Enhancement of Agrobacterium-mediated transformation efficiency of model plant using quorum sensing molecule, N-3-oxo-octanoyl-l-homoserine-lactone

Chen, Chin-Fong; Chan, Kok‐Gan; Tan, Boon Chin

doi:10.1007/s11240-015-0718-2

Cited by 3 publications

(3 citation statements)

References 36 publications

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“…Despite the successes achieved in the past few years, genetic engineering of plants is still in its infancy. This may be due to the limited understanding of most secondary metabolite biosynthesis pathways (Weathers et al 2010), including incomplete pathways and key regulatory mechanisms controlling the pathway fluxes (Oksman-Caldentey & Inzé 2004), as well as the low success rates in introducing multiple genes into plants (Chen et al 2015). …”

Section: Introductionmentioning

confidence: 99%

Stable integration of mgfp5 transgenes following Agrobacterium-mediated transformation in Boesenbergia rotunda cell suspension culture

Wong

Zoolkefli

Karim

et al. 2015

Frontiers in Life Science

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Boesenbergia rotunda, a herb in the ginger family, contains numerous beneficial compounds, such as flavonoids, flavones and cyclohexenyl chalcone derivatives, that have great potential for pharmaceutical applications. However, the low concentration of the bioactive compounds limits their commercial application. In this study, a simple and reliable Agrobacterium-mediated transformation protocol for B. rotunda cell suspension culture was successfully developed. The minimal inhibitory concentration and natural tolerance of the selective agent, hygromycin, against the cells were 20 mg l −l and 30 mg l −l in liquid media and solid media, respectively. The highest number of transformed regenerants (18 ± 0.00 per ml settled cell volume) was recorded when cells were infected with Agrobacterium tumefaciens harbouring pCAMBIA1304 for 10 min and co-cultivated for 2 days. Prolonged infection time ( > 10 min) and co-cultivation period ( > 2 days), however, did not increase the transformation efficiencies. The results clearly show that infection and co-cultivation periods strongly influenced the transformation efficiency in ginger. The transformed cells were recovered and showed green fluorescent signals under ultraviolet excitation. An intense blue colour was observed in the transformed cells after β-glucuronidase (GUS) histochemical staining, further confirming the functionality of the GUS enzymes in the regenerants. Polymerase chain reaction analysis of 3-, 6-, 9-and 12-month-old transformed cells confirmed that the protocol enabled stable integration of the mgfp5 gene. Moreover, the comparatively high number of transformed regenerants in this study made it possible to generate a large number of transgenic cells in a short period, which would be useful for high-throughput functional screening of enzymes involved in the biosynthetic pathways of bioactive compounds.

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Section: Introductionmentioning

confidence: 99%

Stable integration of mgfp5 transgenes following Agrobacterium-mediated transformation in Boesenbergia rotunda cell suspension culture

Wong

Zoolkefli

Karim

et al. 2015

Frontiers in Life Science

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“…20). This difference probably arises because the A. tumefaciens relaxase has evolved to facilitate nuclear entry and integration into the genome, whereas the Bartonella relaxase has not.…”

Section: Bacterial Secretion Systemsmentioning

confidence: 99%

“…However, these efficiencies are still low compared with some A. tumefaciens protocols, which can be as high as 90% (Ref. 20 ). This difference probably arises because the A. tumefaciens relaxase has evolved to facilitate nuclear entry and integration into the genome, whereas the Bartonella relaxase has not.…”

Section: Bacterial Secretion Systemsmentioning

confidence: 99%

Intracellular delivery of biologic therapeutics by bacterial secretion systems

Walker

Stan

Polizzi

2017

Expert Rev. Mol. Med.

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Biologics are a promising new class of drugs based on complex macromolecules such as proteins and nucleic acids. However, delivery of these macromolecules into the cytoplasm of target cells remains a significant challenge. Here we present one potential solution: bacterial nanomachines that have evolved over millions of years to efficiently deliver proteins and nucleic acids across cell membranes and between cells. In this review, we provide a brief overview of the different bacterial systems capable of direct delivery into the eukaryotic cytoplasm and the medical applications for which they are being investigated, along with a perspective on the future directions of this exciting field.

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Expression of Talaromyces thermophilus lipase gene in Trichoderma reesei by homologous recombination at the cbh1 locus

Zhang

Xia

2017

Journal of Industrial Microbiology and Biotechnology

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CBH1 (cellobiohydrolase) comprises the majority of secreted proteins by Trichoderma reesei. For expression of Talaromyces thermophilus lipase gene in T. reesei, a self-designed CBH1 promoter was applied to drive the lipase gene expression cassette which was bracketed by flanking sequences of cbh1 gene for homologous recombination. Protoplast and Agrobacterium-mediated plasmid transformations were performed and compared, resultantly, transformation mediated by Agrobacterium was overall proved to be more efficient. Stable integration of lipase gene into chromosomal DNA of T. reesei transformants was verified by PCR. After shaking flask fermentation, lipase activity of transformant reached 375 IU mL, whereas no cellobiohydrolase activity was detected. SDS-PAGE analysis further showed an obvious protein band about 39 kDa and no CBH1 band in fermentation broth, implying lipase gene was successfully extracellularly expressed in T. reesei via homologous recombination at cbh1 locus. This study herein would benefit genetic engineering of filamentous fungi and industrial application of thermo-alkaline lipase like in paper making and detergents addition.

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Enhancement of Agrobacterium-mediated transformation efficiency of model plant using quorum sensing molecule, N-3-oxo-octanoyl-l-homoserine-lactone

Cited by 3 publications

References 36 publications

Stable integration of mgfp5 transgenes following Agrobacterium-mediated transformation in Boesenbergia rotunda cell suspension culture

Stable integration of mgfp5 transgenes following Agrobacterium-mediated transformation in Boesenbergia rotunda cell suspension culture

Intracellular delivery of biologic therapeutics by bacterial secretion systems

Expression of Talaromyces thermophilus lipase gene in Trichoderma reesei by homologous recombination at the cbh1 locus

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