5. Tick the Consensus Sequence option. We will enter the motif
sequence in Fasta format, over two lines—copy the following
to do so, including the return character after the “CACGTG”
motif:
> GBOX
CACGTG
6. Toggle “Only show significantly enriched motifs (slow).” You
can also specify the significance parameters. In this example
analysis, we will use the default parameters, which includes a
Z-score cutoff of greater than 3, a functional depth cutoff of
0.35 and that this motif must be found in at least half of the
genes in the gene set.
7. Perhaps you are also interested in searching additional motifs.
To search for known motifs, return to the input page and keep
all other settings the same but choose “Only Arabidopsis
PLACE elements” in the step 3 “Data set” dropdown menu,
which will use one of two parts of a previously published motif
database,
PLACE
[37],
which
contains
around
100 cis-elements from plants, manually curated from published, small-scale studies (the G-box motif is encompassed in
this set).
8. Click on “Begin Search” and Cistome will display a diagram
with the overrepresented regulatory elements mapped on the
promoters of the genes included in the analysis (this analysis
may take 2–3 min; be patient) (see Fig. 10). Cistome determines overrepresentation by comparing the frequency of
occurrence of each motif against the frequency of occurrence
of the same motif in randomly selected sets of promoters from
the background set. We set that the G-box is indeed overrepresented in our set of promoters, suggesting our hypothesis
regarding PIF-family transcription factor regulation is correct.
9. Some other useful aspects of the Cistome tool: click on “Cluster View” at the tab along the top of the Cistome output.
Cistome will displays a dendrogram of the overrepresented
motifs based on the similarity of the PSSMs generated from
the mapping procedure.
10. Click on “Seq Logo View” to get the frequency of the distinct
nucleotides that are found in the overrepresented binding sites.
Once you have a given sequence motif you can identify other
genes in the genome that may contain this element. You can
then query coexpression databases to see if these genes are
coexpressed with your gene of interest or, in this example, if
they are coexpressed with ABI3 under any other conditions.
This would suggest common regulatory control of a suite of
functionally related genes.
48
G. Alex Mason et al.
sequence in Fasta format, over two lines—copy the following
to do so, including the return character after the “CACGTG”
motif:
> GBOX
CACGTG
6. Toggle “Only show significantly enriched motifs (slow).” You
can also specify the significance parameters. In this example
analysis, we will use the default parameters, which includes a
Z-score cutoff of greater than 3, a functional depth cutoff of
0.35 and that this motif must be found in at least half of the
genes in the gene set.
7. Perhaps you are also interested in searching additional motifs.
To search for known motifs, return to the input page and keep
all other settings the same but choose “Only Arabidopsis
PLACE elements” in the step 3 “Data set” dropdown menu,
which will use one of two parts of a previously published motif
database,
PLACE
[37],
which
contains
around
100 cis-elements from plants, manually curated from published, small-scale studies (the G-box motif is encompassed in
this set).
8. Click on “Begin Search” and Cistome will display a diagram
with the overrepresented regulatory elements mapped on the
promoters of the genes included in the analysis (this analysis
may take 2–3 min; be patient) (see Fig. 10). Cistome determines overrepresentation by comparing the frequency of
occurrence of each motif against the frequency of occurrence
of the same motif in randomly selected sets of promoters from
the background set. We set that the G-box is indeed overrepresented in our set of promoters, suggesting our hypothesis
regarding PIF-family transcription factor regulation is correct.
9. Some other useful aspects of the Cistome tool: click on “Cluster View” at the tab along the top of the Cistome output.
Cistome will displays a dendrogram of the overrepresented
motifs based on the similarity of the PSSMs generated from
the mapping procedure.
10. Click on “Seq Logo View” to get the frequency of the distinct
nucleotides that are found in the overrepresented binding sites.
Once you have a given sequence motif you can identify other
genes in the genome that may contain this element. You can
then query coexpression databases to see if these genes are
coexpressed with your gene of interest or, in this example, if
they are coexpressed with ABI3 under any other conditions.
This would suggest common regulatory control of a suite of
functionally related genes.
48
G. Alex Mason et al.