Methylation-sensitive RFLPs: characterisation of two oil palm markers showing somaclonal variation-associated polymorphism

Abstract.

The occurrence of "mantled" somaclonal variants (approx. 5%), which display alterations in floral organ structure, among populations of somatic embryo-derived oil palms (Elaeis guineensis Jacq.) currently hampers any scaling-up of clonal plant micropropagation. As a first step towards the identification of abnormality-discriminating markers, we have screened a set of 27 oil palm cDNA probes for methylation-sensitive restriction fragment length polymorphisms (RFLPs) using callus genomic DNA digested with the isoschizomeric enzymes MspI and HpaII. Only two probes (CPHO62 and -63) were found to differentiate reproducibly in two different genotypic backgrounds between nodular compact calli (NCC) and fast-growing calli (FGC), which generate 5% and 100% "mantled" plantlets, respectively. Comparative analyses were then conducted on DNA from inflorescences and leaves of normal and abnormal adult regenerants. With both probes, the observed methylation patterns were strongly clone-dependent and monomorphic with respect to the phenotype of the regenerants, except for the type-specific banding pattern obtained with the CPHO62 probe on material from the LMC3 clonal offspring. The results presented here mirror the higher difference in genomic DNA methylation observed between normal and abnormal embryogenic calli when compared to more differentiated plant material. Moreover, they reinforce the paramount interest of NCC and FGC callus lines as a material of choice in the search for early epigenetic markers of the "mantled" somaclonal variation. The potential use of methylation-sensitive RFLPs for the early detection of somaclonal variation at early stages of the micropropagation process is discussed.