title: methBLAST and methPrimerDB: web-tools for PCR based methylation analysis.
abstract: BACKGROUND
  DNA methylation plays an important role in development and tumorigenesis by epigenetic modification and silencing of critical genes. The development of PCR-based methylation assays on bisulphite modified DNA heralded a breakthrough in speed and sensitivity for gene methylation analysis. Despite this technological advancement, these approaches require a cumbersome gene by gene primer design and experimental validation. Bisulphite DNA modification results in sequence alterations (all unmethylated cytosines are converted into uracils) and a general sequence complexity reduction as cytosines become underrepresented. Consequently, standard BLAST sequence homology searches cannot be applied to search for specific methylation primers.
  
  
  RESULTS
  To address this problem we developed methBLAST, a sequence similarity search program, based on the original BLAST algorithm but querying in silico bisulphite modified genome sequences to evaluate oligonucleotide sequence similarities. Apart from the primer specificity analysis tool, we have also developed a public database termed methPrimerDB for the storage and retrieval of validated PCR based methylation assays. The web interface allows free public access to perform methBLAST searches or database queries and to submit user based information. Database records can be searched by gene symbol, nucleotide sequence, analytical method used, Entrez Gene or methPrimerDB identifier, and submitter's name. Each record contains a link to Entrez Gene and PubMed to retrieve additional information on the gene, its genomic context and the article in which the methylation assay was described. To assure and maintain data integrity and accuracy, the database is linked to other reference databases. Currently, the database contains primer records for the most popular PCR-based methylation analysis methods to study human, mouse and rat epigenetic modifications. methPrimerDB and methBLAST are available at http://medgen.ugent.be/methprimerdb and http://medgen.ugent.be/methblast.
  
  
  CONCLUSIONS
  We have developed two integrated and freely available web-tools for PCR based methylation analysis. methBLAST allows in silico assessment of primer specificity in PCR based methylation assays that can be stored in the methPrimerDB database, which provides a search portal for validated methylation assays.
authors.affiliation: Center for Medical Genetics, Ghent University Hospital, De Pintelaan 185, 9000 Ghent, Belgium. Filip.Pattyn@UGent.be <Filip.Pattyn@UGent.be>
authors.forename: Filip
authors.initials: F
authors.lastname: Pattyn
authors.forename: Jasmien
authors.initials: J
authors.lastname: Hoebeeck
authors.forename: Piet
authors.initials: P
authors.lastname: Robbrecht
authors.forename: Evi
authors.initials: E
authors.lastname: Michels
authors.forename: Anne
authors.initials: A
authors.lastname: De Paepe
authors.forename: Guy
authors.initials: G
authors.lastname: Bottu
authors.forename: David
authors.initials: D
authors.lastname: Coornaert
authors.forename: Robert
authors.initials: R
authors.lastname: Herzog
authors.forename: Frank
authors.initials: F
authors.lastname: Speleman
authors.forename: Jo
authors.initials: J
authors.lastname: Vandesompele
mesh_terms: D000465:Algorithms; D000818:Animals; D019295:Computational Biology / Q000379:methods*; D019175:DNA Methylation; D017931:DNA Primers / Q000737:chemistry; D030541:Databases, Genetic; D030562:Databases, Protein; D044127:Epigenesis, Genetic; D006801:Humans; D020407:Internet; D051379:Mice; D016133:Polymerase Chain Reaction / Q000379:methods*; D051381:Rats; D012984:Software; D013447:Sulfites / Q000737:chemistry
chemical_list: D017931:DNA Primers; D013447:Sulfites; C042345:hydrogen sulfite