Molecular methods like (in situ) hybridization and PCR are routinely applied for identifying and monitoring microbes of interest and for studying the composition of polymicrobial communities in various environments, ranging from natural (including the human body) to biotechnological systems. For this purpose rRNA (genes) and/or functional marker genes are targeted. The accuracy of such diagnostic hybridization and PCR assays is intimately connected with specificity and coverage of the probes and primers used in these experiments. These parameters, however, require frequent re-evaluation due to the rapidly increasing sequence information in public databases.
probeCheck provides a freely accessible, central platform for rapid specificity and coverage evaluations of probes and primers against the latest sequence collections of selected phylogenetic and functional marker genes. Using a common user interface a number of established sequence databases can be queried.
The probeCheck server employs the ARB probe match tool, which creates difference alignments of the tested oligonucleotide and complementary sequences (up to five mismatching bases). Difference alignments are ordered according to the number, position and type of mismatching bases of non-target sequences. For each perfectly-matched and mismatched probe-target hybrid the free energy (ΔG) can be calculated using the two-state hybridization tool of the UNAfold software.
In addition, probeCheck is linked with probe match tools of the Ribosomal Database Project (RDP) II and Greengenes, Blast (search for short nearly exact matches), Oligonucleotide Properties Calculator (OligoCalc), the two-state folding tool of the DINAMelt server, and probeBase, thus offering a pivotal forum to different online probe/primer evaluation tools.
Databases behind probeCheck are regularly retrieved from public database projects such as RDP II, Greengenes, SILVA, and FGPR, or are kindly provided by individual curators.
Curators of own nucleic acid sequence databases are strongly encouraged to make their databases available for probe/primer evaluations on the probeCheck server. Note that probeCheck only enables matches against the database. The actual database remains hidden in the background and is NOT available for download. Please contact the probeCheck staff for further details.
Loy A, Arnold R, Tischler P, Rattei T, Wagner M, Horn M. 2008. probeCheck - a central resource for evaluating oligonucleotide probe coverage and specificity. Environ. Microbiol. 10: 2894-2896. PDF
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Last major update: July 2010
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