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Epigenetic diversity of Saccharum spp. accessions assessed by methylation-sensitive amplification polymorphism (MSAP).

ABSTRACT: The epigenetic diversity of six genotype groups (commercial cultivars, S. officinarum, S. spontaneum, S. robustum, S. barberi, and Erianthus sp.) was assessed through methylation-sensitive amplification polymorphism (MSAP). A total of 1341 MSAP loci were analyzed, of which 1117 (83.29%) were susceptible to cytosine methylation and responsible for a higher proportion of overall diversity among genotypes. The MSAP selective primer combinations captured different proportions of internal and external cytosine methylation loci across genotype groups, while the average external cytosine frequency was higher for all genotype groups. The genotypes were divided into two subpopulations with a high differentiation index (φst = 0.086) based on epigenetic loci. The genotypes were clustered in three subgroups for both methylated and unmethylated loci, considering dissimilarity values. Four methylated fragments (MFs) were randomly selected and subsequently sequenced and compared with sugarcane public databases using BLASTN. MF alignments suggest that cytosine methylation occurs in sugarcane near CpG islands and tandem repeats within transcribed regions and putative cis-regulatory sequences, which assigned functions are associated with stress adaptation. These results provide the first insights about the distribution of this epigenetic mark in sugarcane genome, and suggest a biological relevance of methylated loci.

SUBMITTER: Martins AA

PROVIDER: S-EPMC7246279 | biostudies-literature |

REPOSITORIES: biostudies-literature

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Project description:Epigenetic mechanisms, including histone modifications and DNA methylation, mutually regulate chromatin structure, maintain genome integrity, and affect gene expression and transposon mobility. Variations in DNA methylation within plant populations, as well as methylation in response to internal and external factors, are of increasing interest, especially in the crop research field. Methylation Sensitive Amplification Polymorphism (MSAP) is one of the most commonly used methods for assessing DNA methylation changes in plants. This method involves gel-based visualization of PCR fragments from selectively amplified DNA that are cleaved using methylation-sensitive restriction enzymes. In this study, we developed and validated a new method based on the conventional MSAP approach called Methylation Sensitive Amplification Polymorphism Sequencing (MSAP-Seq). We improved the MSAP-based approach by replacing the conventional separation of amplicons on polyacrylamide gels with direct, high-throughput sequencing using Next Generation Sequencing (NGS) and automated data analysis. MSAP-Seq allows for global sequence-based identification of changes in DNA methylation. This technique was validated in Hordeum vulgare. However, MSAP-Seq can be straightforwardly implemented in different plant species, including crops with large, complex and highly repetitive genomes. The incorporation of high-throughput sequencing into MSAP-Seq enables parallel and direct analysis of DNA methylation in hundreds of thousands of sites across the genome. MSAP-Seq provides direct genomic localization of changes and enables quantitative evaluation. We have shown that the MSAP-Seq method specifically targets gene-containing regions and that a single analysis can cover three-quarters of all genes in large genomes. Moreover, MSAP-Seq's simplicity, cost effectiveness, and high-multiplexing capability make this method highly affordable. Therefore, MSAP-Seq can be used for DNA methylation analysis in crop plants with large and complex genomes.

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Epigenetic diversity of Saccharum spp. accessions assessed by methylation-sensitive amplification polymorphism (MSAP).

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