Arabidopsis thaliana small subunit leader and transit peptide enhance the expression ofBacillus thuringiensis proteins in transgenic plants

Wong, Edith Y.; Hironaka, Catherine M.; Fischhoff, David A.

doi:10.1007/bf00029151

Cited by 102 publications

(42 citation statements)

References 25 publications

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“…Specificity of Construct-Specific Methods for NL, NLP and NLY In NL, the e-p-35Ssequence 17) is used to regulate the expression of the cryIIIA gene, whereas the riblose-1,5-bisphosphate carboxylase small subunit ats1A promoter (PArab-SSU1A) 18) is used to control the cryIIIA gene expression in NLP and NLY. Therefore, the construct-specific primer pair, NL 01-5Ј/NL 01-3Ј, was designed in the junction between e-p-35S and cryIIIA to specifically detect NL.…”

Section: Specificity Of Screening Methods For the Detection Of Gm Potamentioning

confidence: 99%

New Qualitative Detection Methods of Genetically Modified Potatoes

Watanabe

Kuribara

Mishima

et al. 2004

Biological & Pharmaceutical Bulletin

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Many kinds of genetically modified (GM) crops, which include GM soy, maize, rapeseed, cotton and potato, have already been developed and the cultivated acreage of these crops has continued to grow year by year. It was reported that the global area of GM crops for 2003 was 67.7 million hectares with a growth rate of 15% compared to that in 2002. This growth is estimated to rapidly increase, since the planting has been spread all over the world in addition to the nations such as United States (U.S.) and Canada. 1) On the other hand, public concern has been raised in terms of food safety and environmental effects of the GM crops. Especially, consumers are concerned about the negative effects of GM food on their health by their consumption and scientific information has been strongly required.2) Therefore, many governments have now been considering regulations for the use and implementing a labeling system for GM crops as food and feed. Thus, new labeling systems have been introduced for GM foods in the European Union (EU), Australia, Korea, Japan and other countries.The commercialization of fifty-five lines of safety-assessed GM crops including soy, maize, potato, rapeseed, cotton and sugar beet, have already been approved by the Ministry of Health, Labour and Welfare (HMLW) in Japan. To monitor the labeling system, it is necessary to develop reliable and practical methods for the detection and identification of GM crops. The polymerase chain reaction (PCR) is one of the widely used systems for the quantitative or qualitative detection of GM crops and we also have previously reported PCR methods for the detection of GM soy, maize, papaya and potatoes. 3-10)The tetraploid cultivated potato (Solanum tuberosum) is one of the world's four major crops and an important feedstuff, but it is easily infected by many kinds of pests and pathogens.11) Therefore, molecular biology techniques has been attempted to improve the potato varieties which ended with the breeding of GM potatoes commercialized by the Monsanto Co. We have developed polymerase chain reaction (PCR) methods for the qualitative detection of the GM potatoes for the screening and the identification of NL, NLP and NLY. The gene encoding uridine diphosphate (UDP)-glucose pyrophosphorylase (UGPase) was used as a taxon specific gene. We designed the primer pair to detect the cryIIIA genes as a screening method for GM potatoes because the gene should be inserted in all 8 lines of the GM potatoes. For identification of NL, NLP and NLY, we further designed three specific primer pairs for the different recombinant DNAs (r-DNA) specifically introduced into NL, NLP, or NLY. In addition, to identify the 3 lines of NLY that have been introduced with the same r-DNA, the three line-specific primer pairs for the border sequence between the r-DNA and genomic DNA of NLY 3 lines were designed. Six lines of GM potato used as the test material were specifically identified using the each primer pair under the same PCR condition. The detection limits of all the GM potatoes should b...

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Section: Specificity Of Screening Methods For the Detection Of Gm Potamentioning

confidence: 99%

New Qualitative Detection Methods of Genetically Modified Potatoes

Watanabe

Kuribara

Mishima

et al. 2004

Biological & Pharmaceutical Bulletin

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“…In the accompanying paper (Diehn et al, 1998) One approach used in the design of synthetic B.t.-toxin genes has been to modify A/T-rich sequences resembling known sequences involved in RNA processing or stability by converting codons in the target sequences to synonymous codons typically used in plant genes (Perlak et al, 1990(Perlak et al, , 1991(Perlak et al, , 1993Sutton et al, 1992;van der Salm et al, 1994). It has been suggested that the abundance of rare codons in B.t.-toxin genes could cause inefficient translation, resulting in poor expression in plants (Vaeck et al, 1987;Murray et al, 1991;Perlak et al, 1991;Wong et al, 1992), although this remains unproven. Based on this latter hypothesis, however, other synthetic B.t.-toxin genes giving high expression in plants have been constructed that primarily incorporate sequence modifications intended to convert rare codons to plant-preferred codons (Adang et al, 1993;Fujimoto et al, 1993;Koziel et al, 1993).…”

Section: Discussionmentioning

confidence: 99%

Direct Evidence for Rapid Degradation ofBacillus thuringiensis Toxin mRNA as a Cause of Poor Expression in Plants1

Rocher

Vargo-Gogola²,

Diehn

et al. 1998

Plant Physiology

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It is well established that the expression of Bacillus thuringiensis (B.t.) toxin genes in higher plants is severely limited at the mRNA level, but the cause remains controversial. Elucidating whether mRNA accumulation is limited transcriptionally or posttranscriptionally could contribute to effective gene design as well as provide insights about endogenous plant gene-expression mechanisms. To resolve this controversy, we compared the expression of an A/Urich wild-type cryIA(c) gene and a G/C-rich synthetic cryIA(c) B.t.-toxin gene under the control of identical 5 and 3 flanking sequences. Transcriptional activities of the genes were equal as determined by nuclear run-on transcription assays. In contrast, mRNA half-life measurements demonstrated directly that the wildtype transcript was markedly less stable than that encoded by the synthetic gene. Sequences that limit mRNA accumulation were located at more than one site within the coding region, and some appeared to be recognized in Arabidopsis but not in tobacco (Nicotiana tabacum). These results support previous observations that some A/U-rich sequences can contribute to mRNA instability in plants. Our studies further indicate that some of these sequences may be differentially recognized in tobacco cells and Arabidopsis.Gene transfer into plants has become a relatively simple and routine process in many species. However, successful integration of a transgene into the plant genome does not automatically result in expression. For example, introduction of an additional copy of an endogenous gene can trigger cosuppression mechanisms, resulting in silencing of both the introduced and the endogenous genes (for review, see Meyer and Saedler, 1996). In the case of foreign genes transferred into plants, it is not uncommon to find that the introduced gene is incompatible with plant geneexpression mechanisms. Bacillus thuringiensis (B.t.)-toxin genes are perhaps the most widely known example of foreign genes for which it has been difficult to obtain useful levels of expression in transgenic plants (Diehn et al., 1996). Although a variety of mechanisms controlling gene expression have been blamed for poor B.t.-toxin expression, the exact cause remains controversial because of the limited amount of data gathered to address the problem. This is in part due to the inherent difficulties associated with studying a gene expressed at very low levels. It is also due to the fact that, for practical applications in agriculture, this problem can be circumvented by the brute-force approach of constructing highly modified synthetic versions of B.t.-toxin genes that yield high levels of expression. Nevertheless, elucidating the mechanisms that limit B.t.-toxin expression could make the design of new genes more efficient and provide insight about steps in gene expression that are relevant to endogenous plant genes.A widely observed characteristic of B.t.-toxin gene expression in plants has been that little or no mRNA accumulates, even when transcription is under the control of strong pr...

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“…HD73, under control of the Arabidopsis thaliana ribulose-l,5-bisphosphate carboxylase (RuBisCO) small subunit atsiA promoter with and without its associated transit peptide was analysed in transgenic tobacco plants. The examination of leaf tissue revealed that the atslA promoter with its transit peptide sequence fused to the truncated cryiA(c) protein provided a 10-fold to 20-fold increase in cryiA(c) mRNA and protein levels compared to gene constructs in which the cauliflower mosaic virus 35S promoter with a duplication of the enhancer region (CaMV-En35S) was used to express the same cryiA(c) gene (Wong et al 1992). Fujimoto et al (1993) Gittins et al (2000) studied the ability of heterologous RuBisCO small-subunit gene promoters, Rbss3CP (0.8 kbp) from tomato (Lycopersicon esculentum Mill.)…”

Section: Rubisco Small Subunit Promoter As a Green Tissue-specific Prmentioning

confidence: 99%

Endeavours of RuBisCO small subunit promoter as a tool of green tissue specific expression

Bakhsh¹,

Husnain²

2012

CZECH J GENET PLANT

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Transcriptomics has the potential to rapidly increase our knowledge of spatial and temporal gene expression and contributes to the characterization of new promoters for research and development. The successful application of transgenic technology has been further strengthened by the availability of a broad spectrum of promoters having the ability to regulate the temporal and spatial expression patterns of the transgene. A variety of promoters is necessary at all levels of genetic engineering in plants, from basic research discoveries, to development of economically viable crops and plant commodities, to addressing legitimate concerns raised about the safety and containment of transgenic plants in the environment. Compared with the temporalor spatial-specific expression of the toxin, constitutive expression of foreign proteins in transgenic plants may cause adverse effects. Constitutive overexpression of transgenes that interfere with normal processes in a plant underscores the need for refinement of transgene expression. The development of tissue-specific promoters to drive transgene expression has helped us to fulfil that need. Therefore, in certain circumstances, it is desirable to use expression-specific promoters which express only the foreign gene in specific plant tissues or organs. This review highlights the uses and benefits reaped by researchers by using a green tissue specific promoter, RuBisCO small subunit promoter, in different crops and systems and thus establishing a broad range of tissue specific promoters. Such plant promoters that are activated precisely when and where they are needed would be ideal for genetic engineering strategies.

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Arabidopsis thaliana small subunit leader and transit peptide enhance the expression ofBacillus thuringiensis proteins in transgenic plants

Cited by 102 publications

References 25 publications

New Qualitative Detection Methods of Genetically Modified Potatoes

New Qualitative Detection Methods of Genetically Modified Potatoes

Direct Evidence for Rapid Degradation ofBacillus thuringiensis Toxin mRNA as a Cause of Poor Expression in Plants1

Endeavours of RuBisCO small subunit promoter as a tool of green tissue specific expression

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