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Development of EST derived-SSRs of Theobroma grandiflorum for molecular breeding : S04P05

Santos L.F., Moreira R.F., Falcão L.L., Silva-Werneck J.O., Alves R.M., Gramacho K.P., Micheli F., Marcellino L.H.. 2013. In : Brasileiro Ana Christina Miranda (ed.), Fortes Ferreira Claudia (ed.), Fernandez Diana (ed.), Micheli Fabienne (ed.), Coelho Filho M.A. (ed.), Marraccini Pierre (ed.). Biotic and Abiotic Stress Tolerance in Plants: the Challenge for the 21st Century : Book of abstracts of the CIBA 2013. Brasilia : EMBRAPA, p. 56-57. Workshop on Biotic and Abiotic Stress Tolerance in Plants: the Challenge for the 21st Century, 2013-11-06/2013-11-08, Ilhéus-Bahia (Brésil).

Simple sequence repeats (SSRs) markers are widely used in breeding programs, and SSRs associated to expressed sequence tags (ESTs) could be quite efficient in the selection of genotypes of interest. The breeding program of cupuassu (Theobroma grandiflorum) lacks molecular tools to assist the selection of genotypes with desirable characteristics, such as disease resistance, fruit quality or productivity. The objective of this study was to identify SSRs in cupuassu ESTs, and to design and validate primers for molecular marker development. First, 441 ESTs containing 498 EST-SSRs were identified in 8,330 contigs from cupuassu 454-sequencing. The distribution of the SSRs was: 171 (34.33%) mono-, 142 (28.51%) di-, 171 (34.33%) tri-, 7 (1.4%) tetra-, 1 (0.2%) penta- and 6 (1.2%) hexanucleotides. From the 441 ESTs, 70 were selected based on their putative function (e.g. metabolism of sugars, resistance to biotic and abiotic stresses) and 77 primer pairs flanking the SSRs where defined and tested on 16 cupuassu genotypes presenting differences in pulp quality and/or resistance to witches' broom disease. The analysis of the 77 loci allowed the identification of 22 polymorphic loci with a total of 125 alleles. The number the alleles ranged from 2 to 8 with an average of 5.17. The observed heterozygosity (HO) values ranged from 0.21 to 1 with an average of 0.71, and the expected heterozygosity (HE) values ranged from 0.33 to 0.87. The PIC values ranged from 0.30 to 0.77 with an average of 0.71. The 22 polymorphic primers from cupuassu were used in transferability test on DNA from cocoa (T. cacao). Among the 22 primers analyzed, 20 amplified DNA from cocoa, with a transferability rate of 90%. In conclusion, we identified EST-SSRs markers that may be used for cupuassu as well as cocoa breeding programs. Financial Support: Capes/Embrapa; Cirad; UESC. (Texte intégral)

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