Biomaterial Database

Bisulfite sequencing for cytosine-methylation analysis in plants. 2011

Nazmul Haque, and Masamichi Nishiguchi

Faculty of Agriculture, Ehime University, 790-8566 Matsuyama, Japan.

RNA silencing is a sequence-specific RNA degradation mechanism conserved in eukaryotes including fungi, plants, and animals. One of the three RNA silencing pathways is DNA methylation which is the result of interaction between DNA and siRNA, a hallmark of RNA silencing. Bisulfite sequencing can be very powerful for DNA methylation analysis in this context. This method includes DNA extraction, digestion of DNA with restriction enzyme, treatment of DNA with bisulfite, PCR amplification of DNA, cloning of amplified DNA fragments, sequencing of DNA fragments, and analysis of DNA sequences. Based on this method, increased levels of cytosine methylation were obtained in both symmetrical (CpG, CpNpG) and non-symmetrical (CpHpH) contexts in silenced lines of transgenic plants (CG>CNG>CHH), while the methylation levels were low in nonsilenced, over-expressing lines. Through grafting, RNA silencing was induced in the non-silenced scions from silenced rootstocks; however, the methylation level of DNA in the scions did not increase.

UI	MeSH Term	Description	Entries
D008969	Molecular Sequence Data	Descriptions of specific amino acid, carbohydrate, or nucleotide sequences which have appeared in the published literature and/or are deposited in and maintained by databanks such as GENBANK, European Molecular Biology Laboratory (EMBL), National Biomedical Research Foundation (NBRF), or other sequence repositories.	Sequence Data, Molecular,Molecular Sequencing Data,Data, Molecular Sequence,Data, Molecular Sequencing,Sequencing Data, Molecular
D003596	Cytosine	A pyrimidine base that is a fundamental unit of nucleic acids.
D001483	Base Sequence	The sequence of PURINES and PYRIMIDINES in nucleic acids and polynucleotides. It is also called nucleotide sequence.	DNA Sequence,Nucleotide Sequence,RNA Sequence,DNA Sequences,Base Sequences,Nucleotide Sequences,RNA Sequences,Sequence, Base,Sequence, DNA,Sequence, Nucleotide,Sequence, RNA,Sequences, Base,Sequences, DNA,Sequences, Nucleotide,Sequences, RNA
D013447	Sulfites	Inorganic salts of sulfurous acid.	Sulfite,Sulfites, Inorganic,Inorganic Sulfites
D016133	Polymerase Chain Reaction	In vitro method for producing large amounts of specific DNA or RNA fragments of defined length and sequence from small amounts of short oligonucleotide flanking sequences (primers). The essential steps include thermal denaturation of the double-stranded target molecules, annealing of the primers to their complementary sequences, and extension of the annealed primers by enzymatic synthesis with DNA polymerase. The reaction is efficient, specific, and extremely sensitive. Uses for the reaction include disease diagnosis, detection of difficult-to-isolate pathogens, mutation analysis, genetic testing, DNA sequencing, and analyzing evolutionary relationships.	Anchored PCR,Inverse PCR,Nested PCR,PCR,Anchored Polymerase Chain Reaction,Inverse Polymerase Chain Reaction,Nested Polymerase Chain Reaction,PCR, Anchored,PCR, Inverse,PCR, Nested,Polymerase Chain Reactions,Reaction, Polymerase Chain,Reactions, Polymerase Chain
D017343	Genes, Plant	The functional hereditary units of PLANTS.	Plant Genes,Gene, Plant,Plant Gene
D017422	Sequence Analysis, DNA	A multistage process that includes cloning, physical mapping, subcloning, determination of the DNA SEQUENCE, and information analysis.	DNA Sequence Analysis,Sequence Determination, DNA,Analysis, DNA Sequence,DNA Sequence Determination,DNA Sequence Determinations,DNA Sequencing,Determination, DNA Sequence,Determinations, DNA Sequence,Sequence Determinations, DNA,Analyses, DNA Sequence,DNA Sequence Analyses,Sequence Analyses, DNA,Sequencing, DNA
D019175	DNA Methylation	Addition of methyl groups to DNA. DNA methyltransferases (DNA methylases) perform this reaction using S-ADENOSYLMETHIONINE as the methyl group donor.	DNA Methylations,Methylation, DNA,Methylations, DNA
D030821	Plants, Genetically Modified	PLANTS, or their progeny, whose GENOME has been altered by GENETIC ENGINEERING.	Genetically Modified Plants,Plants, Transgenic,Transgenic Plants,GMO Plants,Genetically Engineered Plants,Engineered Plant, Genetically,Engineered Plants, Genetically,GMO Plant,Genetically Engineered Plant,Genetically Modified Plant,Modified Plant, Genetically,Modified Plants, Genetically,Plant, GMO,Plant, Genetically Engineered,Plant, Genetically Modified,Plant, Transgenic,Plants, GMO,Plants, Genetically Engineered,Transgenic Plant
D034622	RNA Interference	A gene silencing phenomenon whereby specific dsRNAs (RNA, DOUBLE-STRANDED) trigger the degradation of homologous mRNA (RNA, MESSENGER). The specific dsRNAs are processed into SMALL INTERFERING RNA (siRNA) which serves as a guide for cleavage of the homologous mRNA in the RNA-INDUCED SILENCING COMPLEX. DNA METHYLATION may also be triggered during this process.	Gene Silencing, Post-Transcriptional,Post-Transcriptional Gene Silencing,Co-Suppression,Cosuppression,Posttranscriptional Gene Silencing,RNA Silencing,RNAi,Co Suppression,Gene Silencing, Post Transcriptional,Gene Silencing, Posttranscriptional,Gene Silencings, Posttranscriptional,Interference, RNA,Post Transcriptional Gene Silencing,Post-Transcriptional Gene Silencings,Silencing, Post-Transcriptional Gene