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Positive Selection

The use of a new mode of selection—positive selection—has been demonstrated to be successful in a large variety of monocot and dicot species. This selection differs from more traditional modes of selection in which compounds such as antibiotics or herbicides are used to kill nontransformed ...

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Elimination of Marker Genes From Transgenic Plants Using MAT Vector Systems

We have developed an efficient system (Multi-Auto-Transformation vectors) for the removal of marker genes and to increase the regeneration frequency of transgenic crops without using antibiotic selection, reducing their possible environmental impact. The MAT vector system is ...

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Simple and Efficient Removal of Marker Genes From Plastids by Homologous Recombination

Removal of marker genes improves the design of transgenic plants. Homologous recombination between direct repeats provides a simple method for excising marker genes after transgenic cells and shoots have been isolated. Efficient implementation of the method requires high rates of ...

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The Study of Transgene Copy Number and Organization

The development of efficient crop transformation systems has necessitated the development of efficient methods for detailed molecular characterization of putative events. This chapter details the routine use of quantitative real-time polymerase chain reaction to determ ...

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Analysis of Gene Expression in Transgenic Plants

Recent years have seen a huge increase and improvement in techniques for analysis of transgene expression in plants. The analysis of RNA frequently provides a vital link between changes in enzyme levels and/or metabolites and the phenotype. This chapter focuses on RNA-based techniques for ...

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Transgene Integration: Use of Matrix Attachment Regions

Matrix attachment regions (MARs) are operationally defined as DNA elements that bind specifically to the nuclear matrix in vitro. When MARs are positioned at the 5′- and 3′-ends of a transgene higher more predictable expression of the transgene results. MARs are increasingly being applied to ...

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Fluorescence In Situ Hybridization to Localize Transgenes in Plant Chromosomes

Production of transgenic plants is now routine for many of our crop species. Methods for the detailed molecular analysis of transgenic plants are available, but often the exact location of the transgene within the crop genome is poorly understood. As a starting point to understanding more abo ...

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Agrobacterium Persistence in Plant Tissues After Transformation

Agrobacterium spp. are routinely used in plant transformation to introduce genes of interest in valuable economic species. However, several agrobacteria species are also plant pathogens with ability to survive in different environments including the inner part of the plants. To avo ...

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Amplification of Genomic Sequences Flanking T-DNA Insertions by Thermal Asymmetric Interlaced Polymerase Chain Reaction

Thermal asymmetric interlaced polymerase chain reaction (TAIL-PCR) is an efficient tool for the recovery of DNA fragments adjacent to known sequences. A protocol is presented for the amplification of genomic sequences flanking DNA (T-DNA or transposon) insertions using the TAIL-PCR ...

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Transgene Dispersal Through Pollen

Techniques used for the transfer of novel genes into host plant genomes have created new possibilities for crop improvement. The implementation of transgenic crop species into agriculture has introduced the possibility of transgene escape into the environment via pollen dispers ...

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Transgenic Crops: The Current and Next Generations

This chapter describes the present status and future prospects for transgenic (genetically modified) crops. It concentrates on the most recent data obtained from patent databases and field trial applications, as well as the usual scientific literature. By these means, it is possible to o ...

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Comparative Development and Impact of Transgenic Papayas in Hawaii, Jamaica, and Venezuela

We present data concerning the creation of transgenic papayas resistant to Papaya ringspot virus (PRSV) and their adoption by three different countries: the United States (e.g., Hawaii), Jamaica, and Venezuela. Although the three sets of transgenic papayas showed effective resistan ...

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Carrot (Daucus carota L.)

Plants are susceptible to infection by a broad range of fungal pathogens. Many horticulturally important crop species lack adequate genetic resistance to disease. Studies on potential mechanisms of disease resistance in plants have revealed the importance of a range of pathogenesi ...

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Potato (Solanum tuberosum L.)

Potato (Solanum tuberosum L.) is a globally important crop plant producing high yields of nutritionally valuable food in the form of tubers. It has been the focus of substantial study because of its use both as a staple food crop and as a potentially significant source of compounds of interest. This has i ...

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Cassava (Manihot esculenta Crantz)

During the last three years the generation of stably transformed cassava plants having value-added traits has become a reality. Currently, two Agrobacterium-mediated transformation systems are routinely used to engineer cassava. These systems use either somatic embryos or fri ...

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Bermudagrass (Cynodon spp.)

Bermudagrass is an important warm-season forage and turf species widely grown in the southern United States. This chapter describes a rapid and efficient protocol that allows for the generation of a large number of transgenic bermudagrass plants, bypassing the callus formation phase. S ...

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Sweet Potato [Ipomoea batatas (L.) Lam.]

Among the available transformation methods reported on sweet potato, Agrobacterium tumefaciens-mediated transformation is more successful and desirable. Stem explants have shown to be ideal for the transformation of sweet potato because of their ready availability as expla ...

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Perennial Ryegrass (Lolium perenne L.)

A protocol that facilitates rapid establishment of Agrobacterium-mediated transformation for perennial ryegrass is described. The synthetic green fluorescent protein (sgfpS65T) reporter gene is introduced in combination with the nptII selectable marker gene into axil ...

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Tall Fescue (Festuca arundinacea Schreb.)

Tall fescue (Festuca arundinacea Schreb.) is the predominant cool-season perennial grass in the United States. It is widely used for both forage and turf purposes. This chapter describes a protocol that allows for the generation of large number of transgenic tall fescue plants by Agrobacte ...

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Switchgrass (Panicum virgatum L.)

During the last decade, Agrobacterium-mediated transformation of more than a dozen monocotyledonous plants, including forage and turf grasses, has been achieved. So far, switchgrass is the only warm season grass that has been transformed with A. tumefaciens. We have developed a highly ef ...

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