This contains one or more sequences that are used by RNAhybrid to hybridize
the miRNA(s) on. RNAhybrid uses all this sequences to find minimal free energy
hybridisations between miRNA(s) and target sequence(s). Sequences should be in
RNA.fasta format but RNAhybrid can also use DNA.fasta files. A single Sequences one
can use can contain up to 50000 basepairs.
contains one or more micro RNA(s) that RNAhybrid uses to hybridize with the
RNA sequences and to find the minimal free energy hybridization. A single micro
RNA sequence can contain up to 2000 basepairs.
RNAhybrid computes the minimal free energy hybridization's the results are
shown by increasing energy. By using the parameters given by bibiserv2 you can
specialize your results the way you want them to be.
helix constraint from |
Forces all structures to have a helix from position a to position b in
respect to the query. The first base has position 1. The parameter "Helix
constrain from" has to be lower or equal to the parameter "Helix constraint to".
You can not use Helix constraint and approximate p-values at the same time.
|
hits per target |
This Parameter defines how many hits are shown by RNAhybrid. The hits are
shown by increasing minimal free energy ( the lower the energy the better the
result)
|
Compact output |
When this parameter is used RNAhybrid gives you only one line of output
instead of the whole output it normally generates.
|
Generate graphics |
Generates a graphical representation of the output in jpg, png and ps format, if
less than 6 hits choosen. If RNAhybrid breaks with an unexpected error, it is often a good idea not to enable
the graphical representation generation.
|
Max internal loop length |
The maximal number of unpaired nucleotides in either side of an internal
loop.
|
energy Threshold |
Shows the hits with all minimal free energy's lower then the threshold (the
lower the result the better). The value has to be lower or equal to
zero.
Notice that the output only shows the results that exceed the energy
threshold or the maximal hits per target.
|
Max bulge loop length |
the maximal number of unpaired nucleotides in a bulge loop.
|
No G:U in seed |
If you click on this you choose weather their are no G:U
bindings allowed in the seed or not. This parameter can only be chosen if you also
use the parameters "Helix constraint from" and "Helix constraint
to". |
helix constraint to |
see helix constraint this is position b you have to use both parameters to
use Helix constraints.
|
approximate p-value |
Used for a quick estimate of extreme value distribution Parameters. You can
choose between nothing, 3utr_fly, 3utr_worm and 3utr_human for better equitation
within these species. You can not use Helix constraint and approximate p-values
at the same time.
|
Usage
Sequences data
The first step of using RNAhybrid is to choose which Sequences you want to use.
There are two windows that you use for your submission. The first window is for the
DNA/RNA sequence(s) and the second for the miRNA(s) you want to hybridize. RNAhybrid
will map the miRNA(s) on the DNA/RNA sequence(s). Its your choice whether you use a
file or you want to copy and paste the input. The server also provides an example,
this example uses a bacterial sequence and a miRNA of the same bacteria. and can be
used to learn how this tool is working. After submitting your sequence data you go
on to step 2.
Parameters
In the second step you choose the parameter set you want to use for your
analyses. RNAhybrid provides you with a lot of useful parameters. The first
parameter is named "hits per target" it is used to calibrate how many hybridisations
you get to see as output. This is competing to the "energy threshold" parameter,
here you choose the maximal MFE (minimum free energy) hybridisations that RNAhybrid
displays, of course if you have only displays the output that has less
hybridisations. For example if you choose a "energy threshold" of -20 and 30 "hits
per target" RNAhybrid will not display 30 hits if their are only 10 hits with less
or equal then the energy threshold.
You can decide if you want an approximate estimate of p-values for the target
sequence therefore RNAhybrid lets you choose between three sequence groups
(3utf_fly, 3utf_human, 3utf_worm). RNAhybrid can calculate better results with this
p-values if you search in sequences of the given groups. Otherwise if you don't want
to use these sequences you have the opportunity to use the "helix constraint"
parameter. This parameter has a "from" and a "to" field here you can say that your
structure should have a helix between those two parameters. This starts counting by
one on your target sequence. When using this parameter you are also allowed to use
the "no G:U in seed" parameter. Using this RNAhybrid tries not to calibrate
structures that have G:U bindings in the seed.
Now its possible to confine special loop structures. The first possibility is
given with the "max internal loop size" parameter. Depending on how you choose the
length of the loop RNAhybrid displays only structures with this as the maximal
internal loop length. For example use 0 as "max internal loop size" and you get
structures with no internal loops at all. Internal loops are loops with both strands
having no binding side, whereas bulge loops are loops where only one strand has no
binding. As said before the other opportunity is using the "max bulge loop size".
Basically this parameter is used in the same way as the "max internal loop size"
parameter only that it delimits the size of bulge loops instead of internal loops.
So if you for example set this parameter to 0 you get structures with only internal
loops.
Last but not least there is the "compact output" parameter
using this parameter the displayed output is only one line of output for each hit. I
would suggest to use this parameter if you only want to know if your results are the
same as RNAhybrid would calibrate. By submitting the combination of parameters you
want to use RNAhybrid starts calibrating the results.
Get the result(s)
The results are shown in a very specific way. First of all you see the Header
which includes the version of RNAhybrid and the Dataset which is shown. Right under
the separation line the first result is shown. First of all the target sequence
length and the miRNA length are pointed out. Furthermore the minimal-free-energy is
shown for the displayed structure, right below this information you can see whether
a p-value is used or which p-value is used. Then the position is marked in which
this structure could be found on the target sequence, below that position mark
RNAhybrid shows how the structure looks like. If you have less or equal to ten
results you get a picture of the structure in addition. This picture can be
downloaded from the results by clicking on the links. RNAhybrid separates all
different Results with a line, so this line is the indicator for going on to the
next Result.