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 - 3.36

Isolating genes potentially involved in the ABA pathway in a tomato mutant by using cDNA-AFLP

Sunseri F.*, Sciancalepore A.*^,**, Martelli G.*, Greco I.*, Gallitelli M.**

*) Dipartimento di Biologia, Difesa e Biotecnologie AgroForestali, Università della Basilicata, Contrada Macchia Romana, 85100 Potenza

**) Metapontum Agrobios, S.S. Jonica 106 Km 448,2, 75010 Metaponto (MT)

Lycopersicon esculentum Mill., TDFs, transcript-derived fragments, ABA-deficient mutant, sitiens

Tomato is a relevant economic plant species that is well characterized among crops. The Solanaceuos species has a relatively small genome (Arumuganathan & Earle, 1991 - Plant Mol Biol Rep 9, 208–219) and efforts are in progress to obtain the complete sequence (www.tigr.org/tdb). Cultivated tomato is known for its low level of polymorphism, and Amplified fragment length polymorphism (AFLP) is a PCR‑based technique capable of detecting more than 50 independent loci in a single PCR reaction.

On the other hand, abscisic acid plays an important role in many aspects of plant growth and development as well as in adaptation to abiotic stresses (Zeevaart & Creelman 1988 - Ann Rev Plant Phys Plant Mol Biol 39, 439–473). The isolation of the genes related to ABA pathway will be essential for determining the role of ABA.

Tomato sit mutant was obtained by X-ray (Stubbe, 1957 - Kulturpflanze 5, 190) and it is deficient in ABA biosynthesis. The objective of the present study was to assess the polymorphic AFLP fragments between two isogenic lines (IL) of tomato (Lycopersicon esculentum Mill.), obtained from a backcross of an ABA-deficient mutant in a genetic background cv. “Money Maker”, both at gDNA and cDNA level.

Ninety primer pairs were tested on cDNA detecting a total of ~ 9.000 AFLP fragments in the IL, with an average of 100 fragments obtained per primer pair, 271 fragments (3% of the total observed) were polymorphic, between the genotypes tested. Thirty primer pairs tested on genomic DNA resulted in ~ 3.200 AFLP fragments (an average of 103 fragments per primer pair), only 9 fragments (0.28%) were polymorphic between the wild type and the isogenic ABA-deficient mutant sitiens. The isolated transcript-derived fragments (TDFs) which show differential profiles between the IL were used to characterize cDNA clones potentially involved in the ABA pathway.

Generation of ESTs as differentially expressed sequences, between IL, is proposed as a way of developing ESTs around specific trait for an integrated approach to isolate genes of interest. Cloned TDFs with similarity to sequences linked to sit gene will be mapped for chromosome walking experiments. For this purpose, an interspecific F₂ population between sit and Lycopersicon pennellii was obtained. Until now, starting from the 271 transcript-derived fragments (TDFs) detected, 38 fragments were isolated and 4 sequenced. The obtained sequences, compared against public databases, showed similarity with several EST tomato sequences of unknown functions.