An efficient method for the production of marker-free transgenic plants of peanut (Arachis hypogaea L.). 2010

Madhurima Bhatnagar, and Kalyani Prasad, and Pooja Bhatnagar-Mathur, and M Lakshmi Narasu, and Farid Waliyar, and Kiran K Sharma
Genetic Transformation Laboratory, International Crops Research Institute for the Semi-Arid Tropics (ICRISAT), Patancheru, Andhra Pradesh, India.

Recombinant genes conferring resistance to antibiotics or herbicides are widely used as selectable markers in plant transformation for selecting the primary transgenic events. However, these become redundant once the transgenic plants have been developed and identified. Although, there is no evidence that the selectable marker genes are unsafe for consumers and the environment, it would be desirable if the marker genes can be eliminated from the final transgenic events. The availability of efficient transformation methods can enable the possibility of developing transgenic events that are devoid of the marker gene/s upfront. Taking advantage of the high and consistent transformation potential of peanut, we report a technique for developing its transgenics without the use of any selectable marker gene. Marker-free binary vectors harboring either the phytoene synthase gene from maize (Zmpsy1) or the chitinase gene from rice (Rchit) were constructed and used for Agrobacterium tumefaciens-mediated transformation of peanut. The putative transgenic events growing in vitro were initially identified by PCR and further confirmed for gene integration and expression by dot blots assays, Southern blots, and RT-PCR where they showed a transformation frequency of over 75%. This system is simple, efficient, rapid, and does not require the complex segregation steps and analysis for selection of the transgenic events. This approach for generation of marker-free transgenic plants minimizes the risk of introducing unwanted genetic changes, allows stacking of multiple genes and can be applicable to other plant species that have high shoot regeneration efficiencies.

UI MeSH Term Description Entries
D010367 Arachis A plant genus of the family FABACEAE that includes peanuts. Arachis hypogaea,Peanuts,Peanut
D003313 Zea mays A plant species of the family POACEAE. It is a tall grass grown for its EDIBLE GRAIN, corn, used as food and animal FODDER. Corn,Indian Corn,Maize,Teosinte,Zea,Corn, Indian
D005818 Genetic Engineering Directed modification of the gene complement of a living organism by such techniques as altering the DNA, substituting genetic material by means of a virus, transplanting whole nuclei, transplanting cell hybrids, etc. Genetic Intervention,Engineering, Genetic,Intervention, Genetic,Genetic Interventions,Interventions, Genetic
D005822 Genetic Vectors DNA molecules capable of autonomous replication within a host cell and into which other DNA sequences can be inserted and thus amplified. Many are derived from PLASMIDS; BACTERIOPHAGES; or VIRUSES. They are used for transporting foreign genes into recipient cells. Genetic vectors possess a functional replicator site and contain GENETIC MARKERS to facilitate their selective recognition. Cloning Vectors,Shuttle Vectors,Vectors, Genetic,Cloning Vector,Genetic Vector,Shuttle Vector,Vector, Cloning,Vector, Genetic,Vector, Shuttle,Vectors, Cloning,Vectors, Shuttle
D012275 Oryza A genus of grass family (Poaceae) that include several rice species. Oryza sativa,Rice,Rices
D014170 Transformation, Genetic Change brought about to an organisms genetic composition by unidirectional transfer (TRANSFECTION; TRANSDUCTION, GENETIC; CONJUGATION, GENETIC, etc.) and incorporation of foreign DNA into prokaryotic or eukaryotic cells by recombination of part or all of that DNA into the cell's genome. Genetic Transformation,Genetic Transformations,Transformations, Genetic
D015139 Blotting, Southern A method (first developed by E.M. Southern) for detection of DNA that has been electrophoretically separated and immobilized by blotting on nitrocellulose or other type of paper or nylon membrane followed by hybridization with labeled NUCLEIC ACID PROBES. Southern Blotting,Blot, Southern,Southern Blot
D016960 Agrobacterium tumefaciens A species of gram-negative, aerobic bacteria isolated from soil and the stems, leafs, and roots of plants. Some biotypes are pathogenic and cause the formation of PLANT TUMORS in a wide variety of higher plants. The species is a major research tool in biotechnology. Alcaligenes radiobacter,Bacillus radiobacter,Bacterium radiobacter,Bacterium tumefaciens,Phytomonas tumefaciens,Polymonas tumefaciens,Pseudomonas radiobacter,Pseudomonas tumefaciens,Rhizobium radiobacter,Agrobacterium radiobacter
D020133 Reverse Transcriptase Polymerase Chain Reaction A variation of the PCR technique in which cDNA is made from RNA via reverse transcription. The resultant cDNA is then amplified using standard PCR protocols. Polymerase Chain Reaction, Reverse Transcriptase,Reverse Transcriptase PCR,PCR, Reverse Transcriptase,Transcriptase PCR, Reverse
D030821 Plants, Genetically Modified PLANTS, or their progeny, whose GENOME has been altered by GENETIC ENGINEERING. Genetically Modified Plants,Plants, Transgenic,Transgenic Plants,GMO Plants,Genetically Engineered Plants,Engineered Plant, Genetically,Engineered Plants, Genetically,GMO Plant,Genetically Engineered Plant,Genetically Modified Plant,Modified Plant, Genetically,Modified Plants, Genetically,Plant, GMO,Plant, Genetically Engineered,Plant, Genetically Modified,Plant, Transgenic,Plants, GMO,Plants, Genetically Engineered,Transgenic Plant

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