Articles
DEFENSE AGAINST PATHOGENIC BACTERIA IN TRANSGENIC POTATO PLANTS
Article number
447_86
Pages
423 – 430
Language
Abstract
Humoral antibacterial substances were reported to be induced in the giant silk-moth larvae over two decades ago.
Since then such substances, mostly identified as short proteins, were found in many insects of various groups (e.g.
Lepidoptera, Diptera, Hymenoptera). Bactericidal proteins were identified in larvae of the flesh-fly (Sarcophaga peregrina) by Natori and associates in 1977 and the cDNA coding for Sarcotoxin IA (sarco) was subsequently isolated by these investigators.
The respective protein was also characterized and the mature proteins mass is about 5 kd.
We recruited sarco to confer tolerance against bacterial rots in tubers and roots of potato, that are caused by Erwinia spp and Pseudomonas solanacearum. Consequently we constructed transformation vectors that contained the cDNA of sarco or the gus reporter gene.
In each case we engineered the root-specific promoter (Tob) or the tuber promoter (Pat), upstream of the respective coding sequence.
Such cassettes, that also included the omega translational enhancer and a terminator sequence were transferred to Agrobacteria and genetic transformation of tuber discs was performed.
Five potato cultivars and breeding lines were used: Desiree, Achirana INTA, LT-9, TS-10, TS-15. Potato plants that regenerated from Agrobacterium infected tuber-discs and rooted on selective medium were regarded putative transformants and were further analyzed.
We found that putative transformants that resulted from transformation of vectors that contained Gus driven by Tob and Pat indeed expressed the reporter gene in their roots and tubers, respectively.
This verified the potency and specificity of the chimeric genes in the transformation vectors.
Polyclonal anti-sarco antibodies were produced and used to evaluate the expression of sarco in the putative transgenic potato plants.
Preliminary western-blot assays indicated that indeed the roots of some of the plants that were transformed with the chimeric-gene that contained the Tob promoter and the sarco cDNA showed bands that reacted with the anti–sarco antibodies.
Since then such substances, mostly identified as short proteins, were found in many insects of various groups (e.g.
Lepidoptera, Diptera, Hymenoptera). Bactericidal proteins were identified in larvae of the flesh-fly (Sarcophaga peregrina) by Natori and associates in 1977 and the cDNA coding for Sarcotoxin IA (sarco) was subsequently isolated by these investigators.
The respective protein was also characterized and the mature proteins mass is about 5 kd.
We recruited sarco to confer tolerance against bacterial rots in tubers and roots of potato, that are caused by Erwinia spp and Pseudomonas solanacearum. Consequently we constructed transformation vectors that contained the cDNA of sarco or the gus reporter gene.
In each case we engineered the root-specific promoter (Tob) or the tuber promoter (Pat), upstream of the respective coding sequence.
Such cassettes, that also included the omega translational enhancer and a terminator sequence were transferred to Agrobacteria and genetic transformation of tuber discs was performed.
Five potato cultivars and breeding lines were used: Desiree, Achirana INTA, LT-9, TS-10, TS-15. Potato plants that regenerated from Agrobacterium infected tuber-discs and rooted on selective medium were regarded putative transformants and were further analyzed.
We found that putative transformants that resulted from transformation of vectors that contained Gus driven by Tob and Pat indeed expressed the reporter gene in their roots and tubers, respectively.
This verified the potency and specificity of the chimeric genes in the transformation vectors.
Polyclonal anti-sarco antibodies were produced and used to evaluate the expression of sarco in the putative transgenic potato plants.
Preliminary western-blot assays indicated that indeed the roots of some of the plants that were transformed with the chimeric-gene that contained the Tob promoter and the sarco cDNA showed bands that reacted with the anti–sarco antibodies.
Authors
E. Galun, D. Aviv, Y. Mahler-Slasky, S. Wolf, Y. Mahler-Slasky, S. Galili, A. Perl, R. Aly
Keywords
Erwinia spp., Pseudomonas solanacearum, Solanum tuberosum, Sarcophaga peregrina, Sarcotoxin, Transformation
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