboxylic acid oxidase gene in Eksotika papaya. One-month old embryogenic calli were separately transformed with Agrobacterium strain LBA 4404 harbouring the three different RNAi pOpOff2 constructs bearing the 1-aminocyclopropane-1-carboxylic acid oxidase gene. A total of 176 putative transformed lines were produced from 15,000 calli transformed, selected, then regenerated on medium supplemented with kanamycin. Integration and expression of the targeted gene in putatively transformed lines were verified by PCR and real-time RT-PCR. Confined field evaluation of a total of 31 putative transgenic lines planted showed a knockdown expression of the targeted ACO1 and ACO2 genes in 13 lines, which required more than 8 days to achieve the full yellow colour (Index 6). Fruits harvested from lines pRNAiACO2 L2-9 and pRNAiACO1 L2 exhibited about 20 and 14 days extended post-harvest shelf life to reach Index 6, respectively. The total soluble solids contents of the fruits ranged from 11 to 14° Brix, a range similar to fruits from non-transformed, wild type seed-derived plants. Full article
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