Optimization of Agrobacterium-mediated transient gene expression and endogenous gene silencing in Piper colubrinum Link. by vacuum infiltration

Mani, Tomson; Manjula, S.

doi:10.1007/s11240-010-9836-z

Cited by 12 publications

(5 citation statements)

References 27 publications

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“…However, the transformation efficiency and regeneration efficiency are too low and time-consuming to characterize multiple genes. In recent years, the techniques of gene silencing and transient expression based on agroinfiltration have become important methods for gene function research, because of their short time consumption and high throughput [ 47 , 48 , 49 ]. In previous study, ScCAT1 has been shown to play a positive role in HR and immune responses by transient overexpression in N. benthamiana leaves [ 23 ].…”

Section: Discussionmentioning

confidence: 99%

The Role of Sugarcane Catalase Gene ScCAT2 in the Defense Response to Pathogen Challenge and Adversity Stress

Sun

Liu

Wang

et al. 2018

IJMS

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Catalases, which consist of multiple structural isoforms, catalyze the decomposition of hydrogen peroxide in cells to prevent membrane lipid peroxidation. In this study, a group II catalase gene ScCAT2 (GenBank Accession No. KF528830) was isolated from sugarcane genotype Yacheng05-179. ScCAT2 encoded a predicted protein of 493 amino acid residues, including a catalase active site signature (FARERIPERVVHARGAS) and a heme-ligand signature (RVFAYADTQ). Subcellular localization experiments showed that the ScCAT2 protein was distributed in the cytoplasm, plasma membrane, and nucleus of Nicotiana benthamiana epidermal cells. Quantitative real-time polymerase chain reaction (qRT-PCR) analysis indicated that the ScCAT2 gene was ubiquitously expressed in sugarcane tissues, with expression levels from high to low in stem skin, stem pith, roots, buds, and leaves. ScCAT2 mRNA expression was upregulated after treatment with abscisic acid (ABA), sodium chloride (NaCl), polyethylene glycol (PEG), and 4 °C low temperature, but downregulated by salicylic acid (SA), methyl jasmonate (MeJA), and copper chloride (CuCl2). Moreover, tolerance of Escherichia coli Rosetta cells carrying pET-32a-ScCAT2 was enhanced by NaCl stress, but not by CuCl2 stress. Sporisorium scitamineum infection of 10 different sugarcane genotypes showed that except for YZ03-258, FN40, and FN39, ScCAT2 transcript abundance in four smut-resistant cultivars (Yacheng05-179, YZ01-1413, YT96-86, and LC05-136) significantly increased at the early stage (1 day post-inoculation), and was decreased or did not change in the two smut-medium-susceptibility cultivars (ROC22 and GT02-467), and one smut-susceptible cultivar (YZ03-103) from 0 to 3 dpi. Meanwhile, the N. benthamiana leaves that transiently overexpressed ScCAT2 exhibited less severe disease symptoms, more intense 3,3′-diaminobenzidine (DAB) staining, and higher expression levels of tobacco immune-related marker genes than the control after inoculation with tobacco pathogen Ralstonia solanacearum or Fusarium solani var. coeruleum. These results indicate that ScCAT2 plays a positive role in immune responses during plant–pathogen interactions, as well as in salt, drought, and cold stresses.

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Section: Discussionmentioning

confidence: 99%

The Role of Sugarcane Catalase Gene ScCAT2 in the Defense Response to Pathogen Challenge and Adversity Stress

Sun

Liu

Wang

et al. 2018

IJMS

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“…It is used to indicate transient and stable transformations (Serres et al 1992), and to estimate the relative contribution and strength of promoter elements in the transcriptional activation. Most recently, GUS histochemical assay was utilized in the production of markerfree Petunia hybrida and tomato (Khan et al , 2011, and in the production of a more efficient Agrobacterium tumefaciens-mediated transformation protocol in melon (Ntui et al 2010), in Lilium (Azadi et al 2010), in Petunia hybrida (Thirukkumaran et al 2009), in pineapple (Gangopadhyay et al 2009), in Piper colubrinum (Mani and Manjula 2011), in sesame (Yadav et al 2010), in…”

Section: Discussionmentioning

confidence: 99%

Characterization of inhibitor(s) of β-glucuronidase enzyme activity in GUS-transgenic wheat

Ramadan

Eissa

El-Domyati

et al. 2011

Plant Cell Tiss Organ Cult

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The uidA gene, encoding for b-glucuronidase (GUS), is the most frequently used reporter gene in plants.As a reporter enzyme, GUS can be assayed both qualitatively and quantitatively. In wheat, there are numerous reports of failure in detecting GUS enzyme activity in tissues of transgenic plants, while other reports have suggested presence of b-glucuronidase inhibitor(s) in wheat tissues. In the present study, we show that the b-glucuronidase enzyme activity is not only tissue-specific but also genotype-dependent. Our data demonstrate that the glucuronic acid could be the candidate inhibitor for b-glucuronidase enzyme activity in wheat leaves and roots. It should be noted that the assays to detect b-glucuronidase enzyme activity in wheat should be interpreted carefully. Based on the data of our present study, we recommend studying the chemical pathways, the unintended effects and the possible loss-of-function of any candidate transgene prior to transformation experiments.

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“…Genes are successfully transiently expressed in various tissues of different plants via syringe infiltration, such as Arabidopsis , soybean leaves ( Kim et al, 2009 ; King et al, 2015 ), and melon fruit ( Han et al, 2015 ). Vacuum infiltration has also been utilized for transient transformation in different plant tissues, including tobacco, black pepper leaves ( Mani and Manjula, 2011 ; Fujiuchi et al, 2016 ), and lisianthus pollen ( Sung, 2004 ). Although specialized equipment is required, vacuum infiltration exhibits improved transformation potential or yield ( Zhao et al, 2017 ).…”

Section: Introductionmentioning

confidence: 99%

An Easy and Rapid Transformation Protocol for Transient Expression in Cotton Fiber

et al. 2022

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Cotton fiber is the most important natural textile material in the world. Identification and functional characterization of genes regulating fiber development are fundamental for improving fiber quality and yield. However, stable cotton transformation is time-consuming, low in efficiency, and technically complex. Moreover, heterologous systems, such as Arabidopsis and tobacco, did not always work to elucidate the function of cotton fiber specifically expressed genes or their promoters. For these reasons, constructing a rapid transformation system using cotton fibers is necessary to study fiber’s specifically expressed genes. In this study, we developed an easy and rapid Agrobacterium-mediated method for the transient transformation of genes and promoters in cotton fibers. First, we found that exogenous genes could be expressed in cotton fibers via using β-glucuronidase (GUS) and green fluorescence protein (GFP) as reporters. Second, parameters affecting transformation efficiency, including LBA4404 Agrobacterium strain, 3 h infection time, and 2-day incubation time, were determined. Third, four different cotton genes that are specifically expressed in fibers were transiently transformed in cotton fibers, and the transcripts of these genes were detected ten to thousand times increase over the control. Fourth, GUS staining and activity analysis demonstrated that the activity profiles of GhMYB212 and GhFSN1 promoters in transformed fibers are similar to their native activity in developmental fibers. Furthermore, the transient transformation method was confirmed to be suitable for subcellular localization studies. In summary, the presented Agrobacterium-mediated transient transformation method is a fast, simple, and effective system for promoter characterization and protein expression in cotton fibers.

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Optimization of Agrobacterium-mediated transient gene expression and endogenous gene silencing in Piper colubrinum Link. by vacuum infiltration

Cited by 12 publications

References 27 publications

The Role of Sugarcane Catalase Gene ScCAT2 in the Defense Response to Pathogen Challenge and Adversity Stress

The Role of Sugarcane Catalase Gene ScCAT2 in the Defense Response to Pathogen Challenge and Adversity Stress

Characterization of inhibitor(s) of β-glucuronidase enzyme activity in GUS-transgenic wheat

An Easy and Rapid Transformation Protocol for Transient Expression in Cotton Fiber

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