Marcadores moleculares baseados em retrotransposons e marcadores EST-SSR para a caracterização genética e análise da variação somaclonal de mandioca (Manihot esculenta Crantz)

Abstract

Current study establishes appropriate conditions for the clonal multiplication of cassava varieties (Manihot esculenta Crantz) with agronomic and industrial interest, free of the Cassava Common Mosaic Virus (CsCMV), and monitors the genetic stability of clones by molecular markers. Meristem-tips from seven cassava varieties (Baianinha, Fécula Branca, Icaraíma, IPR União, Olho junto, Pioneira e Tamboara) were employed to obtain the clones. They were inoculated in culture medium with different concentrations, types of auxin and different concentrations of 6- benzyladenine and gibberellin. The clones of the different varieties were established in culture media with different auxins. Texts PTA-ELISA and IC-RT-PCR (Immunocapture of CsCMV virions) were used for virus indexing. Twelve primers based on the LTR (Long Terminal Repeat) of retrotransposons were designed to monitor genetic stability: IRAP (Inter-Retrotransposon Amplified Polymorphism) and REMAP (Retrotransposon-Microssatellite Amplified Polymorphism), suitable for analyzing the DNA sequences of varieties Olho Junto, Fécula Branca, Tamboara and IPR-União. The REMAP and IRAP primers were employed with primers to a simple sequence repeat contained in expressed sequences of DNA (EST-SSR) in the cassava genome, to investigate the possibility of induction of somaclonal variation in the clones. The polymorphism in DNA sequences, amplified with primers IRAP and REMAP, was more than three times greater than the polymorphism detected by the EST-SSR primers in clones of the varieties Fécula Branca, Olho Junto and IPRUnião. Genetic variability was greater with the increasing number of subcultures. IRAP and REMAP primers designed for current study may be recommended to monitor the stability propagated in in vitro cassava clones.