The Effect of Cucumber mosaic virus 2b Protein to Transient Expression and Transgene Silencing Mediated by Agro-infiltration

Choi, Min Sue; Yoon, In Sun; Rhee, Yong; Choi, Seung‐Kook; Lim, Sun Hyung; Won, So Youn; Lee, Yeon Hee; Choi, Hong Soo; Lee, Suk Chan; Kim, Kook Hyung; Lomonossoff, George P.; Sohn, Seong Han

doi:10.5423/ppj.2008.24.3.296

Cited by 11 publications

(17 citation statements)

References 18 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…2A). The final concentrations of each Agrobacterium in the mixture was adjusted to OD 600nm = 0.2 to minimize possible stress on leaf and maximize GFP expression as reported (Choi et al, 2008). The strength of GFP expression was found in the order of GFP+NS3 > GFP+2b > GFP alone; GFP expression remained until 15 dpi in the presence of NS3 (Fig.…”

Section: Resultsmentioning

confidence: 99%

“…Agroinfiltration was carried out by using a previously reported protocol (Choi et al, 2008). Wildtype N. benthamiana was used to examine the enhancing effect of NS3 on transient expression, and two lines of GFP-transgenic N.benthamiana were used for analyzing the suppressive effect on GFP-transgene co-silencing.…”

Section: Methodsmentioning

confidence: 99%

“…In addition, pGreen0229-35S:GFP (35S-GFP) was used as a marker vector for transient expression and a initiator of GFPtransgene co-silencing. These two vectors were previously constructed by Choi et al (2008). GFP-transgenic plants were produced using leaf-disc transformation with A. tumefaciens harboring 35S-GFP (Sohn et al, 2011).…”

Section: Methodsmentioning

confidence: 99%

“…For western blotting, total soluble protein (TSP) was extracted from the agroinfiltrated leaves as described by Choi et al (2008). Protein extracts (5 µg) were separated by 12.5% SDS-PAGE, and subsequently blotted onto polyvinylidene fluoride (PVDF) membranes (GE Healthcare).…”

Section: Methodsmentioning

confidence: 99%

“…The NS3 gene has been identified as a gene silencing suppressor not only in plants and cultured insect cell (Hemmes et 2007), but also in mammalian cells (Schnettler et al, 2008), through the binding and sequestering of siRNAs (Xiong et al, 2009). Here, we isolated the NS3 gene from a Korean isolate and reported its effect on both transient gene expression and transgene co-silencing in comparison with CMV 2b (Choi et al, 2008). As a result, we have demonstrated that the NS3 gene can be one of the most potent enhancers for transient expression and suppressors of gene silencing.…”

mentioning

confidence: 90%

See 4 more Smart Citations

Effect of Rice stripe virus NS3 on Transient Gene Expression and Transgene Co-Silencing

Sohn¹,

Huh²,

Kim³

et al. 2011

The Plant Pathology Journal

Self Cite

View full text Add to dashboard Cite

Nonstructural protein 3 (NS3) encoded by RNA3 of Rice stripe virus (RSV), known to be a suppressor of gene silencing, was cloned and sequenced. The cloned NS3 gene is composed of 636 nucleotides encoding 211 deduced amino acids, and showed a high degree of similarity with the equivalent genes isolated from Korea, Japan and China. The NS3 gene promoted the enhancement of transient gene expression and suppressed transgene co-silencing. In the transient GFP expression via agroinfiltration, GFP expression was dramatically enhanced in terms of both protein yield and expression period in the presence of NS3. The highest accumulation of GFP protein reached to 6.8% of total soluble proteins, which corresponded to a two-fold increase compared to that obtained in the absence of NS3. In addition, NS3 significantly suppressed the initiation of GFP co-silencing induced by the additive GFP infiltration in GFP-transgenic Nicotiana benthamiana. The NS3 gene was also found to be a stronger suppressor than Cucumber mosaic virus 2b. These observations are believed to be derived from the strong suppressive effect of NS3 on gene silencing, and indicate that NS3 could be used as an effective enhancer for the rapid production of foreign proteins in plants.

show abstract

Section: Resultsmentioning

confidence: 99%

Section: Methodsmentioning

confidence: 99%

Section: Methodsmentioning

confidence: 99%

Section: Methodsmentioning

confidence: 99%

mentioning

confidence: 90%

See 3 more Smart Citations

Effect of Rice stripe virus NS3 on Transient Gene Expression and Transgene Co-Silencing

Sohn¹,

Huh²,

Kim³

et al. 2011

The Plant Pathology Journal

Self Cite

View full text Add to dashboard Cite

show abstract

Regulation and function of the pepper pectin methylesterase inhibitor (CaPMEI1) gene promoter in defense and ethylene and methyl jasmonate signaling in plants

Choi

Hong

et al. 2009

Planta

View full text Add to dashboard Cite

Analysis of the promoters of defense-related genes is valuable for determining stress signaling and transcriptional activation during pathogen infection. Here, we have isolated and functionally characterized the promoter region of the pepper (Capsicum annuum) pectin methylesterase inhibitor 1 (CaPMEI1) gene in transiently transformed tobacco plants and stably transformed Arabidopsis plants. Among four 5' deletion constructs analyzed, the -958-bp CaPMEI1 promoter induced a high level of GUS reporter activity in tobacco leaf tissue, driven by pathogen infection as well as by ethylene and methyl jasmonate (MeJA) treatment. The 204-bp region from -958 bp to -754 bp of the CaPMEI1 promoter is responsible for the stress-responsive expression. In addition, the pepper transcription factor CARAV1 activated the CaPMEI1 promoter in tobacco leaves, whereas the transcription factor CAbZIP1 did not. In the transgenic Arabidopsis plants, the -958 bp CaPMEI1 promoter was functionally regulated by developmental cues, bacterial and oomycete pathogen infections, and treatment with ethylene and MeJA. Histochemical GUS staining analyses of Arabidopsis tissues revealed that the CaPMEI1 promoter was mainly activated in leaf veins in response to various biotic and abiotic stimuli. Together, these results suggest that CaPMEI1 promoter activation may be a critical molecular event for host defense response and ethylene- and MeJA-mediated CaPMEI1 gene expression.

show abstract

The epigenetic phenotypes in transgenic Nicotiana benthamiana for CaMV 35S-GFP are mediated by spontaneous transgene silencing

Sohn

Choi²,

Kim

et al. 2011

Plant Biotechnol Rep

Self Cite

View full text Add to dashboard Cite

Diverse epigenetic phenotypes are frequently found during research on transgenic plants. To understand the factors underlying such diversity, hundreds of independent 35S-GFP transgenic N. benthamiana plants were analyzed. The diverse GFP-expression phenotypes of the transgenic plants were classified into three major types based on the GFP expression patterns and their response to 35S-GFP agroinfiltration: steady-green, silenced and non-uniform phenotype. The non-uniform phenotype was further sub-divided into five minor phenotypes: variegated, reddropped, on-silencing, partitioned and misty, according to the distribution of GFP expression on the leaves. Many of transgenic plants continuously generated diverse phenotypes over several generations despite the transgene identity. Such epigenetic GFP phenotyping was found to be the result of spontaneous transgene silencing mediated by either or both of post-transcriptional gene silencing (PTGS) and transcriptional gene silencing (TGS). This finding was verified by the detection of 21-and 24-nt small interfering RNA (siRNA) molecules, and DNA methylation in the transgenic plants that showed repeated epigenetic variation. Agroinfiltration demonstrated that irregular distribution of GFP on a leaf was the result of erratic transgene silencing, and the technique also proved to be a rapid and effective method for selecting fully silenced plants within 3 days. Furthermore, two novel phenotypes described are potential materials for in-depth investigations into the genes and mechanisms responsible for spontaneous transgene silencing.

show abstract

The Effect of Cucumber mosaic virus 2b Protein to Transient Expression and Transgene Silencing Mediated by Agro-infiltration

Cited by 11 publications

References 18 publications

Effect of Rice stripe virus NS3 on Transient Gene Expression and Transgene Co-Silencing

Effect of Rice stripe virus NS3 on Transient Gene Expression and Transgene Co-Silencing

Regulation and function of the pepper pectin methylesterase inhibitor (CaPMEI1) gene promoter in defense and ethylene and methyl jasmonate signaling in plants

The epigenetic phenotypes in transgenic Nicotiana benthamiana for CaMV 35S-GFP are mediated by spontaneous transgene silencing

Contact Info

Product

Resources

About