Reporter assay using the beta-galactosidase (lacZ) gene. The lacZ gene is typically fused to the promoter of interest. Differential regulation of the promoter mediated by the TF is assessed by induction of the system and evaluation of lacZ expression. Bacteria expressing lacZ appear blue when grown on a X-gal medium. The assay is often performed using a plasmid borne construction on a lacZ(def) strain.

The DNAse foot-printing method starts by focusing on a given region of interest (e.g. a promoter region) and amplifying it by PCR to obtain lots of sample. It then throws in the TF and then the DNAse. The mix is left to stir for a short time and then gel electrophoresis is run to compare the pattern of fragments in a control (no TF) and in the sample. If the TF has bound the sample, it will have protected a stretch of DNA (encompassing some fragments of the control) and thus those fragments will not appear in the sample gel. The fragments can then be cut-out from the gel, purified and sequenced to obtain the sequence of the protected region. This is often used to identify the binding motif of a TF for the first time. The foot-printing will typically resolve the protected region down to 50-100 bp, and the sequence can be then examined for possible TF-binding sites either by eye of using a computer search.

Electro-mobility shift-assays (or gel retardation assays) are a standard way of assessing TF-binding. A fragment of DNA of interest is amplified and labeled with a fluorophore. The fragment is left to incubate in a solution containing abundant TF and non-specific DNA (e.g. randomly cleaved DNA from salmon sperm, of all things) and then a gel is run with the incubated sample and a control (sample that has not been in contact with the TF). If the TF has bound the sample, the complex will migrate more slowly than unbound DNA through the gel, and this retarded band can be used as evidence of binding. The unspecific DNA ensures that the binding is specific to the fragment of interest and that any non-specific DNA-binding proteins left-over in the TF purification will bind there, instead of on the fragment of interest. EMSAs are typically carried out in a bunch of fragments, shown as multiple double (control+experiment) lanes in a wide picture. Certain additional controls are run in at least one of the fragments to ascertain specificity. In the most basic of these, specific competitor (the fragment of interest or a known positive control, unlabelled) is added to the reaction. This should sequester the TF and hence make the retardation band disappear, proving that the binding is indeed specific

Visual inspection of promoter region to identify putative binding sites based on previous knowledge

This is a weak form of in-silico search, in which the consensus sequence for the motif is compared to genomic positions and the number of mismatches (between candidate site and consensus) is used as a measure of site-quality.

A type of reporter assay . The idea is identical to a betagalactosidase assay, but here fluorescence of luciferase gene product is used as the reporter.

This is a weak form of in-silico search, in which the consensus sequence for the motif is compared to genomic positions and the number of mismatches (between candidate site and consensus) is used as a measure of site-quality.

Electro-mobility shift-assays (or gel retardation assays) are a standard way of assessing TF-binding. A fragment of DNA of interest is amplified and labeled with a fluorophore. The fragment is left to incubate in a solution containing abundant TF and non-specific DNA (e.g. randomly cleaved DNA from salmon sperm, of all things) and then a gel is run with the incubated sample and a control (sample that has not been in contact with the TF). If the TF has bound the sample, the complex will migrate more slowly than unbound DNA through the gel, and this retarded band can be used as evidence of binding. The unspecific DNA ensures that the binding is specific to the fragment of interest and that any non-specific DNA-binding proteins left-over in the TF purification will bind there, instead of on the fragment of interest. EMSAs are typically carried out in a bunch of fragments, shown as multiple double (control+experiment) lanes in a wide picture. Certain additional controls are run in at least one of the fragments to ascertain specificity. In the most basic of these, specific competitor (the fragment of interest or a known positive control, unlabelled) is added to the reaction. This should sequester the TF and hence make the retardation band disappear, proving that the binding is indeed specific

A type of reporter assay . The idea is identical to a betagalactosidase assay, but here fluorescence of luciferase gene product is used as the reporter.

This is a weak form of in-silico search, in which the consensus sequence for the motif is compared to genomic positions and the number of mismatches (between candidate site and consensus) is used as a measure of site-quality.

The DNAse foot-printing method starts by focusing on a given region of interest (e.g. a promoter region) and amplifying it by PCR to obtain lots of sample. It then throws in the TF and then the DNAse. The mix is left to stir for a short time and then gel electrophoresis is run to compare the pattern of fragments in a control (no TF) and in the sample. If the TF has bound the sample, it will have protected a stretch of DNA (encompassing some fragments of the control) and thus those fragments will not appear in the sample gel. The fragments can then be cut-out from the gel, purified and sequenced to obtain the sequence of the protected region. This is often used to identify the binding motif of a TF for the first time. The foot-printing will typically resolve the protected region down to 50-100 bp, and the sequence can be then examined for possible TF-binding sites either by eye of using a computer search.

Electro-mobility shift-assays (or gel retardation assays) are a standard way of assessing TF-binding. A fragment of DNA of interest is amplified and labeled with a fluorophore. The fragment is left to incubate in a solution containing abundant TF and non-specific DNA (e.g. randomly cleaved DNA from salmon sperm, of all things) and then a gel is run with the incubated sample and a control (sample that has not been in contact with the TF). If the TF has bound the sample, the complex will migrate more slowly than unbound DNA through the gel, and this retarded band can be used as evidence of binding. The unspecific DNA ensures that the binding is specific to the fragment of interest and that any non-specific DNA-binding proteins left-over in the TF purification will bind there, instead of on the fragment of interest. EMSAs are typically carried out in a bunch of fragments, shown as multiple double (control+experiment) lanes in a wide picture. Certain additional controls are run in at least one of the fragments to ascertain specificity. In the most basic of these, specific competitor (the fragment of interest or a known positive control, unlabelled) is added to the reaction. This should sequester the TF and hence make the retardation band disappear, proving that the binding is indeed specific

A type of reporter assay . The idea is identical to a betagalactosidase assay, but here fluorescence of luciferase gene product is used as the reporter.

Rapid Amplification of cDNA Ends is a technique to obtain the full length sequence of an RNA transcript. Most succinctly, the technique exploits the fact that PCR will stop at the ends of a mRNA. Basically, a primer mapping a "center" region of the mRNA is used to perform RT-PCR and terminal deoxynucleotidyl transferase (TdT) is used to add identical nucleotides to the 3' end of the cDNA. Standard PCR using another primer and the TdT-oligomer is performed in the other sense. The technique is frequently used to verify transcription start sites, which are relevant to the function (e.g. repression) of transcription factor binding sites.

Target-specific mutation, as opposed to non-specific mutation. In the context of TF-binding sites, site-directed mutagenesis is typically used to establish/confirm the specific sequence and location of a site, often in tandem with EMSA. Different positions of a putative binding site are mutated to non-consensus (or random) bases and binding to the mutated site is evaluated through EMSA or other means. Often implemented only in conserved motif positions or serially through all positions of a site.

As with motif discovery, TF-binding sites search benefits from a comparative genomics approach. Searching a single genome for TFBS will yield very noisy results. If a number of related genomes are searched, then the search results can be compared and strengthened by requiring that a site be located, for instance, in the promoter region of the same gene for at least two or three species. As in the case of motif discovery, these methods are not often applied to verify experimental results, but can be used to guide experimental research. For instance, comparative genomics searches can be implemented to detect good candidate sites, which are then verified using an experimental technique.

Electro-mobility shift-assays (or gel retardation assays) are a standard way of assessing TF-binding. A fragment of DNA of interest is amplified and labeled with a fluorophore. The fragment is left to incubate in a solution containing abundant TF and non-specific DNA (e.g. randomly cleaved DNA from salmon sperm, of all things) and then a gel is run with the incubated sample and a control (sample that has not been in contact with the TF). If the TF has bound the sample, the complex will migrate more slowly than unbound DNA through the gel, and this retarded band can be used as evidence of binding. The unspecific DNA ensures that the binding is specific to the fragment of interest and that any non-specific DNA-binding proteins left-over in the TF purification will bind there, instead of on the fragment of interest. EMSAs are typically carried out in a bunch of fragments, shown as multiple double (control+experiment) lanes in a wide picture. Certain additional controls are run in at least one of the fragments to ascertain specificity. In the most basic of these, specific competitor (the fragment of interest or a known positive control, unlabelled) is added to the reaction. This should sequester the TF and hence make the retardation band disappear, proving that the binding is indeed specific

A type of reporter assay . The idea is identical to a betagalactosidase assay, but here fluorescence of luciferase gene product is used as the reporter.

MSA can be used to align promoter regions and highlight regions of conservation that may be indicative of transcription factor binding sites.

Once the binding motif for a TF is known, this motif (which essentially defines a pattern) can be used to scan sequences in order to search for putative TF-binding site. This is useful, for instance, when trying to identify TF-binding site in ChIP-chip data. Searching for TF-binding site can be done in numerous ways. The most basic method is consensus search, in sequences are scored according to how many mismatches they have with the consensus sequence for the motif. A more elaborate way of searching involves using regular expressions, which allow to search for more loosely defined motifs [e.g. C(C/G)AT]. Common algorithms for this type of search include Pattern Locator and the DNA Pattern Find method of the SMS2 suite, but also some word processors. Finally, the mainstream way of conducting TF-binding site search is through the use of position-specific scoring matrices, which basically count the occurrences of each base at each position of the motif and use the inferred frequencies to score candidate sites. Algorithms in this last category include TFSEARCH, FITOM, CONSITE, TESS and MatInspector.

Electro-mobility shift-assays (or gel retardation assays) are a standard way of assessing TF-binding. A fragment of DNA of interest is amplified and labeled with a fluorophore. The fragment is left to incubate in a solution containing abundant TF and non-specific DNA (e.g. randomly cleaved DNA from salmon sperm, of all things) and then a gel is run with the incubated sample and a control (sample that has not been in contact with the TF). If the TF has bound the sample, the complex will migrate more slowly than unbound DNA through the gel, and this retarded band can be used as evidence of binding. The unspecific DNA ensures that the binding is specific to the fragment of interest and that any non-specific DNA-binding proteins left-over in the TF purification will bind there, instead of on the fragment of interest. EMSAs are typically carried out in a bunch of fragments, shown as multiple double (control+experiment) lanes in a wide picture. Certain additional controls are run in at least one of the fragments to ascertain specificity. In the most basic of these, specific competitor (the fragment of interest or a known positive control, unlabelled) is added to the reaction. This should sequester the TF and hence make the retardation band disappear, proving that the binding is indeed specific

A type of reporter assay . The idea is identical to a betagalactosidase assay, but here fluorescence of luciferase gene product is used as the reporter.

Once the binding motif for a TF is known, this motif (which essentially defines a pattern) can be used to scan sequences in order to search for putative TF-binding site. This is useful, for instance, when trying to identify TF-binding site in ChIP-chip data. Searching for TF-binding site can be done in numerous ways. The most basic method is consensus search, in sequences are scored according to how many mismatches they have with the consensus sequence for the motif. A more elaborate way of searching involves using regular expressions, which allow to search for more loosely defined motifs [e.g. C(C/G)AT]. Common algorithms for this type of search include Pattern Locator and the DNA Pattern Find method of the SMS2 suite, but also some word processors. Finally, the mainstream way of conducting TF-binding site search is through the use of position-specific scoring matrices, which basically count the occurrences of each base at each position of the motif and use the inferred frequencies to score candidate sites. Algorithms in this last category include TFSEARCH, FITOM, CONSITE, TESS and MatInspector.

As with motif discovery, TF-binding sites search benefits from a comparative genomics approach. Searching a single genome for TFBS will yield very noisy results. If a number of related genomes are searched, then the search results can be compared and strengthened by requiring that a site be located, for instance, in the promoter region of the same gene for at least two or three species. As in the case of motif discovery, these methods are not often applied to verify experimental results, but can be used to guide experimental research. For instance, comparative genomics searches can be implemented to detect good candidate sites, which are then verified using an experimental technique.

Electro-mobility shift-assays (or gel retardation assays) are a standard way of assessing TF-binding. A fragment of DNA of interest is amplified and labeled with a fluorophore. The fragment is left to incubate in a solution containing abundant TF and non-specific DNA (e.g. randomly cleaved DNA from salmon sperm, of all things) and then a gel is run with the incubated sample and a control (sample that has not been in contact with the TF). If the TF has bound the sample, the complex will migrate more slowly than unbound DNA through the gel, and this retarded band can be used as evidence of binding. The unspecific DNA ensures that the binding is specific to the fragment of interest and that any non-specific DNA-binding proteins left-over in the TF purification will bind there, instead of on the fragment of interest. EMSAs are typically carried out in a bunch of fragments, shown as multiple double (control+experiment) lanes in a wide picture. Certain additional controls are run in at least one of the fragments to ascertain specificity. In the most basic of these, specific competitor (the fragment of interest or a known positive control, unlabelled) is added to the reaction. This should sequester the TF and hence make the retardation band disappear, proving that the binding is indeed specific

A type of reporter assay . The idea is identical to a betagalactosidase assay, but here fluorescence of luciferase gene product is used as the reporter.

MSA can be used to align promoter regions and highlight regions of conservation that may be indicative of transcription factor binding sites.

Once the binding motif for a TF is known, this motif (which essentially defines a pattern) can be used to scan sequences in order to search for putative TF-binding site. This is useful, for instance, when trying to identify TF-binding site in ChIP-chip data. Searching for TF-binding site can be done in numerous ways. The most basic method is consensus search, in sequences are scored according to how many mismatches they have with the consensus sequence for the motif. A more elaborate way of searching involves using regular expressions, which allow to search for more loosely defined motifs [e.g. C(C/G)AT]. Common algorithms for this type of search include Pattern Locator and the DNA Pattern Find method of the SMS2 suite, but also some word processors. Finally, the mainstream way of conducting TF-binding site search is through the use of position-specific scoring matrices, which basically count the occurrences of each base at each position of the motif and use the inferred frequencies to score candidate sites. Algorithms in this last category include TFSEARCH, FITOM, CONSITE, TESS and MatInspector.

Electro-mobility shift-assays (or gel retardation assays) are a standard way of assessing TF-binding. A fragment of DNA of interest is amplified and labeled with a fluorophore. The fragment is left to incubate in a solution containing abundant TF and non-specific DNA (e.g. randomly cleaved DNA from salmon sperm, of all things) and then a gel is run with the incubated sample and a control (sample that has not been in contact with the TF). If the TF has bound the sample, the complex will migrate more slowly than unbound DNA through the gel, and this retarded band can be used as evidence of binding. The unspecific DNA ensures that the binding is specific to the fragment of interest and that any non-specific DNA-binding proteins left-over in the TF purification will bind there, instead of on the fragment of interest. EMSAs are typically carried out in a bunch of fragments, shown as multiple double (control+experiment) lanes in a wide picture. Certain additional controls are run in at least one of the fragments to ascertain specificity. In the most basic of these, specific competitor (the fragment of interest or a known positive control, unlabelled) is added to the reaction. This should sequester the TF and hence make the retardation band disappear, proving that the binding is indeed specific

A type of reporter assay . The idea is identical to a betagalactosidase assay, but here fluorescence of luciferase gene product is used as the reporter.

Once the binding motif for a TF is known, this motif (which essentially defines a pattern) can be used to scan sequences in order to search for putative TF-binding site. This is useful, for instance, when trying to identify TF-binding site in ChIP-chip data. Searching for TF-binding site can be done in numerous ways. The most basic method is consensus search, in sequences are scored according to how many mismatches they have with the consensus sequence for the motif. A more elaborate way of searching involves using regular expressions, which allow to search for more loosely defined motifs [e.g. C(C/G)AT]. Common algorithms for this type of search include Pattern Locator and the DNA Pattern Find method of the SMS2 suite, but also some word processors. Finally, the mainstream way of conducting TF-binding site search is through the use of position-specific scoring matrices, which basically count the occurrences of each base at each position of the motif and use the inferred frequencies to score candidate sites. Algorithms in this last category include TFSEARCH, FITOM, CONSITE, TESS and MatInspector.

Electro-mobility shift-assays (or gel retardation assays) are a standard way of assessing TF-binding. A fragment of DNA of interest is amplified and labeled with a fluorophore. The fragment is left to incubate in a solution containing abundant TF and non-specific DNA (e.g. randomly cleaved DNA from salmon sperm, of all things) and then a gel is run with the incubated sample and a control (sample that has not been in contact with the TF). If the TF has bound the sample, the complex will migrate more slowly than unbound DNA through the gel, and this retarded band can be used as evidence of binding. The unspecific DNA ensures that the binding is specific to the fragment of interest and that any non-specific DNA-binding proteins left-over in the TF purification will bind there, instead of on the fragment of interest. EMSAs are typically carried out in a bunch of fragments, shown as multiple double (control+experiment) lanes in a wide picture. Certain additional controls are run in at least one of the fragments to ascertain specificity. In the most basic of these, specific competitor (the fragment of interest or a known positive control, unlabelled) is added to the reaction. This should sequester the TF and hence make the retardation band disappear, proving that the binding is indeed specific

A type of reporter assay . The idea is identical to a betagalactosidase assay, but here fluorescence of luciferase gene product is used as the reporter.

Once the binding motif for a TF is known, this motif (which essentially defines a pattern) can be used to scan sequences in order to search for putative TF-binding site. This is useful, for instance, when trying to identify TF-binding site in ChIP-chip data. Searching for TF-binding site can be done in numerous ways. The most basic method is consensus search, in sequences are scored according to how many mismatches they have with the consensus sequence for the motif. A more elaborate way of searching involves using regular expressions, which allow to search for more loosely defined motifs [e.g. C(C/G)AT]. Common algorithms for this type of search include Pattern Locator and the DNA Pattern Find method of the SMS2 suite, but also some word processors. Finally, the mainstream way of conducting TF-binding site search is through the use of position-specific scoring matrices, which basically count the occurrences of each base at each position of the motif and use the inferred frequencies to score candidate sites. Algorithms in this last category include TFSEARCH, FITOM, CONSITE, TESS and MatInspector.

Electro-mobility shift-assays (or gel retardation assays) are a standard way of assessing TF-binding. A fragment of DNA of interest is amplified and labeled with a fluorophore. The fragment is left to incubate in a solution containing abundant TF and non-specific DNA (e.g. randomly cleaved DNA from salmon sperm, of all things) and then a gel is run with the incubated sample and a control (sample that has not been in contact with the TF). If the TF has bound the sample, the complex will migrate more slowly than unbound DNA through the gel, and this retarded band can be used as evidence of binding. The unspecific DNA ensures that the binding is specific to the fragment of interest and that any non-specific DNA-binding proteins left-over in the TF purification will bind there, instead of on the fragment of interest. EMSAs are typically carried out in a bunch of fragments, shown as multiple double (control+experiment) lanes in a wide picture. Certain additional controls are run in at least one of the fragments to ascertain specificity. In the most basic of these, specific competitor (the fragment of interest or a known positive control, unlabelled) is added to the reaction. This should sequester the TF and hence make the retardation band disappear, proving that the binding is indeed specific

A type of reporter assay . The idea is identical to a betagalactosidase assay, but here fluorescence of luciferase gene product is used as the reporter.

Once the binding motif for a TF is known, this motif (which essentially defines a pattern) can be used to scan sequences in order to search for putative TF-binding site. This is useful, for instance, when trying to identify TF-binding site in ChIP-chip data. Searching for TF-binding site can be done in numerous ways. The most basic method is consensus search, in sequences are scored according to how many mismatches they have with the consensus sequence for the motif. A more elaborate way of searching involves using regular expressions, which allow to search for more loosely defined motifs [e.g. C(C/G)AT]. Common algorithms for this type of search include Pattern Locator and the DNA Pattern Find method of the SMS2 suite, but also some word processors. Finally, the mainstream way of conducting TF-binding site search is through the use of position-specific scoring matrices, which basically count the occurrences of each base at each position of the motif and use the inferred frequencies to score candidate sites. Algorithms in this last category include TFSEARCH, FITOM, CONSITE, TESS and MatInspector.

As with motif discovery, TF-binding sites search benefits from a comparative genomics approach. Searching a single genome for TFBS will yield very noisy results. If a number of related genomes are searched, then the search results can be compared and strengthened by requiring that a site be located, for instance, in the promoter region of the same gene for at least two or three species. As in the case of motif discovery, these methods are not often applied to verify experimental results, but can be used to guide experimental research. For instance, comparative genomics searches can be implemented to detect good candidate sites, which are then verified using an experimental technique.

Electro-mobility shift-assays (or gel retardation assays) are a standard way of assessing TF-binding. A fragment of DNA of interest is amplified and labeled with a fluorophore. The fragment is left to incubate in a solution containing abundant TF and non-specific DNA (e.g. randomly cleaved DNA from salmon sperm, of all things) and then a gel is run with the incubated sample and a control (sample that has not been in contact with the TF). If the TF has bound the sample, the complex will migrate more slowly than unbound DNA through the gel, and this retarded band can be used as evidence of binding. The unspecific DNA ensures that the binding is specific to the fragment of interest and that any non-specific DNA-binding proteins left-over in the TF purification will bind there, instead of on the fragment of interest. EMSAs are typically carried out in a bunch of fragments, shown as multiple double (control+experiment) lanes in a wide picture. Certain additional controls are run in at least one of the fragments to ascertain specificity. In the most basic of these, specific competitor (the fragment of interest or a known positive control, unlabelled) is added to the reaction. This should sequester the TF and hence make the retardation band disappear, proving that the binding is indeed specific

A type of reporter assay . The idea is identical to a betagalactosidase assay, but here fluorescence of luciferase gene product is used as the reporter.

MSA can be used to align promoter regions and highlight regions of conservation that may be indicative of transcription factor binding sites.

Once the binding motif for a TF is known, this motif (which essentially defines a pattern) can be used to scan sequences in order to search for putative TF-binding site. This is useful, for instance, when trying to identify TF-binding site in ChIP-chip data. Searching for TF-binding site can be done in numerous ways. The most basic method is consensus search, in sequences are scored according to how many mismatches they have with the consensus sequence for the motif. A more elaborate way of searching involves using regular expressions, which allow to search for more loosely defined motifs [e.g. C(C/G)AT]. Common algorithms for this type of search include Pattern Locator and the DNA Pattern Find method of the SMS2 suite, but also some word processors. Finally, the mainstream way of conducting TF-binding site search is through the use of position-specific scoring matrices, which basically count the occurrences of each base at each position of the motif and use the inferred frequencies to score candidate sites. Algorithms in this last category include TFSEARCH, FITOM, CONSITE, TESS and MatInspector.

Electro-mobility shift-assays (or gel retardation assays) are a standard way of assessing TF-binding. A fragment of DNA of interest is amplified and labeled with a fluorophore. The fragment is left to incubate in a solution containing abundant TF and non-specific DNA (e.g. randomly cleaved DNA from salmon sperm, of all things) and then a gel is run with the incubated sample and a control (sample that has not been in contact with the TF). If the TF has bound the sample, the complex will migrate more slowly than unbound DNA through the gel, and this retarded band can be used as evidence of binding. The unspecific DNA ensures that the binding is specific to the fragment of interest and that any non-specific DNA-binding proteins left-over in the TF purification will bind there, instead of on the fragment of interest. EMSAs are typically carried out in a bunch of fragments, shown as multiple double (control+experiment) lanes in a wide picture. Certain additional controls are run in at least one of the fragments to ascertain specificity. In the most basic of these, specific competitor (the fragment of interest or a known positive control, unlabelled) is added to the reaction. This should sequester the TF and hence make the retardation band disappear, proving that the binding is indeed specific

A type of reporter assay . The idea is identical to a betagalactosidase assay, but here fluorescence of luciferase gene product is used as the reporter.

Once the binding motif for a TF is known, this motif (which essentially defines a pattern) can be used to scan sequences in order to search for putative TF-binding site. This is useful, for instance, when trying to identify TF-binding site in ChIP-chip data. Searching for TF-binding site can be done in numerous ways. The most basic method is consensus search, in sequences are scored according to how many mismatches they have with the consensus sequence for the motif. A more elaborate way of searching involves using regular expressions, which allow to search for more loosely defined motifs [e.g. C(C/G)AT]. Common algorithms for this type of search include Pattern Locator and the DNA Pattern Find method of the SMS2 suite, but also some word processors. Finally, the mainstream way of conducting TF-binding site search is through the use of position-specific scoring matrices, which basically count the occurrences of each base at each position of the motif and use the inferred frequencies to score candidate sites. Algorithms in this last category include TFSEARCH, FITOM, CONSITE, TESS and MatInspector.

As with motif discovery, TF-binding sites search benefits from a comparative genomics approach. Searching a single genome for TFBS will yield very noisy results. If a number of related genomes are searched, then the search results can be compared and strengthened by requiring that a site be located, for instance, in the promoter region of the same gene for at least two or three species. As in the case of motif discovery, these methods are not often applied to verify experimental results, but can be used to guide experimental research. For instance, comparative genomics searches can be implemented to detect good candidate sites, which are then verified using an experimental technique.

Electro-mobility shift-assays (or gel retardation assays) are a standard way of assessing TF-binding. A fragment of DNA of interest is amplified and labeled with a fluorophore. The fragment is left to incubate in a solution containing abundant TF and non-specific DNA (e.g. randomly cleaved DNA from salmon sperm, of all things) and then a gel is run with the incubated sample and a control (sample that has not been in contact with the TF). If the TF has bound the sample, the complex will migrate more slowly than unbound DNA through the gel, and this retarded band can be used as evidence of binding. The unspecific DNA ensures that the binding is specific to the fragment of interest and that any non-specific DNA-binding proteins left-over in the TF purification will bind there, instead of on the fragment of interest. EMSAs are typically carried out in a bunch of fragments, shown as multiple double (control+experiment) lanes in a wide picture. Certain additional controls are run in at least one of the fragments to ascertain specificity. In the most basic of these, specific competitor (the fragment of interest or a known positive control, unlabelled) is added to the reaction. This should sequester the TF and hence make the retardation band disappear, proving that the binding is indeed specific

MSA can be used to align promoter regions and highlight regions of conservation that may be indicative of transcription factor binding sites.

Once the binding motif for a TF is known, this motif (which essentially defines a pattern) can be used to scan sequences in order to search for putative TF-binding site. This is useful, for instance, when trying to identify TF-binding site in ChIP-chip data. Searching for TF-binding site can be done in numerous ways. The most basic method is consensus search, in sequences are scored according to how many mismatches they have with the consensus sequence for the motif. A more elaborate way of searching involves using regular expressions, which allow to search for more loosely defined motifs [e.g. C(C/G)AT]. Common algorithms for this type of search include Pattern Locator and the DNA Pattern Find method of the SMS2 suite, but also some word processors. Finally, the mainstream way of conducting TF-binding site search is through the use of position-specific scoring matrices, which basically count the occurrences of each base at each position of the motif and use the inferred frequencies to score candidate sites. Algorithms in this last category include TFSEARCH, FITOM, CONSITE, TESS and MatInspector.

Electro-mobility shift-assays (or gel retardation assays) are a standard way of assessing TF-binding. A fragment of DNA of interest is amplified and labeled with a fluorophore. The fragment is left to incubate in a solution containing abundant TF and non-specific DNA (e.g. randomly cleaved DNA from salmon sperm, of all things) and then a gel is run with the incubated sample and a control (sample that has not been in contact with the TF). If the TF has bound the sample, the complex will migrate more slowly than unbound DNA through the gel, and this retarded band can be used as evidence of binding. The unspecific DNA ensures that the binding is specific to the fragment of interest and that any non-specific DNA-binding proteins left-over in the TF purification will bind there, instead of on the fragment of interest. EMSAs are typically carried out in a bunch of fragments, shown as multiple double (control+experiment) lanes in a wide picture. Certain additional controls are run in at least one of the fragments to ascertain specificity. In the most basic of these, specific competitor (the fragment of interest or a known positive control, unlabelled) is added to the reaction. This should sequester the TF and hence make the retardation band disappear, proving that the binding is indeed specific

Once the binding motif for a TF is known, this motif (which essentially defines a pattern) can be used to scan sequences in order to search for putative TF-binding site. This is useful, for instance, when trying to identify TF-binding site in ChIP-chip data. Searching for TF-binding site can be done in numerous ways. The most basic method is consensus search, in sequences are scored according to how many mismatches they have with the consensus sequence for the motif. A more elaborate way of searching involves using regular expressions, which allow to search for more loosely defined motifs [e.g. C(C/G)AT]. Common algorithms for this type of search include Pattern Locator and the DNA Pattern Find method of the SMS2 suite, but also some word processors. Finally, the mainstream way of conducting TF-binding site search is through the use of position-specific scoring matrices, which basically count the occurrences of each base at each position of the motif and use the inferred frequencies to score candidate sites. Algorithms in this last category include TFSEARCH, FITOM, CONSITE, TESS and MatInspector.

Electro-mobility shift-assays (or gel retardation assays) are a standard way of assessing TF-binding. A fragment of DNA of interest is amplified and labeled with a fluorophore. The fragment is left to incubate in a solution containing abundant TF and non-specific DNA (e.g. randomly cleaved DNA from salmon sperm, of all things) and then a gel is run with the incubated sample and a control (sample that has not been in contact with the TF). If the TF has bound the sample, the complex will migrate more slowly than unbound DNA through the gel, and this retarded band can be used as evidence of binding. The unspecific DNA ensures that the binding is specific to the fragment of interest and that any non-specific DNA-binding proteins left-over in the TF purification will bind there, instead of on the fragment of interest. EMSAs are typically carried out in a bunch of fragments, shown as multiple double (control+experiment) lanes in a wide picture. Certain additional controls are run in at least one of the fragments to ascertain specificity. In the most basic of these, specific competitor (the fragment of interest or a known positive control, unlabelled) is added to the reaction. This should sequester the TF and hence make the retardation band disappear, proving that the binding is indeed specific

Electro-mobility shift-assays (or gel retardation assays) are a standard way of assessing TF-binding. A fragment of DNA of interest is amplified and labeled with a fluorophore. The fragment is left to incubate in a solution containing abundant TF and non-specific DNA (e.g. randomly cleaved DNA from salmon sperm, of all things) and then a gel is run with the incubated sample and a control (sample that has not been in contact with the TF). If the TF has bound the sample, the complex will migrate more slowly than unbound DNA through the gel, and this retarded band can be used as evidence of binding. The unspecific DNA ensures that the binding is specific to the fragment of interest and that any non-specific DNA-binding proteins left-over in the TF purification will bind there, instead of on the fragment of interest. EMSAs are typically carried out in a bunch of fragments, shown as multiple double (control+experiment) lanes in a wide picture. Certain additional controls are run in at least one of the fragments to ascertain specificity. In the most basic of these, specific competitor (the fragment of interest or a known positive control, unlabelled) is added to the reaction. This should sequester the TF and hence make the retardation band disappear, proving that the binding is indeed specific

Electro-mobility shift-assays (or gel retardation assays) are a standard way of assessing TF-binding. A fragment of DNA of interest is amplified and labeled with a fluorophore. The fragment is left to incubate in a solution containing abundant TF and non-specific DNA (e.g. randomly cleaved DNA from salmon sperm, of all things) and then a gel is run with the incubated sample and a control (sample that has not been in contact with the TF). If the TF has bound the sample, the complex will migrate more slowly than unbound DNA through the gel, and this retarded band can be used as evidence of binding. The unspecific DNA ensures that the binding is specific to the fragment of interest and that any non-specific DNA-binding proteins left-over in the TF purification will bind there, instead of on the fragment of interest. EMSAs are typically carried out in a bunch of fragments, shown as multiple double (control+experiment) lanes in a wide picture. Certain additional controls are run in at least one of the fragments to ascertain specificity. In the most basic of these, specific competitor (the fragment of interest or a known positive control, unlabelled) is added to the reaction. This should sequester the TF and hence make the retardation band disappear, proving that the binding is indeed specific

Electro-mobility shift-assays (or gel retardation assays) are a standard way of assessing TF-binding. A fragment of DNA of interest is amplified and labeled with a fluorophore. The fragment is left to incubate in a solution containing abundant TF and non-specific DNA (e.g. randomly cleaved DNA from salmon sperm, of all things) and then a gel is run with the incubated sample and a control (sample that has not been in contact with the TF). If the TF has bound the sample, the complex will migrate more slowly than unbound DNA through the gel, and this retarded band can be used as evidence of binding. The unspecific DNA ensures that the binding is specific to the fragment of interest and that any non-specific DNA-binding proteins left-over in the TF purification will bind there, instead of on the fragment of interest. EMSAs are typically carried out in a bunch of fragments, shown as multiple double (control+experiment) lanes in a wide picture. Certain additional controls are run in at least one of the fragments to ascertain specificity. In the most basic of these, specific competitor (the fragment of interest or a known positive control, unlabelled) is added to the reaction. This should sequester the TF and hence make the retardation band disappear, proving that the binding is indeed specific

A type of reporter assay . The idea is identical to a betagalactosidase assay, but here fluorescence of luciferase gene product is used as the reporter.

Once the binding motif for a TF is known, this motif (which essentially defines a pattern) can be used to scan sequences in order to search for putative TF-binding site. This is useful, for instance, when trying to identify TF-binding site in ChIP-chip data. Searching for TF-binding site can be done in numerous ways. The most basic method is consensus search, in sequences are scored according to how many mismatches they have with the consensus sequence for the motif. A more elaborate way of searching involves using regular expressions, which allow to search for more loosely defined motifs [e.g. C(C/G)AT]. Common algorithms for this type of search include Pattern Locator and the DNA Pattern Find method of the SMS2 suite, but also some word processors. Finally, the mainstream way of conducting TF-binding site search is through the use of position-specific scoring matrices, which basically count the occurrences of each base at each position of the motif and use the inferred frequencies to score candidate sites. Algorithms in this last category include TFSEARCH, FITOM, CONSITE, TESS and MatInspector.

Electro-mobility shift-assays (or gel retardation assays) are a standard way of assessing TF-binding. A fragment of DNA of interest is amplified and labeled with a fluorophore. The fragment is left to incubate in a solution containing abundant TF and non-specific DNA (e.g. randomly cleaved DNA from salmon sperm, of all things) and then a gel is run with the incubated sample and a control (sample that has not been in contact with the TF). If the TF has bound the sample, the complex will migrate more slowly than unbound DNA through the gel, and this retarded band can be used as evidence of binding. The unspecific DNA ensures that the binding is specific to the fragment of interest and that any non-specific DNA-binding proteins left-over in the TF purification will bind there, instead of on the fragment of interest. EMSAs are typically carried out in a bunch of fragments, shown as multiple double (control+experiment) lanes in a wide picture. Certain additional controls are run in at least one of the fragments to ascertain specificity. In the most basic of these, specific competitor (the fragment of interest or a known positive control, unlabelled) is added to the reaction. This should sequester the TF and hence make the retardation band disappear, proving that the binding is indeed specific

A type of reporter assay . The idea is identical to a betagalactosidase assay, but here fluorescence of luciferase gene product is used as the reporter.

Once the binding motif for a TF is known, this motif (which essentially defines a pattern) can be used to scan sequences in order to search for putative TF-binding site. This is useful, for instance, when trying to identify TF-binding site in ChIP-chip data. Searching for TF-binding site can be done in numerous ways. The most basic method is consensus search, in sequences are scored according to how many mismatches they have with the consensus sequence for the motif. A more elaborate way of searching involves using regular expressions, which allow to search for more loosely defined motifs [e.g. C(C/G)AT]. Common algorithms for this type of search include Pattern Locator and the DNA Pattern Find method of the SMS2 suite, but also some word processors. Finally, the mainstream way of conducting TF-binding site search is through the use of position-specific scoring matrices, which basically count the occurrences of each base at each position of the motif and use the inferred frequencies to score candidate sites. Algorithms in this last category include TFSEARCH, FITOM, CONSITE, TESS and MatInspector.

Electro-mobility shift-assays (or gel retardation assays) are a standard way of assessing TF-binding. A fragment of DNA of interest is amplified and labeled with a fluorophore. The fragment is left to incubate in a solution containing abundant TF and non-specific DNA (e.g. randomly cleaved DNA from salmon sperm, of all things) and then a gel is run with the incubated sample and a control (sample that has not been in contact with the TF). If the TF has bound the sample, the complex will migrate more slowly than unbound DNA through the gel, and this retarded band can be used as evidence of binding. The unspecific DNA ensures that the binding is specific to the fragment of interest and that any non-specific DNA-binding proteins left-over in the TF purification will bind there, instead of on the fragment of interest. EMSAs are typically carried out in a bunch of fragments, shown as multiple double (control+experiment) lanes in a wide picture. Certain additional controls are run in at least one of the fragments to ascertain specificity. In the most basic of these, specific competitor (the fragment of interest or a known positive control, unlabelled) is added to the reaction. This should sequester the TF and hence make the retardation band disappear, proving that the binding is indeed specific

A type of reporter assay . The idea is identical to a betagalactosidase assay, but here fluorescence of luciferase gene product is used as the reporter.

Once the binding motif for a TF is known, this motif (which essentially defines a pattern) can be used to scan sequences in order to search for putative TF-binding site. This is useful, for instance, when trying to identify TF-binding site in ChIP-chip data. Searching for TF-binding site can be done in numerous ways. The most basic method is consensus search, in sequences are scored according to how many mismatches they have with the consensus sequence for the motif. A more elaborate way of searching involves using regular expressions, which allow to search for more loosely defined motifs [e.g. C(C/G)AT]. Common algorithms for this type of search include Pattern Locator and the DNA Pattern Find method of the SMS2 suite, but also some word processors. Finally, the mainstream way of conducting TF-binding site search is through the use of position-specific scoring matrices, which basically count the occurrences of each base at each position of the motif and use the inferred frequencies to score candidate sites. Algorithms in this last category include TFSEARCH, FITOM, CONSITE, TESS and MatInspector.

Electro-mobility shift-assays (or gel retardation assays) are a standard way of assessing TF-binding. A fragment of DNA of interest is amplified and labeled with a fluorophore. The fragment is left to incubate in a solution containing abundant TF and non-specific DNA (e.g. randomly cleaved DNA from salmon sperm, of all things) and then a gel is run with the incubated sample and a control (sample that has not been in contact with the TF). If the TF has bound the sample, the complex will migrate more slowly than unbound DNA through the gel, and this retarded band can be used as evidence of binding. The unspecific DNA ensures that the binding is specific to the fragment of interest and that any non-specific DNA-binding proteins left-over in the TF purification will bind there, instead of on the fragment of interest. EMSAs are typically carried out in a bunch of fragments, shown as multiple double (control+experiment) lanes in a wide picture. Certain additional controls are run in at least one of the fragments to ascertain specificity. In the most basic of these, specific competitor (the fragment of interest or a known positive control, unlabelled) is added to the reaction. This should sequester the TF and hence make the retardation band disappear, proving that the binding is indeed specific

A type of reporter assay . The idea is identical to a betagalactosidase assay, but here fluorescence of luciferase gene product is used as the reporter.

Once the binding motif for a TF is known, this motif (which essentially defines a pattern) can be used to scan sequences in order to search for putative TF-binding site. This is useful, for instance, when trying to identify TF-binding site in ChIP-chip data. Searching for TF-binding site can be done in numerous ways. The most basic method is consensus search, in sequences are scored according to how many mismatches they have with the consensus sequence for the motif. A more elaborate way of searching involves using regular expressions, which allow to search for more loosely defined motifs [e.g. C(C/G)AT]. Common algorithms for this type of search include Pattern Locator and the DNA Pattern Find method of the SMS2 suite, but also some word processors. Finally, the mainstream way of conducting TF-binding site search is through the use of position-specific scoring matrices, which basically count the occurrences of each base at each position of the motif and use the inferred frequencies to score candidate sites. Algorithms in this last category include TFSEARCH, FITOM, CONSITE, TESS and MatInspector.

Electro-mobility shift-assays (or gel retardation assays) are a standard way of assessing TF-binding. A fragment of DNA of interest is amplified and labeled with a fluorophore. The fragment is left to incubate in a solution containing abundant TF and non-specific DNA (e.g. randomly cleaved DNA from salmon sperm, of all things) and then a gel is run with the incubated sample and a control (sample that has not been in contact with the TF). If the TF has bound the sample, the complex will migrate more slowly than unbound DNA through the gel, and this retarded band can be used as evidence of binding. The unspecific DNA ensures that the binding is specific to the fragment of interest and that any non-specific DNA-binding proteins left-over in the TF purification will bind there, instead of on the fragment of interest. EMSAs are typically carried out in a bunch of fragments, shown as multiple double (control+experiment) lanes in a wide picture. Certain additional controls are run in at least one of the fragments to ascertain specificity. In the most basic of these, specific competitor (the fragment of interest or a known positive control, unlabelled) is added to the reaction. This should sequester the TF and hence make the retardation band disappear, proving that the binding is indeed specific

A type of reporter assay . The idea is identical to a betagalactosidase assay, but here fluorescence of luciferase gene product is used as the reporter.

Once the binding motif for a TF is known, this motif (which essentially defines a pattern) can be used to scan sequences in order to search for putative TF-binding site. This is useful, for instance, when trying to identify TF-binding site in ChIP-chip data. Searching for TF-binding site can be done in numerous ways. The most basic method is consensus search, in sequences are scored according to how many mismatches they have with the consensus sequence for the motif. A more elaborate way of searching involves using regular expressions, which allow to search for more loosely defined motifs [e.g. C(C/G)AT]. Common algorithms for this type of search include Pattern Locator and the DNA Pattern Find method of the SMS2 suite, but also some word processors. Finally, the mainstream way of conducting TF-binding site search is through the use of position-specific scoring matrices, which basically count the occurrences of each base at each position of the motif and use the inferred frequencies to score candidate sites. Algorithms in this last category include TFSEARCH, FITOM, CONSITE, TESS and MatInspector.

Electro-mobility shift-assays (or gel retardation assays) are a standard way of assessing TF-binding. A fragment of DNA of interest is amplified and labeled with a fluorophore. The fragment is left to incubate in a solution containing abundant TF and non-specific DNA (e.g. randomly cleaved DNA from salmon sperm, of all things) and then a gel is run with the incubated sample and a control (sample that has not been in contact with the TF). If the TF has bound the sample, the complex will migrate more slowly than unbound DNA through the gel, and this retarded band can be used as evidence of binding. The unspecific DNA ensures that the binding is specific to the fragment of interest and that any non-specific DNA-binding proteins left-over in the TF purification will bind there, instead of on the fragment of interest. EMSAs are typically carried out in a bunch of fragments, shown as multiple double (control+experiment) lanes in a wide picture. Certain additional controls are run in at least one of the fragments to ascertain specificity. In the most basic of these, specific competitor (the fragment of interest or a known positive control, unlabelled) is added to the reaction. This should sequester the TF and hence make the retardation band disappear, proving that the binding is indeed specific

A type of reporter assay . The idea is identical to a betagalactosidase assay, but here fluorescence of luciferase gene product is used as the reporter.

Once the binding motif for a TF is known, this motif (which essentially defines a pattern) can be used to scan sequences in order to search for putative TF-binding site. This is useful, for instance, when trying to identify TF-binding site in ChIP-chip data. Searching for TF-binding site can be done in numerous ways. The most basic method is consensus search, in sequences are scored according to how many mismatches they have with the consensus sequence for the motif. A more elaborate way of searching involves using regular expressions, which allow to search for more loosely defined motifs [e.g. C(C/G)AT]. Common algorithms for this type of search include Pattern Locator and the DNA Pattern Find method of the SMS2 suite, but also some word processors. Finally, the mainstream way of conducting TF-binding site search is through the use of position-specific scoring matrices, which basically count the occurrences of each base at each position of the motif and use the inferred frequencies to score candidate sites. Algorithms in this last category include TFSEARCH, FITOM, CONSITE, TESS and MatInspector.

Electro-mobility shift-assays (or gel retardation assays) are a standard way of assessing TF-binding. A fragment of DNA of interest is amplified and labeled with a fluorophore. The fragment is left to incubate in a solution containing abundant TF and non-specific DNA (e.g. randomly cleaved DNA from salmon sperm, of all things) and then a gel is run with the incubated sample and a control (sample that has not been in contact with the TF). If the TF has bound the sample, the complex will migrate more slowly than unbound DNA through the gel, and this retarded band can be used as evidence of binding. The unspecific DNA ensures that the binding is specific to the fragment of interest and that any non-specific DNA-binding proteins left-over in the TF purification will bind there, instead of on the fragment of interest. EMSAs are typically carried out in a bunch of fragments, shown as multiple double (control+experiment) lanes in a wide picture. Certain additional controls are run in at least one of the fragments to ascertain specificity. In the most basic of these, specific competitor (the fragment of interest or a known positive control, unlabelled) is added to the reaction. This should sequester the TF and hence make the retardation band disappear, proving that the binding is indeed specific

A type of reporter assay . The idea is identical to a betagalactosidase assay, but here fluorescence of luciferase gene product is used as the reporter.

Once the binding motif for a TF is known, this motif (which essentially defines a pattern) can be used to scan sequences in order to search for putative TF-binding site. This is useful, for instance, when trying to identify TF-binding site in ChIP-chip data. Searching for TF-binding site can be done in numerous ways. The most basic method is consensus search, in sequences are scored according to how many mismatches they have with the consensus sequence for the motif. A more elaborate way of searching involves using regular expressions, which allow to search for more loosely defined motifs [e.g. C(C/G)AT]. Common algorithms for this type of search include Pattern Locator and the DNA Pattern Find method of the SMS2 suite, but also some word processors. Finally, the mainstream way of conducting TF-binding site search is through the use of position-specific scoring matrices, which basically count the occurrences of each base at each position of the motif and use the inferred frequencies to score candidate sites. Algorithms in this last category include TFSEARCH, FITOM, CONSITE, TESS and MatInspector.

Electro-mobility shift-assays (or gel retardation assays) are a standard way of assessing TF-binding. A fragment of DNA of interest is amplified and labeled with a fluorophore. The fragment is left to incubate in a solution containing abundant TF and non-specific DNA (e.g. randomly cleaved DNA from salmon sperm, of all things) and then a gel is run with the incubated sample and a control (sample that has not been in contact with the TF). If the TF has bound the sample, the complex will migrate more slowly than unbound DNA through the gel, and this retarded band can be used as evidence of binding. The unspecific DNA ensures that the binding is specific to the fragment of interest and that any non-specific DNA-binding proteins left-over in the TF purification will bind there, instead of on the fragment of interest. EMSAs are typically carried out in a bunch of fragments, shown as multiple double (control+experiment) lanes in a wide picture. Certain additional controls are run in at least one of the fragments to ascertain specificity. In the most basic of these, specific competitor (the fragment of interest or a known positive control, unlabelled) is added to the reaction. This should sequester the TF and hence make the retardation band disappear, proving that the binding is indeed specific

A type of reporter assay . The idea is identical to a betagalactosidase assay, but here fluorescence of luciferase gene product is used as the reporter.

Once the binding motif for a TF is known, this motif (which essentially defines a pattern) can be used to scan sequences in order to search for putative TF-binding site. This is useful, for instance, when trying to identify TF-binding site in ChIP-chip data. Searching for TF-binding site can be done in numerous ways. The most basic method is consensus search, in sequences are scored according to how many mismatches they have with the consensus sequence for the motif. A more elaborate way of searching involves using regular expressions, which allow to search for more loosely defined motifs [e.g. C(C/G)AT]. Common algorithms for this type of search include Pattern Locator and the DNA Pattern Find method of the SMS2 suite, but also some word processors. Finally, the mainstream way of conducting TF-binding site search is through the use of position-specific scoring matrices, which basically count the occurrences of each base at each position of the motif and use the inferred frequencies to score candidate sites. Algorithms in this last category include TFSEARCH, FITOM, CONSITE, TESS and MatInspector.

Electro-mobility shift-assays (or gel retardation assays) are a standard way of assessing TF-binding. A fragment of DNA of interest is amplified and labeled with a fluorophore. The fragment is left to incubate in a solution containing abundant TF and non-specific DNA (e.g. randomly cleaved DNA from salmon sperm, of all things) and then a gel is run with the incubated sample and a control (sample that has not been in contact with the TF). If the TF has bound the sample, the complex will migrate more slowly than unbound DNA through the gel, and this retarded band can be used as evidence of binding. The unspecific DNA ensures that the binding is specific to the fragment of interest and that any non-specific DNA-binding proteins left-over in the TF purification will bind there, instead of on the fragment of interest. EMSAs are typically carried out in a bunch of fragments, shown as multiple double (control+experiment) lanes in a wide picture. Certain additional controls are run in at least one of the fragments to ascertain specificity. In the most basic of these, specific competitor (the fragment of interest or a known positive control, unlabelled) is added to the reaction. This should sequester the TF and hence make the retardation band disappear, proving that the binding is indeed specific

A type of reporter assay . The idea is identical to a betagalactosidase assay, but here fluorescence of luciferase gene product is used as the reporter.

Once the binding motif for a TF is known, this motif (which essentially defines a pattern) can be used to scan sequences in order to search for putative TF-binding site. This is useful, for instance, when trying to identify TF-binding site in ChIP-chip data. Searching for TF-binding site can be done in numerous ways. The most basic method is consensus search, in sequences are scored according to how many mismatches they have with the consensus sequence for the motif. A more elaborate way of searching involves using regular expressions, which allow to search for more loosely defined motifs [e.g. C(C/G)AT]. Common algorithms for this type of search include Pattern Locator and the DNA Pattern Find method of the SMS2 suite, but also some word processors. Finally, the mainstream way of conducting TF-binding site search is through the use of position-specific scoring matrices, which basically count the occurrences of each base at each position of the motif and use the inferred frequencies to score candidate sites. Algorithms in this last category include TFSEARCH, FITOM, CONSITE, TESS and MatInspector.

Electro-mobility shift-assays (or gel retardation assays) are a standard way of assessing TF-binding. A fragment of DNA of interest is amplified and labeled with a fluorophore. The fragment is left to incubate in a solution containing abundant TF and non-specific DNA (e.g. randomly cleaved DNA from salmon sperm, of all things) and then a gel is run with the incubated sample and a control (sample that has not been in contact with the TF). If the TF has bound the sample, the complex will migrate more slowly than unbound DNA through the gel, and this retarded band can be used as evidence of binding. The unspecific DNA ensures that the binding is specific to the fragment of interest and that any non-specific DNA-binding proteins left-over in the TF purification will bind there, instead of on the fragment of interest. EMSAs are typically carried out in a bunch of fragments, shown as multiple double (control+experiment) lanes in a wide picture. Certain additional controls are run in at least one of the fragments to ascertain specificity. In the most basic of these, specific competitor (the fragment of interest or a known positive control, unlabelled) is added to the reaction. This should sequester the TF and hence make the retardation band disappear, proving that the binding is indeed specific

A type of reporter assay . The idea is identical to a betagalactosidase assay, but here fluorescence of luciferase gene product is used as the reporter.

Once the binding motif for a TF is known, this motif (which essentially defines a pattern) can be used to scan sequences in order to search for putative TF-binding site. This is useful, for instance, when trying to identify TF-binding site in ChIP-chip data. Searching for TF-binding site can be done in numerous ways. The most basic method is consensus search, in sequences are scored according to how many mismatches they have with the consensus sequence for the motif. A more elaborate way of searching involves using regular expressions, which allow to search for more loosely defined motifs [e.g. C(C/G)AT]. Common algorithms for this type of search include Pattern Locator and the DNA Pattern Find method of the SMS2 suite, but also some word processors. Finally, the mainstream way of conducting TF-binding site search is through the use of position-specific scoring matrices, which basically count the occurrences of each base at each position of the motif and use the inferred frequencies to score candidate sites. Algorithms in this last category include TFSEARCH, FITOM, CONSITE, TESS and MatInspector.

Electro-mobility shift-assays (or gel retardation assays) are a standard way of assessing TF-binding. A fragment of DNA of interest is amplified and labeled with a fluorophore. The fragment is left to incubate in a solution containing abundant TF and non-specific DNA (e.g. randomly cleaved DNA from salmon sperm, of all things) and then a gel is run with the incubated sample and a control (sample that has not been in contact with the TF). If the TF has bound the sample, the complex will migrate more slowly than unbound DNA through the gel, and this retarded band can be used as evidence of binding. The unspecific DNA ensures that the binding is specific to the fragment of interest and that any non-specific DNA-binding proteins left-over in the TF purification will bind there, instead of on the fragment of interest. EMSAs are typically carried out in a bunch of fragments, shown as multiple double (control+experiment) lanes in a wide picture. Certain additional controls are run in at least one of the fragments to ascertain specificity. In the most basic of these, specific competitor (the fragment of interest or a known positive control, unlabelled) is added to the reaction. This should sequester the TF and hence make the retardation band disappear, proving that the binding is indeed specific

A type of reporter assay . The idea is identical to a betagalactosidase assay, but here fluorescence of luciferase gene product is used as the reporter.

Once the binding motif for a TF is known, this motif (which essentially defines a pattern) can be used to scan sequences in order to search for putative TF-binding site. This is useful, for instance, when trying to identify TF-binding site in ChIP-chip data. Searching for TF-binding site can be done in numerous ways. The most basic method is consensus search, in sequences are scored according to how many mismatches they have with the consensus sequence for the motif. A more elaborate way of searching involves using regular expressions, which allow to search for more loosely defined motifs [e.g. C(C/G)AT]. Common algorithms for this type of search include Pattern Locator and the DNA Pattern Find method of the SMS2 suite, but also some word processors. Finally, the mainstream way of conducting TF-binding site search is through the use of position-specific scoring matrices, which basically count the occurrences of each base at each position of the motif and use the inferred frequencies to score candidate sites. Algorithms in this last category include TFSEARCH, FITOM, CONSITE, TESS and MatInspector.

As with motif discovery, TF-binding sites search benefits from a comparative genomics approach. Searching a single genome for TFBS will yield very noisy results. If a number of related genomes are searched, then the search results can be compared and strengthened by requiring that a site be located, for instance, in the promoter region of the same gene for at least two or three species. As in the case of motif discovery, these methods are not often applied to verify experimental results, but can be used to guide experimental research. For instance, comparative genomics searches can be implemented to detect good candidate sites, which are then verified using an experimental technique.

Electro-mobility shift-assays (or gel retardation assays) are a standard way of assessing TF-binding. A fragment of DNA of interest is amplified and labeled with a fluorophore. The fragment is left to incubate in a solution containing abundant TF and non-specific DNA (e.g. randomly cleaved DNA from salmon sperm, of all things) and then a gel is run with the incubated sample and a control (sample that has not been in contact with the TF). If the TF has bound the sample, the complex will migrate more slowly than unbound DNA through the gel, and this retarded band can be used as evidence of binding. The unspecific DNA ensures that the binding is specific to the fragment of interest and that any non-specific DNA-binding proteins left-over in the TF purification will bind there, instead of on the fragment of interest. EMSAs are typically carried out in a bunch of fragments, shown as multiple double (control+experiment) lanes in a wide picture. Certain additional controls are run in at least one of the fragments to ascertain specificity. In the most basic of these, specific competitor (the fragment of interest or a known positive control, unlabelled) is added to the reaction. This should sequester the TF and hence make the retardation band disappear, proving that the binding is indeed specific

MSA can be used to align promoter regions and highlight regions of conservation that may be indicative of transcription factor binding sites.

Once the binding motif for a TF is known, this motif (which essentially defines a pattern) can be used to scan sequences in order to search for putative TF-binding site. This is useful, for instance, when trying to identify TF-binding site in ChIP-chip data. Searching for TF-binding site can be done in numerous ways. The most basic method is consensus search, in sequences are scored according to how many mismatches they have with the consensus sequence for the motif. A more elaborate way of searching involves using regular expressions, which allow to search for more loosely defined motifs [e.g. C(C/G)AT]. Common algorithms for this type of search include Pattern Locator and the DNA Pattern Find method of the SMS2 suite, but also some word processors. Finally, the mainstream way of conducting TF-binding site search is through the use of position-specific scoring matrices, which basically count the occurrences of each base at each position of the motif and use the inferred frequencies to score candidate sites. Algorithms in this last category include TFSEARCH, FITOM, CONSITE, TESS and MatInspector.

As with motif discovery, TF-binding sites search benefits from a comparative genomics approach. Searching a single genome for TFBS will yield very noisy results. If a number of related genomes are searched, then the search results can be compared and strengthened by requiring that a site be located, for instance, in the promoter region of the same gene for at least two or three species. As in the case of motif discovery, these methods are not often applied to verify experimental results, but can be used to guide experimental research. For instance, comparative genomics searches can be implemented to detect good candidate sites, which are then verified using an experimental technique.

Electro-mobility shift-assays (or gel retardation assays) are a standard way of assessing TF-binding. A fragment of DNA of interest is amplified and labeled with a fluorophore. The fragment is left to incubate in a solution containing abundant TF and non-specific DNA (e.g. randomly cleaved DNA from salmon sperm, of all things) and then a gel is run with the incubated sample and a control (sample that has not been in contact with the TF). If the TF has bound the sample, the complex will migrate more slowly than unbound DNA through the gel, and this retarded band can be used as evidence of binding. The unspecific DNA ensures that the binding is specific to the fragment of interest and that any non-specific DNA-binding proteins left-over in the TF purification will bind there, instead of on the fragment of interest. EMSAs are typically carried out in a bunch of fragments, shown as multiple double (control+experiment) lanes in a wide picture. Certain additional controls are run in at least one of the fragments to ascertain specificity. In the most basic of these, specific competitor (the fragment of interest or a known positive control, unlabelled) is added to the reaction. This should sequester the TF and hence make the retardation band disappear, proving that the binding is indeed specific

MSA can be used to align promoter regions and highlight regions of conservation that may be indicative of transcription factor binding sites.

Once the binding motif for a TF is known, this motif (which essentially defines a pattern) can be used to scan sequences in order to search for putative TF-binding site. This is useful, for instance, when trying to identify TF-binding site in ChIP-chip data. Searching for TF-binding site can be done in numerous ways. The most basic method is consensus search, in sequences are scored according to how many mismatches they have with the consensus sequence for the motif. A more elaborate way of searching involves using regular expressions, which allow to search for more loosely defined motifs [e.g. C(C/G)AT]. Common algorithms for this type of search include Pattern Locator and the DNA Pattern Find method of the SMS2 suite, but also some word processors. Finally, the mainstream way of conducting TF-binding site search is through the use of position-specific scoring matrices, which basically count the occurrences of each base at each position of the motif and use the inferred frequencies to score candidate sites. Algorithms in this last category include TFSEARCH, FITOM, CONSITE, TESS and MatInspector.

As with motif discovery, TF-binding sites search benefits from a comparative genomics approach. Searching a single genome for TFBS will yield very noisy results. If a number of related genomes are searched, then the search results can be compared and strengthened by requiring that a site be located, for instance, in the promoter region of the same gene for at least two or three species. As in the case of motif discovery, these methods are not often applied to verify experimental results, but can be used to guide experimental research. For instance, comparative genomics searches can be implemented to detect good candidate sites, which are then verified using an experimental technique.

Electro-mobility shift-assays (or gel retardation assays) are a standard way of assessing TF-binding. A fragment of DNA of interest is amplified and labeled with a fluorophore. The fragment is left to incubate in a solution containing abundant TF and non-specific DNA (e.g. randomly cleaved DNA from salmon sperm, of all things) and then a gel is run with the incubated sample and a control (sample that has not been in contact with the TF). If the TF has bound the sample, the complex will migrate more slowly than unbound DNA through the gel, and this retarded band can be used as evidence of binding. The unspecific DNA ensures that the binding is specific to the fragment of interest and that any non-specific DNA-binding proteins left-over in the TF purification will bind there, instead of on the fragment of interest. EMSAs are typically carried out in a bunch of fragments, shown as multiple double (control+experiment) lanes in a wide picture. Certain additional controls are run in at least one of the fragments to ascertain specificity. In the most basic of these, specific competitor (the fragment of interest or a known positive control, unlabelled) is added to the reaction. This should sequester the TF and hence make the retardation band disappear, proving that the binding is indeed specific

MSA can be used to align promoter regions and highlight regions of conservation that may be indicative of transcription factor binding sites.

Once the binding motif for a TF is known, this motif (which essentially defines a pattern) can be used to scan sequences in order to search for putative TF-binding site. This is useful, for instance, when trying to identify TF-binding site in ChIP-chip data. Searching for TF-binding site can be done in numerous ways. The most basic method is consensus search, in sequences are scored according to how many mismatches they have with the consensus sequence for the motif. A more elaborate way of searching involves using regular expressions, which allow to search for more loosely defined motifs [e.g. C(C/G)AT]. Common algorithms for this type of search include Pattern Locator and the DNA Pattern Find method of the SMS2 suite, but also some word processors. Finally, the mainstream way of conducting TF-binding site search is through the use of position-specific scoring matrices, which basically count the occurrences of each base at each position of the motif and use the inferred frequencies to score candidate sites. Algorithms in this last category include TFSEARCH, FITOM, CONSITE, TESS and MatInspector.

Electro-mobility shift-assays (or gel retardation assays) are a standard way of assessing TF-binding. A fragment of DNA of interest is amplified and labeled with a fluorophore. The fragment is left to incubate in a solution containing abundant TF and non-specific DNA (e.g. randomly cleaved DNA from salmon sperm, of all things) and then a gel is run with the incubated sample and a control (sample that has not been in contact with the TF). If the TF has bound the sample, the complex will migrate more slowly than unbound DNA through the gel, and this retarded band can be used as evidence of binding. The unspecific DNA ensures that the binding is specific to the fragment of interest and that any non-specific DNA-binding proteins left-over in the TF purification will bind there, instead of on the fragment of interest. EMSAs are typically carried out in a bunch of fragments, shown as multiple double (control+experiment) lanes in a wide picture. Certain additional controls are run in at least one of the fragments to ascertain specificity. In the most basic of these, specific competitor (the fragment of interest or a known positive control, unlabelled) is added to the reaction. This should sequester the TF and hence make the retardation band disappear, proving that the binding is indeed specific

Target-specific mutation, as opposed to non-specific mutation. In the context of TF-binding sites, site-directed mutagenesis is typically used to establish/confirm the specific sequence and location of a site, often in tandem with EMSA. Different positions of a putative binding site are mutated to non-consensus (or random) bases and binding to the mutated site is evaluated through EMSA or other means. Often implemented only in conserved motif positions or serially through all positions of a site.

Visual inspection of promoter region to identify putative binding sites based on previous knowledge