Stable transformation of soybean by electroporation and root formation from transformed callus

Christou, Paul; Murphy, Jean E.; Swain, William F.

doi:10.1073/pnas.84.12.3962

Cited by 60 publications

(21 citation statements)

References 19 publications

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“…pAcX1100P and pAcX1021P were constructed by inserting the chimeric 8-glucuronidase (GUS) (2) and neomycin phosphotransferase II (NPT II) (3) genes from pCMC1100 (1) and pCMC1021 (4), respectively, into the unique Xho I site ofpAc3 (5). The insertions in these plasmids are oriented such that the direction of transcription is the same as that of the Ac element (6).…”

Section: Methodsmentioning

confidence: 99%

Inheritance and expression of foreign genes in transgenic soybean plants

Christou¹,

Swain²,

Yang³

et al. 1989

Proc. Natl. Acad. Sci. U.S.A.

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188

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DNA-coated gold particles were introduced into meristems of immature soybean seeds using electric discharge particle acceleration to produce transgenic fertile soybean plants. The lineages of integrated foreign DNA in two independently transformed plants were followed in the first (R1) and second (R2) generation of self-pollinated progeny. One plant (4615) was transformed with the Escherichia coli genes for 8-glucuronidase and neomycin phosphotransferase II; the other (3993) was transformed only with the gene for 18-glucuronidase. Segregation ratios for the introduced gene(s) were approximately 3:1 for plant 4615 and 1:1 for plant 3993 in the R1 generation. DNA analysis showed 100% concordance between presence of the foreign gene sequences and enzyme activity. Moreover, all copies of the foreign genes are inherited as a unit in each plant. Plant 3993 segregated in a 1:1 ratio in the R2 generation. RI plants derived from plant 4615, which expressed both genes, gave either 100% or 3:1 expression of both genes in the R2 generation, demonstrating recovery of both homozygous and heterozygous R, plants. Our results show that foreign DNA introduced into soybean plants using electric discharge particle acceleration can be inherited in a Mendelian manner. Results also demonstrate cotransformation of tandem markers and show that both markers are inherited as closely linked genes in subsequent generations. These results indicate that whole plants can be derived from single transformed cells by a de novo organogenic pathway.Technologies are being developed to transform the more important agronomic crops since Agrobacterium-or protoplast-based transformation methods are currently unsuccessful or inefficient with these plants. We described (1) how electric discharge particle acceleration can stably transform a major commercial cultivar of soybean. This method bypasses traditional tissue culture operations involving protoplast and callus cultures and may thereby reduce varietal restrictions and problems of somaclonal variation frequently associated with regeneration of plants from tissue culture. To evaluate and characterize the stability of introduced genes in transgenic plants obtained by this method, the expression and organization of two foreign marker genes were analyzed in sexually derived progeny over two generations. MATERIALS AND METHODSPlasmids. pAcX1100P and pAcX1021P were constructed by inserting the chimeric 8-glucuronidase (GUS) (2) and neomycin phosphotransferase II (NPT II) (3) genes from pCMC1100 (1) and pCMC1021 (4), respectively, into the unique Xho I site ofpAc3 (5). The insertions in these plasmids are oriented such that the direction of transcription is the same as that of the Ac element (6). Plasmids were prepared as described (1).Preparation of DNA and Particle Delivery. DNA-coated gold particles were prepared by mixing equal molar amounts of pAcX1100P, pAcX1021P, and pAc3 with 1.5-pim gold beads at a rate of 1 mg of beads to 0.01-0.1 ;kg of DNA as described (1, 7). Particle delivery into the ta...

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Section: Methodsmentioning

confidence: 99%

Inheritance and expression of foreign genes in transgenic soybean plants

Christou¹,

Swain²,

Yang³

et al. 1989

Proc. Natl. Acad. Sci. U.S.A.

Self Cite

188

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“…Protoplasts from Phaseolus vulgaris cv Negro jamapa were isolated 3 (Fig. 1), were established for both PEG-and electroporation-mediated transformation.…”

Section: Materials and Methods Protoplast Isolationmentioning

confidence: 99%

Transient Gene Expression in Protoplasts of Phaseolus vulgaris Isolated from a Cell Suspension Culture

León

Planckaert²,

Walbot³

1991

Plant Physiol.

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“…Gene electrotransfer of plant protoplasts was successfully applied for transformation of several crop species such as maize, rice, wheat, sorghum, soybean, and rye (Fromm et al 1986;Lee et al 1986;Christou et al 1987;Pitt et al 1997;Quecini et al 2002). Cell viability preservation is crucial for production of transgenic plants, as transformed protoplast should maintain the ability of normal organogenesis.…”

Section: Plant Protoplast Electroporationmentioning

confidence: 99%

Electroporation in Biological Cell and Tissue: An Overview

Kandušer

Miklavčič²

2008

Electrotechnologies for Extraction From Food Plants and Biomaterials

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In this chapter, basics and mechanisms of electroporation are presented. Most important electric pulse parameters for electroporation efficiency for different applications that involve introduction of small molecules and macromolecules into the cell or cell membrane electrofusion are described. In all these applications, cell viability has to be preserved. However, in some biotechnological applications, such as liquid food sterilization or water treatment, electroporation is used as a method for efficient cell killing. For all the applications mentioned above, besides electric pulse parameters, other factors, such as electroporation medium composition and osmotic pressure, play significant roles in electroporation effectiveness. For controlled use of the method in all applications, the basic mechanisms of electroporation need to be known. The phenomenon was studied from the single-cell level and dense cell suspension that represents a simplified homogenous tissue model, to complex biological tissues. In the latter, different cell types and electric conductivity that change during the course of electric pulse application can significantly affect the effectiveness of the treatment. For such a complex situation, the design and use of suitable electrodes and theoretical modeling of electric field distribution within the tissue are essential. Electroporation as a universal method applicable to different cell types is used for different purposes. In medicine it is used for electrochemotherapy and genetherapy. In biotechnology it is used for water and liquid food sterilization and for transfection of bacteria, yeast, plant protoplast, and intact plant tissue. Understanding the phenomenon of electroporation, its mechanisms and optimization of all the parameters that affect electroporation is a prerequisite for successful treatment. In addition to the parameters mentioned above, different biological characteristics of treated cell affect the outcome of the treatment. Electroporation, gene electrotransfer and electrofusion are affected by cell membrane fluidity, cytoskeleton, and the presence of the cell wall in bacteria yeast and plant cells. Thus, electroporation parameters need to be specifically optimized for different cell types.

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Stable transformation of soybean by electroporation and root formation from transformed callus

Cited by 60 publications

References 19 publications

Inheritance and expression of foreign genes in transgenic soybean plants

Inheritance and expression of foreign genes in transgenic soybean plants

Transient Gene Expression in Protoplasts of Phaseolus vulgaris Isolated from a Cell Suspension Culture

Electroporation in Biological Cell and Tissue: An Overview

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