DNA methylation changes after cryopreservation: MSAP analysis by multivariant statistical methods

Cryopreservation allows long-term conservation, and the genetic stability of the preserved material is one of its most important features compared to other conservation techniques. However, this stability has been questioned and in some cases epi-genetic instability after cryopreservation has been detected. Epigenetic information includes heritable signals that modulate gene expression but are not encoded in the primary nucleotide sequence. DNA methylation is the most extensively studied epigenetic modification and it has been connected to many crucial biological processes. The central issue of this work was to detect epigenetic changes in the cytosine methylation pattern throughout the whole cryopreservation process in an encapsulation-dehydration protocol in Mentha xpiperita apices. Although many techniques have been described for analysing DNA methylation, MSAP (methylation sensitive amplified polymorphism) is widely used. The global DNA methylation status has been stud ied after each step of the cryopreservation protocol. The effect of the addition during the procedure of vitamin E or ascorbic acid was also evaluated. AII the methylation variations detected were statistically analysed via multinomial logistic regression in order to test their significance. This study intends to introduce a new statistical approach to address methylation data in this contexto The results obtained showed and statistically proved that significant differences in the methylation pattern existed among the steps of the protocol.