Genomic changes associated with somaclonal variation in banana (Musa spp.)

The molecular basis of somaclonal variation is not precisely known, but both genetic and epigenetic mechanisms have been proposed. The available evidence points toward the existence of labile portions of the genome that can be modulated when the cells undergo the stress of tissue culture. Therefore, the hypothesis that there are identifiable and predictable DNA markers for the early diagnosis of somaclonal variation has been tested. Representational difference analysis was used to isolate unique fragments of DNA (difference products) between visible culture-induced off-type and normal banana plants. Markers generated from six difference products differentiated between some of the off-type and normal pairs. The genomic region around one of these difference products has been extensively characterized and has a high degree of polymorphism, with variation in up to 10% of the nucleotides sequenced in the region. This same region has been shown to vary in other pairs of off-type and normal banana plants derived from tissue culture as well as in plants propagated commercially in vitro. The data are consistent with the hypothesis that there is at least one particularly labile portion of the genome that is especially susceptible to the stress imposed during tissue culture and that is associated with higher rearrangement and mutation rates than other portions of the genome. Consequently, the regions that are reported here have the potential to be used as early detection tools for identifying somaclonal variants

IRAP, a retrotransposon-based marker system for the detection of somaclonal variation in barley

The retrotransposon-based marker system, inter-retrotransposon amplified polymorphism (IRAP), and inter-simple sequence repeats (ISSRs) were used to detect somaclonal variation induced by tissue culture. IRAPs use a single primer designed to amplify out from the 5′ LTR sequence of the BARE-1 retrotransposon combined with a degenerate 3′ anchor, similar to that of ISSR primers. We analysed DNA polymorphisms in 147 primary regenerants and parental controls from three cultivars of barley (Hordeum vulgare). The ISSR marker system generated an average of 218 bands per primer, with 29 polymorphisms of which 12 were novel non-parental bands. In comparison, the IRAP system generated an average of 121 bands per primer, with 15 polymorphisms of which nine were novel non-parental bands. Polymorphism detected for IRAP and ISSR markers was more than twofold higher in Golden Promise than Mackay and Tallon cultivars. However, there was no significant difference in the frequency of novel non-parental bands. Cluster analysis revealed that the level of polymorphism and genetic variability detected was comparable between IRAP and ISSR markers. This suggests that retrotransposon-based marker systems, such as IRAP, based on retrotransposons such as BARE-1, are valuable tools for the detailed characterisation of mutation profiles that arise during tissue culture. Their use should improve our understanding of processes influencing mutation and somaclonal variation and allow for the design of methods that yield fewer genome changes in applications where maintaining clonal integrity is important