Desenvolvimento e validação de marcadores moleculares tipo microssatélites a partir do transcriptoma de Vigna unguiculata (L.) Walp.

Abstract

Genetic improvement of plants has potentiated continuous gains in the productivity of several crops. In this context, molecular markers stand out as a promising tool, which allows direct access to the genotype of the individual, thus differentiating it self from the morphological markers. Microsatellites (SSR) are codominant markers and are among the most used for this purpose. Thus, the present work aimed to develop and validate SSR markers for use in genetic improvement programs of cowpea [Vigna unguiculata (L.) Walp.]. For this, three transcripts of the species, from experiments by CPSMV (Cowpea Severe Mosaic Virus), CABMV (Cowpea Aphid-born Mosaic Virus) and root dehydration were analyzed. A computational pipeline was used to identify and annotate SSRs. The selection of the candidates for the development of the markers was performed considering the polymorphism of size between the motifs, followed for the design of the primers using the SnapGene tool. 45,776 sequences containing SSRs were identified, among which the di- and trinucleotide motifs prevailed, together representing a total of 60% of the sequences identified. Of these, 3,493 sequences were present in the three transcripts analyzed, where 160 were selected based on the most significant difference in size (12-42) among the motifs, followed by the design of 30 SSRs. In the conventional PCR step, it was observed that 73% of the markers confirmed polymorphism between the BR 14-Mulato and IT85F-2687 accessions, while 57% were polymorphic between Pingo de Ouro and Santo Inácio. A total of 23 markers were transferable to V. unguiculata subsp. sesquipedalis and 16 for V. radiata (L.). Finally, the analysis allowed the identification of SSRs in the transcriptomes of cowpea, submitted to biotic and abiotic stresses, revealing polymorphisms between different accessions. The data identified, analyzed and validated will contribute to studies of genetic diversity and pre-breeding, aiming to broaden the available genetic base of cowpea.