关键词: DNA 甲基化, 大花蕙兰, 甲基化敏感扩增多态性(MSAP) , 子房发育

Abstract: The best known and most thoroughly studied epigenetic phenomena is DNA methylation, which plays an important role in regulating gene expression during plant regeneration and development. In this study, MSAP (methylation sensitive amplified polymorphism) technique was carried out to analyze differences of the methylation status between before and after pollination in ovaries of Cymbidium hybridium. 72 selective primer combinations were used to check the status of cytosine methylation DNA samples and a total of 5892 fragments were obtained . Thereinto, 748 fragments, each representing a recognition site cleaved by either or both of the isoschizomers ( Hpa Ⅱ and Msp Ⅰ ), were amplified , which were significant differences between the non-pollinated and the pollinated ovaries. The results demonstrated DNA methylation events occured in ovaries from Cymbidium hybridium . Both total and full methylation levels in the pollinated ovaries ( 11.4% ,7.8% ) were lower than those in the non-pollinated ovaries (14%, 9.5% ), which suggested some demethylations occurred. Furthermore, methylation patterns varied between the two ovaries. 14 types of MSAP patterns detected in the study belonged to two classes, type Ⅰ and Ⅱ. 25. 6% of detected fragments belonged to Type Ⅰ and 74.4% were type Ⅱ. Distinct patterns of DNA methylation arising through demethylation or de novo methylation might have specialized functions. This suggested the significance of epigenetic function in the development of orchid ovaries. The hypothesis that DNA methylation played a role in the C. hybridium ovary development would help to lucubrate the methylation status of special genes, clone the fragments with different methylation patterns, and shed novel insights into the molecular mechanisms of the ovary and floral development of orchids from the point of view of epigenetics.

Key words: DNA methylation