Patterns of cytosine methylation in an elite rice hybrid and its parental lines, detected by a methylation-sensitive amplification polymorphism technique |
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Authors: | L Z Xiong C G Xu M A Saghai Maroof and Qifa Zhang |
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Institution: | (1) National Key Laboratory of Crop Genetic Improvement Huazhong Agricultural University, Wuhan 430070, China e-mail: qifazh@public.wh.hb.cn; Fax: +86-27-87393392, CN;(2) Department of Crop and Soil Environmental Sciences Virginia Polytechnic Institute and State University Blacksburg, VA 24061, USA, US |
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Abstract: | DNA methylation is known to play an important role in the regulation of gene expression in eukaryotes. In this study, we
assessed the extent and pattern of cytosine methylation in the rice genome, using the technique of methylation-sensitive amplified
polymorphism (MSAP), which is a modification of the amplified fragment length polymorphism (AFLP) method that makes use of
the differential sensitivity of a pair of isoschizomers to cytosine methylation. The tissues assayed included seedlings and
flag leaves of an elite rice hybrid, Shanyou 63, and the parental lines Zhenshan 97 and Minghui 63. In all, 1076 fragments,
each representing a recognition site cleaved by either or both of the isoschizomers, were amplified using 16 pairs of selective
primers. A total of 195 sites were found to be methylated at cytosines in one or both parents, and the two parents showed
approximately the same overall degree of methylation (16.3%), as revealed by the incidence of differential digestion by the
isoschizomers. Four classes of patterns were identified in a comparative assay of cytosine methylation in the parents and
hybrid; increased methylation was detected in the hybrid compared to the parents at some of the recognition sites, while decreased
methylation in the hybrid was detected at other sites. A small proportion of the sites was found to be differentially methylated
in seedlings and flag leaves; DNA from young seedlings was methylated to a greater extent than that from flag leaves. Almost
all of the methylation patterns detected by MSAP could be confirmed by Southern analysis using the isolated amplified fragments
as probes. The results clearly demonstrate that the MSAP technique is highly efficient for large-scale detection of cytosine
methylation in the rice genome. We believe that the technique can be adapted for use in other plant species.
Received: 23 October 1998 / Accepted: 11 January 1999 |
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Keywords: | DNA methylation Hybrid rice Tissue specificity Isoschizomers PCR amplification |
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