Proceedings of the XLVI Italian Society of Agricultural Genetics - SIGA Annual Congress

Giardini Naxos, Italy - 18/21 September, 2002

ISBN 88-900622-3-1

Poster Abstract - 4.34

MOLECULAR CHARACTERIZATION OF POTATO TRANSGENIC PLANTS

CARTOLANO M.*, CONSIGLIO F.**, COZZOLINO A.*, FILIPPONE E.*, BARONE A.*

*) Dept. of Soil, Plant and Environmental Sciences, School of Biotechnology Sciences, Via Università 100, 80055 Portici, Italy

**) CNR-IMOF, Via Università 133, 80055 Portici, Italy

transgenic tetraploid potatoes, AFLP markers, T-DNA junctions, TAIL-PCR

Transgenic potato plants, transformed with the Trichoderma harzianum endochitinase gene, were studied in order to 1) analyse the genetic variability induced by the transformation event 2) characterize the genomic regions flanking the T-DNA insert.

An AFLP analysis was carried out on 10 transgenic S.tuberosum cv. Desirèe plants, 2 micropropagated genotypes, 2 vegetatively propagated genotypes and 5 callus regenerated in vitro genotypes of the same cv. Desirèe. Twenty primer combinations allowed 1069 total AFLP markers to be clearly identified. The average percentage of polymorphic markers was 1.12% among the transgenic genotypes, 4.49% among the regenerated genotypes and 1.03% between the two that were micropropagated. The c2 statistical analysis provided evidence of a not significant level of polymorphism between the transgenic and vegetatively propagated genotypes (c2= 0.023, P<0.05), whereas a significant difference between the regenerated and transgenic genotypes (c2 =5.44, P<0.05) was found. The analysis performed allowed the 0.02% of tetraploid potato genome to be covered. Further AFLP markers are being tested in order to increase the portion of analysed genome.

Moreover, in order to monitor where in the potato genome of our transgenic plants the endochitinase gene has been inserted, a modified TAIL-PCR method (Liu et al.; 1995) was successfully pointed out to isolate the T-DNA junctions in tetraploid potatoes. The essential modifications are related to the necessity to amplify a target sequence in the more complex cultivated potato genome (1730 Mbp/1C) than the Arabidopsis one, for which the original TAIL-PCR was developed.

A DNA fragment of 400 bp outside of the T-DNA left border was isolated in one transgenic genotype and was preliminary characterized through bioinformatic tools. Further analyses are in progress to get more information about this region and to also identify the junction at the T-DNA right border. This molecular analysis will allow us to understand if functional genes or regulatory sequences have been interrupted due to the transformation events.

Liu et al., 1995 Plant Journal 8: 457-463.