Genetic Engineering and Gene Transformation
Saeed Soheilivand; Amir Mousavi; Mohammadreza Safarnejad
Abstract
Sour lime (Citrus aurantifolia L.) is one of the most important woody plants is widely known for its recalcitrance to genetic transformation. We aimed herein to evaluate effective factors influencing the transformation efficiency and the reduction of chimeric transgenic shoots in sour lime. Epicotyl ...
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Sour lime (Citrus aurantifolia L.) is one of the most important woody plants is widely known for its recalcitrance to genetic transformation. We aimed herein to evaluate effective factors influencing the transformation efficiency and the reduction of chimeric transgenic shoots in sour lime. Epicotyl and internode explants were genetically transformed with different Agrobacterium tumefaciens strains e.g., LBA4404, GV3850, and GV3101, harboring the vectors pBI121 and pCAMBIA3301 containing β-glucuronidase (GUS) as a reporter gene. The effect of the following factors was evaluated: Agrobacterium concentration (OD600=0.3, 0.5 and 1), during inoculation (5 seconds, 10 minutes and 30 minutes), co-culture (2 and 3 days), and the selection regime (phosphinothricin at 1, 3, 5 and 10 mg/l and kanamycin at 25, 50, 75 and 100 mg/l). In following, transformation efficiency and the chimeric transgenic shoots rate were respectively confirmed by PCR and GUS assays. The results showed that Agrobacterium strain LBA4404, at the OD600 of 0.5, with 5 seconds (for epicotyl) and 10 minutes (for internode) inoculation at two-day co-culture period, were identified the most suitable treatments for both explants. The transformation frequencies ranged from 0.93% for internode on DKW medium containing 1.0 mg/l of phosphinothricin to 14.29% for epicotyl on DKW medium containing 50 mg/l of kanamycin. Inclusion of the high-level of selective treatments, improved the transformation rate through decreasing frequency escape and chimeric transgenic shoots. These findings provide novel insights into the appropriate procedure to constitute non-chimeric lime transgenic shoots.
Molecular Genetics and Genetic Engineering
Zahra Fathi; Katayoun Zamani; Mohammad Malboobi
Abstract
Phosphite is a reduced form of phosphate, wherein an oxygen replaces a hydrogen atom, and this substitution has a significant effect on its performance in living organisms. Phosphite is readily transfered into plant cells through phosphate transporters. However, plants do not have the ability to use ...
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Phosphite is a reduced form of phosphate, wherein an oxygen replaces a hydrogen atom, and this substitution has a significant effect on its performance in living organisms. Phosphite is readily transfered into plant cells through phosphate transporters. However, plants do not have the ability to use phosphite as a phosphorus resourc such that this property has limited the use of phosphite as fertilizer; however, phosphite has been used as a fungicide and biostimulant in agriculture. Some bacteria are able to oxidize phosphite into phosphate to cover for various cellular functions. In the last decade, the molecular mechanism of this biological oxidation has been elucidated to occure by the enzyme phosphite oxidoreductase or phosphite dehydrogenase. Phosphite is produced in large quantities in various chemical industries as a by-product or waste that is not recycled. The identification of the enzyme phosphite dehydrogenase, that catalyses the oxidation of phosphite to phosphate, has opened a new path for the recycle of this waste. Recently, there have been reports for the production of transgenic plants expressing ptxD gene. In practice, ptxD gene can be used as a marker in the selection of transgenic plants. By producing these transgenic plants, phosphite can be used as a herbicide and even as a phosphorus fertilizer.