Articles
STUDIES TO IMPROVE TRANSIENT GUSA EXPRESSION IN A. TUMEFACIENS-MEDIATED TRANSFORMATION OF RICE
Article number
461_47
Pages
409 – 416
Language
Abstract
The direct-DNA uptake methods has been routinely used in transforming rice.
The recent development of rice genetic transformation using A. tumefaciens as a natural vector necessitate the investigation of factors that will promote optimum gene expression and T-DNA transfer in the recalcitrant and economically important indica rice varieties.
Activities of three different promoters were investigated for transient expression using gusA after particle bombardment of suspension cells.
Actin and ubiquitin promoters were found to induce higher GUS activity compared to the chimeric promoter of mannopine synthethase and a truncated cauliflower mosaic virus 35S promoter.
A binary vector pCGN1558 was modified to contain these promoters in front of the gusA-intron. A. tumefaciens strain EHA101 and A136 were transformed with the derivatives of the binary vector and transformed colonies were tested for infectivity on coleoptile bases of IR54 and IR72 seedlings.
GUS activity of the infected coleoptile bases showed that the A. tumefaciens colonies were comparable to but not better than the positive control At656. Possible reasons for the obtained results were explained with emphasis on the importance of the compatibility of the A. tumefaciens strains with the binary vector, the border sequences, and the Ti-plasmid present.
The recent development of rice genetic transformation using A. tumefaciens as a natural vector necessitate the investigation of factors that will promote optimum gene expression and T-DNA transfer in the recalcitrant and economically important indica rice varieties.
Activities of three different promoters were investigated for transient expression using gusA after particle bombardment of suspension cells.
Actin and ubiquitin promoters were found to induce higher GUS activity compared to the chimeric promoter of mannopine synthethase and a truncated cauliflower mosaic virus 35S promoter.
A binary vector pCGN1558 was modified to contain these promoters in front of the gusA-intron. A. tumefaciens strain EHA101 and A136 were transformed with the derivatives of the binary vector and transformed colonies were tested for infectivity on coleoptile bases of IR54 and IR72 seedlings.
GUS activity of the infected coleoptile bases showed that the A. tumefaciens colonies were comparable to but not better than the positive control At656. Possible reasons for the obtained results were explained with emphasis on the importance of the compatibility of the A. tumefaciens strains with the binary vector, the border sequences, and the Ti-plasmid present.
Authors
R. R. Aldemita
Keywords
Agrobacterium tumefaciens, transformation, binary vector, β-glucuronidase
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