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 Plant Breeding
M Abkar; F Sanjarian; A Mousavi
Volume 2, Issue 3 , January 2013, , Pages 13-23
Abstract
Fusarium head blight (FHB) is a disease that causes major economic losses in wheat and other cereal crops production worldwide. Contamination of food with the trichothecene mycotoxin deoxynivalenol (DON) produced by Fusarium graminearum is a major health concern for humans and animals because trichothecenes ...
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Fusarium head blight (FHB) is a disease that causes major economic losses in wheat and other cereal crops production worldwide. Contamination of food with the trichothecene mycotoxin deoxynivalenol (DON) produced by Fusarium graminearum is a major health concern for humans and animals because trichothecenes are potent cytotoxins of eukaryotic cells. Trichothecene mycotoxins inhibit translation by targeting ribosomal protein L3 at the peptidyl transferase center. In this study, we modified a Tomato (Lycopersicon esculentum) cDNA encoding the ribosomal protein RPL3 so that amino acid residue 258 is changed from tryptophan to cysteine and amino acid residue 259 is change from histidine to tyrosine. All version of the tomato RPL3 were introduced to DON-sensitive pdr5 and ayt1 mutant strain Saccharomyces cerevisiae. When transgenic yeast were compared for growth in the presence of DON, a difference in growth rate and survival was observed among those yeasts expressing the modified versions of the tomato RPL3 genes, compared to those expressing the wild-type yeast RPL3 gene. These results can create a new field in developing FHB resistance varieties of wheat through genetic manipulation.