An improved method for Agrobacterium-mediated genetic transformation from cotyledon explants of Brassica juncea

Bhuiyan, Mohammed Shafi Ullah; Min, Sung Ran; Jeong, Won Joong; Sultana, Sayeda; Choi, Kwan Sam; Lim, Yong Pyo; Song, Won Yong; Lee, Youngsook; Liu, Jang R.

doi:10.5511/plantbiotechnology.10.0921a

Cited by 23 publications

(13 citation statements)

References 23 publications

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“…Approximately 8.7% survival was obtained when the hygromycin concentration was increased to 4 mg/L. These results are not consistent with those of Bhuiyan et al (2011), who reported that 15 mg/L hygromycin was optimal for the primary selection of transgenic shoots. However, differences exist among various cultivars of B. napus.…”

Section: Effect Of Hygromycin On Survival Of Hypocotyl Segmentscontrasting

confidence: 50%

Improved Agrobacterium-mediated transformation and high efficiency of root formation from hypocotyl meristem of spring Brassica napus ‘Precocity’ cultivar

Liu

Lang

et al. 2015

Genet. Mol. Res.

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ABSTRACT. Rape seed (Brassica napus L.) is one of the most important oil seed crops in the world. Genetic manipulation of rapeseed requires a suitable tissue culture system and an efficient method for plant regeneration, as well as an efficient transformation procedure. However, development of transgenic B. napus has been problematic, and current studies are limited to cultivated varieties. In this study, we report a protocol for regeneration of transgenic rape after Agrobacterium-mediated transformation of hypocotyls from the spring B. napus 'Precocity' cultivar. We analyzed the effects of plant growth regulators in the medium on regeneration. Additionally, factors affecting the transformation efficiency, including seedling age, Agrobacterium concentration, infection time, and co-cultivation time, were assessed by monitoring GUS expression. Results from these experiments revealed that transformation was optimized when the meristematic parts of the 16841 Transformation and root formation of Brassica napus ©FUNPEC-RP www.funpecrp.com.br Genetics and Molecular Research 14 (4): 16840-16855 (2015) hypocotyls were taken from 8 day-old seedlings, cultured on Murashinge and Skoog basal media containing 0.1 mg/L 1-naphthaleneacetic acid and 2.5 mg/L 6-benzylaminopurine, and incubated in Agrobacterium suspension (OD 600 = 0.5) for 3 to 5 min, followed by 2 days of cocultivation. Integration of T-DNA into the plant genome was confirmed by polymerase chain reaction (PCR), b-glucuronidase histochemical staining, and quantitative real-time PCR. The protocols developed for regeneration, transformation, and rooting described in this study could help to accelerate the development of transgenic spring rape varieties with novel features.

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Section: Effect Of Hygromycin On Survival Of Hypocotyl Segmentscontrasting

confidence: 50%

Improved Agrobacterium-mediated transformation and high efficiency of root formation from hypocotyl meristem of spring Brassica napus ‘Precocity’ cultivar

Liu

Lang

et al. 2015

Genet. Mol. Res.

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“…Rai-5 seeds were surface sterilized by soaking for 10 min in a 10% bleach solution, thoroughly rinsed with sterile distilled water, and germinated on MS medium (Murashige and Skoog 1962) containing 20 g l -1 sucrose and 6 g l -1 Phytoagar (Duchefa Biochemie, Haarlem, The Netherlands). Cotyledon explants from 4-day-old seedlings were used for transformation and hygromycin-resistant shoots were regenerated as previously described by Bhuiyan et al (2011b). Putative transgenic plants were grown in potting soil and transferred to a greenhouse to produce T 1 seeds.…”

Section: Plasmid Constructs Plant Transformation and Plant Regeneramentioning

confidence: 99%

Overexpression of AtATM3 in Brassica juncea confers enhanced heavy metal tolerance and accumulation

Bhuiyan

Min

Jeong

et al. 2011

Plant Cell Tiss Organ Cult

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AtATM3, a member of the ATP-binding cassette transporter family, is localized at the mitochondrial membrane of Arabidopsis thaliana and is involved in the biogenesis of Fe-S clusters and iron homeostasis in plants.Through Agrobacterium-mediated genetic transformation, the AtATM3 gene driven by the cauliflower mosaic virus 35S promoter (CaMV35S) was introduced into Brassica juncea (Indian mustard), a plant species suitable for phytoremediation, with the aim of improving heavy metal tolerance and accumulation in plants. The presence of the AtATM3 gene in transgenic plants was confirmed by polymerase chain reaction (PCR). Reverse transcriptase-PCR analysis confirmed AtATM3 expression in transgenic plants, but the level of AtATM3 expression varied between lines. AtATM3 overexpression in B. juncea conferred enhanced tolerance to cadmium [Cd(II)] and lead [Pb (II)] stresses. Importantly, the shoot tissues of transgenic seedlings contained about 1.5-to 2.5-fold higher Cd(II) and Pb(II) levels than wild type (WT) seedlings, demonstrating significantly-increased accumulation of both Cd(II) and Pb(II) in transgenic plants. The enhanced capacity of heavy metal tolerance and accumulation by AtATM3 transgenic plants was attributed to higher BjGSHII (B. juncea glutathione synthetase II) and BjPCS1 (phytochelatin synthase 1) expression levels induced by AtATM3 overexpression. In addition, AtATM3 overexpression regulated the expression of several metal transporters in the transgenic B. juncea plants under heavy metal stress conditions. Therefore, AtATM3 transgenic plants are more tolerant of and can accumulate more heavy metals to enhance phytoremediation of contaminated soils.

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“…We are currently using the transgenic the BRRI dhan29 lines for heavy metal phytoremediation purposes. A researcher has got same results in Brassica juncea (Bhuiyan et al, 2011).…”

Section: Segregation Of T1 Progenymentioning

confidence: 54%

Development of Transgenic Rice (Oryza sativa L.) Plant Using Cadmium Tolerance Gene (YCFI) through Agrobacterium Mediated Transformation for Phytoremediation

Islam¹,

Khalekuzzaman²

2015

Asian J. of Agricultural Research

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Cadmium has adverse effects on plant development and ultimately reduces production. The investigation was undertaken for successful gene transfer and phytoremediation through an efficient Agrobacterium-mediated genetic transformation method using cadmium tolerance gene (YCF1). Embryogenic calli induced after 20 days of highly regenerating rice cultivar BRRI dhan29 and Agrobacterium strain GV 3101 was transformed with binary vector pCAMBIA 1303-YCF1 which contained the hygromycin phosphotransferase (HPT) gene as a selectable marker and the yeast cadmium factor 1 (YCF1) gene, were used for genetic transformation in the experiment. The transformed colonies were selected on 15 mg LG 1 hygromycin and 50 mg LG 1 rifampicin to select hygromycin resistant shoots. Hygromycin-resistant shoots were subsequently rooted on root induction medium. Rooted plantlets were transferred to pot-soil, hardened and grown in a greenhouse until maturity and stable integration, expression of YCF1 gene was also confirmed by using PCR analysis. The maximum transformation efficiency of 22% was obtained using 500 mg LG 1 cefotaxime as a bacteriostatic agent to inhibit growth of Agrobacterium and 100 mM acetosyringone in co-cultivation medium. Southern blot analysis was performed to confirm that transgenes (HPT and YCF1) were stably integrated into the plant genome. All transgenic plants showed single-copy of transgene integration in the host genome. This transgenic rice plant will uptake cadmium from soil and will protect rice grain from cadmium and store into cell vacuoles of rice plants. As a result soil will be free from cadmium through phytoremediation process.

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An improved method for Agrobacterium-mediated genetic transformation from cotyledon explants of Brassica juncea

Cited by 23 publications

References 23 publications

Improved Agrobacterium-mediated transformation and high efficiency of root formation from hypocotyl meristem of spring Brassica napus ‘Precocity’ cultivar

Improved Agrobacterium-mediated transformation and high efficiency of root formation from hypocotyl meristem of spring Brassica napus ‘Precocity’ cultivar

Overexpression of AtATM3 in Brassica juncea confers enhanced heavy metal tolerance and accumulation

Development of Transgenic Rice (Oryza sativa L.) Plant Using Cadmium Tolerance Gene (YCFI) through Agrobacterium Mediated Transformation for Phytoremediation

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