PJB-2006-129
THE METHYLATION-SENSITIVE AMPLIFICATION POLYMORPHISM IN JUVENILE AND ADULT PHASE CRAB APPLE (MALUS MICROMALUS)
ISHFAQ A. HAFIZ, NADEEM A. ABBASI, AZHAR HUSSAIN, ABDUL WAHEED* AND S.M. SAQLAN NAQVI**
Abstract
Cytosine methylation has been implicated in regulation of gene expression, genomic imprinting and chromatin remodeling, resulting both in temporal and developmental regulation. Keeping in view its importance in development, this study was carried out to explore the methylation-sensitive amplification polymorphism (MSAP) associated with the transition from juvenile to adult phase in Malus micromalus. For detection of methylation in the genome of M. micromalus, a pair of restriction endo-nucleases HpaII-MspI was used. Genomic DNA from juvenile and adult phase leaves of the seedling tree, that was undigested (control) and digested with each enzyme HpaII and MspI, was amplified using selected primers. In total 72 bands were amplified with the help of 12 primers. Post amplification digestion of these bands with MspI or HpaII revealed 35 bands containing CCGG. Ten bands absent in amplified profiles from J phase digested DNA, appeared in amplified products from digested A phase DNA, indicating de novo methylation at CCGG. Five bands disappeared in A phase while these bands were present in J phase DNA amplified profiles, revealing presence of restriction site without methylation in A phase. These results are suggestive that demethylation may have occurred in A phase. Seventeen motifs of DNA methylation at CCGG remaining similar in both phases seem to have been maintained from basal to crown part of the seedling tree. Amplified profiles produced from restricted DNA from both phases showed polymorphism due to differential methylation.
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