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Production of transgenic carnation with antisense ACS (1-aminocyclopropane44-carboxy late synthase) gene
104-107.Views:167Dianthus chinensis and Dianthus caryophyllus varieties were tested for shoot regeneration from leaf and petal explants and transformed with Agrobacterium tuniefaciens strains (EHA 105 and LBA 4404) harbouring an apple derived ACS cDNA in antisense orientation in order to reduce ethylene production and influence the ethylene dependant traits in carnation. After transformation regenerating shoots were selected on MS medium containing 50-75-100-125-150 mg/1 kanamycin and supplemented with 1 mg/1 BA, 0.2 mg/1 NAA. Transgene integration was proved by PCR analysis with npt II spcific primers followed by Southern hybridisation of DNA isolated from green shoots on medium containing 150 mg/1 kanamycin. Several putative transformants were subjected to RT-PCR in order to examine the npt 11 expression at mRNA level. Both the transformant and the non-transformant plants were potted into glasshouse to observe the effect of changed ethylene production on flowering time, petal senescence and vase life.
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Review of the self-incompatibility in apple (Malus x domestica Borkh., syn.: Malus pumila Mill.)
31-36.Views:508Apple (Malus x domestica Borkh.) is one of the most important fruit crops showing ribonuclease-mediated self-incompatibility, and no self-compatible apple cultivars are known. Twenty-nine S-alleles were identified in apple and many more incompatibility groups are present compared to sweet cherry. Results from a Belgian, English and a Japanese research group are combined and the S-genotypes of the most important world cultivars are collected. Two different allele labelling system are reconciled and detection methods used in case of the specific alleles are shown. Effects of the resistance breeding programmes are discussed; and scientific efforts involving transgenic technology to create self-compatible genotypes are shown. This review covers the most interesting issues regarding self-incompatibility in apple.
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The effects of ACS (1-aminocyclopropane-l-carboxylate synthase) gene down regulation on ethylene production and fruit softening in transgenic apple
65-70.Views:113A detailed examination of the production of ethylene and other ripening parameters during storage period has been undertaken in transgenic apple fruits, where the ethylene biosynthesis was inhibited by antisense ACS (l-aminocyclopropane-l-carboxylate synthase) gene. Data indicate down regulation of ethylene production, softening and spoilage in some transgenic lines. In some cases ethylene production was inhibited for over 90 percent, considerable reduction of softening and spoilage was observed probably due to the reduced activity of cell wall degradable enzymes. ACS activity was also monitored during ripening. The fruits of the best transgenic lines could be stored for minimum 4-5 months longer under 5 °C cold room storage conditions and one month longer at normal room temperature. This molecular approach can provide an alternative way to replace the commonly used and costly atmospheric storage of fruits.
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Genetic engineering of apple (Malus domestica Borkh.) for resistance to fungal diseases using g2ps1 gene from Gerbera hybrida (Asteraceae)
15-12.Views:267In the present study, g2ps1 gene from Gerbera hybrida coding for 2-pyrone synthase which contribute for fungal and insect resistance was used. The aim was to work out an efficient approach of genetic transformation for apple cvs. ‘Golden Delicious’, ‘Royal Gala’ and ‘MM111’, ‘M26’ rootstocks for improving their fungal resistance using genetic engineering techniques. Adventitious shoot formation from leaf pieces of apples studied was achieved using middle leaf segments taken from the youngest leaves from in vitro-grown plants.
Optimum conditions for ‚direct’ shoot organogenesis resulted in high regeneration efficiency of 0%, 95%, 92%, 94% in the studied apples respectively. Putative transgenic shoots could be obtained on MS media with B5 Vitamins, 5.0 mg l-1 BAP, or 2.0 mg l-1 TDZ with 0.2 mg l-1 NAA in the presence of the selection agent “PPT” at 3.0-5.0 mgl-1. Shoot multiplication of transgenic shoots was achieved on: MS + B5 vitamins + 1.0 mg l-1 BAP + 0.3 mg l-1 IBA, 0.2 mg l-1 GA3+1.0 g/l MES+ 30 g/l sucrose + 7.0 g/l Agar, with the selection agent PPT at 5.0 mg l-1 and were subcultured every 4 weeks in order to get sufficient material to confirm transformation of the putative shoots obtained. Six, seven, one and six transgenic clones of the apples studied respectively have been obtained and confirmed by selection on the media containing the selection agent “PPT” and by PCR analysis using the suitable primers in all clones obtained for the presence of the selection” bar gene (447 bp) and the gene-of- interest “g2PS1” (1244 bp), with transformation efficiency of 0.4%, 0.6%, 0.1% and 0.3% respectively. These transgenic clones were multiplied further in vitro in the presence of the selection agent ‘PPT’ and rooted in vitro. Rooted transgenic plantlets were successfully acclimatized and are being kept under-containment conditions according to the biosafety by-law in Syria to evaluate their performance for fungal resistance . -
Down-regulation of ethylene production in carnation (Dianthus Caryphyllus L.) by an apple derived ACC-cDNA
101-104.Views:128Transgenic carnations were produced with an apple derived antisense ACC-synthase cDNA. Transgenic carnation regenerants were potted in glasshouse. All transformed plants showed normal growth and were true-to-type. Ethylene production — measured at full opening stage — lowered by 30-60 %, no plant with 100 % decrease was identified. The vase-life has been observed for 5 years. 38 % of the transformant carnations showed a higher a relative value in days by more than 2 days to 6 days. Twenty six plants were found exhibiting the most marked alterations in the tested trait. In these plants ethylene production decreased by 37-67 %, they have longer vase-life (by 4 days or more). Since the fragrance variety 'Bíbor' was the plant material for genetic modification of vase-life, this trait has been conserved after transformation in spite of the fact that the position of transgene integration cannot be directed.