Submitting data to the IRESite
The IRESite database is currently accessible only through its web
interface at http://www.iresite.org. The aim of
this database is to help the scientific community with analysis of the
vast amount of experimental data which accumulated from hundreds of
experiments during past two decades. Although the experimental results
are not directly comparable we believe the analysis is possible when
certain parameters are evaluated together. Anyone is welcome to submit
new data and if the data meet certain minimal criteria (like the
sequence is properly labeled, gene names and references are supplied,
etc.) the record is made visible to the public at the main
web-site. To submit a new record you need to login first using
your web browser with session cookies enabled so you do not have to
enter the password after every mouse click. The email address is
necessary because we must be able to contact you in case of questions
related to your submission. Finally, you have your own profile where
you can store your Unfinished data (submissions) and restart
working with them any time later.
To sort the data at the highest level we distinguish two types
of records. Imagine you would like to learn something about the
IRES element of the Hepatitis C virus. In the first round you
want to read some summarized information and not to get lost in
details of the individual hundreds of molecules used in the
experiments to prove that this IRES segment exists at all! So, you
would go for the entry named HCV of natural_transcript type containing
basic overview of the element present in the viral genomic RNA,
accommodated with its sequence, relevant citations and where known also
containing its secondary structure, read the list of known interacting
proteins with their descriptions, interacting regions, etc. If
you get really interested in the HCV IRES you can easily lookup the
HCV records of the plasmid_with_promoter_and_putative_IRES_translationally_characterized type and judge yourself whether
the primary data convince you.
Let us explain our "nomenclature" of these two types of records:
natural_transcript and plasmid_with_promoter_and_putative_IRES_translationally_characterized although it was also
described in the Introduction text reachable through the main menu:
IRESite covers a wide range of experimental data and therefore you have to choose the right record type matching exactly your case.
- If you are about to submit data about IRES containg gene or virus
(you will have to provide sequence of the mRNA or viral genomic +RNA
or whatever was subject to translation by ribosomes) select
natural_transcript entry type. This record type is probably the
simplest-one
- If you are about to submit data about a plasmid vector containing
putative IRES select the plasmid_with_promoter_and_putative_IRES_translationally_characterized record type (your choice
in most cases). Such plasmid_with_promoter_and_putative_IRES_translationally_characterized molecules contain at least a part of
it's sequence artificially inserted or modified (like a pRF plasmid
containing EMCV IRES) and in other words you cannot find such a
sequence in any genome (there is no sequence of pRF plasmid with EMCV
IRES integrated in any genome, right?). That is why the record is
called plasmid_with_promoter_and_putative_IRES_translationally_characterized (such records also include cases when IRESs
was constructed assembled de novo, like IRESs selected from
randomly generated sequence libraries). Be prepared to provide values
describing efficiency of the putative IRES in scale to positive and
negative controls. If you do not have the numbers pick-up the
plasmid_with_promoter_and_putative_IRES_without_translational_characterization (described further below) record type instead.
- If you are about to submit data about a negative control plasmid
used within your experiments (which obviously lacks the IRES segment,
like an empty pRF plasmid) select negative_control_plasmid_with_promoter_and_without_putative_IRES
datatype.
- If you are about to submit data about a promoter-less plasmid
select promoter-less_plasmid_with_putative_IRES datatype.
- Finally, some plasmid sequences cannot be entered into the
database in certain cases as plasmid_with_promoter_and_putative_IRES_translationally_characterized records while they do
contain putative IRES segment their experimental characterization was
so incomplete no useful data showing IRES function are
available. plasmid_with_promoter_and_putative_IRES_translationally_characterized records must have Translation
section filled out. If there are no data showing strength/function of
IRES in experiments involving in vitro or in vivo
translation select plasmid_with_promoter_and_putative_IRES_without_translational_characterization entry instead of
plasmid_with_promoter_and_putative_IRES_translationally_characterized. Beware such records are rather useless and are
included just for the completeness (as acquiring their sequences often
took enormous time and might help the reader of the article answering
some questions). In other words, the datatype plasmid_with_promoter_and_putative_IRES_without_translational_characterization
differs from the type plasmid_with_promoter_and_putative_IRES_translationally_characterized only in the way that the
section Translation is not facultative.
When you choose from the web menu above Records/Submit new
entry you will receive a web form. The web form should guide you
through the submission process. The form might look too large for the
first time visit but not everything must be filled-out. Some fields
are optional and it is the only form you have to fill-out during the
process. Typically you will resubmit the form several times using the
Continue and save a copy ... button to our webserver which will
perform some necessary syntactic controls of the data and check values
of some fields too. Whenever there is an error found in the form the
webserver returns the form back to you with all your data and with
highlighted field containing the problematic data (in red
color). Every form field is surrounded by a basic explanatory text
which we try to keep short but informative. An additional help text is
accessible by pointing the computer mouse cursor over the graphical
symbol containing the question mark icon.
In which order should one submit the data?
The best way is to submit negative_control_plasmid_with_promoter_and_without_putative_IRES
sequences first, then positive_control_plasmid sequences,
then plasmid_with_promoter_and_putative_IRES_translationally_characterized entries and finally natural_transcript record
summarizing the results and utilizing cellular mRNA sequence or viral
genomic sequence. In case still other plasmid data are unsubmitted yet
probably the plasmid_with_promoter_and_putative_IRES_without_translational_characterization record type is the fail-over
choice.
The reason for such order is the fact that when inserting
plasmid_with_promoter_and_putative_IRES_translationally_characterized data you have to correlate IRES activity in
translation relatively to positive/negative controls and for
convenience of others you should refer to IRESite Id: number of their
records which is assigned after you submit new data. The number is in
the leftmost column in the tabular output of your Pending
entries.
How long does it take?
In theory 7 minutes per record.
Is it possible to postpone the data submission if I hit a problem?
Yes. The submission process is time consuming and many
things can happen in the meantime. Therefore, after clicking the
Continue and save a copy ... button the data is not only
analyzed/checked (before being returned back to you with possible
error messages in red) but the contents are also saved into your
account on our server. Therefore, after any Continue and save a
copy ... mouse click, you may ignore any errors returned and
select "Logout" from the web menu. Next time you login you can
continue and fill-up the form you have not finished before. You may
have several unfinished forms in your folder with postponed data! For
the first attempts expect half an hour as you might realize you have
to read the documentation, figure out transcription start sites from your
plasmids, position of ORFs in the transcribed mRNA sequence, position
of IRES within the mRNA, etc. The actual time really depends whether
you have sequence data and annotation readily available or not.
Shall I prepare some data in advance?
We recommend you to collect in advance all the necessary data
in a plain-text file which can be edited by the simplest text
editor you have available (like Notepad on MS Windows). Then, you can
simply cut-and-paste the data into this web form and click the "Submit
to curators" button. The Remarks fields can hold unlimited
amount of additional information. Typically, you may want to include
additional information about the experiment itself, about the IRES
element, about the translational setup, about the proteins interacting
with the mRNA/+RNA molecule or about the secondary structures found in
a certain region. Please note there is a separate Remarks field
for each of these fields, which can keep the data connected only to
the relevant sections. Therefore, you can conveniently split all
your notes and put them in pieces just into the right section! The
same applies to References, which are again available as a part
of every of these distinct sections and thus it is very convenient for
a reader to find in the section called "Protein interactions" only
those references where the actual interactions were described and not
intermixed with the publications describing the plasmid assembly or
mixed with the publications describing the secondary structures found
in whatever region of the molecule.
Is there a list of minimal requirements for a record?
As explained above there are five types of data. All types of
records have certain requirements and most of them do
overlap. However, it is not possible to enumerate here the very long
list of the facultative and optional values. The best overview one can
get when browsing the database and inspecting any record of the
specific datatype in question. Let us emphasize just a few common
attributes:
full-length sequence of the mRNA or the viral genomic +RNA
molecule accompanied by one-line description
positions and description of all open-reading-frames available in
that sequence
exact position of IRES element in that RNA molecule
if a plasmid construct was used to express the mRNA molecule, also
sequence of the plasmid including its characteristics like promoter
used to express mRNA, whether transcription was in
vitro/vivo.
The natural_transcript records are a simple summary of current
knowledge.
The natural_transcript datatype is simpler compared to the
plasmid_with_promoter_and_putative_IRES_translationally_characterized records. For example, natural_transcript datatype
does not have associated any information about the translation of the
concrete IRES element. Of course the IRES is involved in translation
within its host but the translation efficiencies are not known, they
are known only based on experiments and the results are only recorded
as plasmid_with_promoter_and_putative_IRES_translationally_characterized records. Thus, the natural_transcript records are
thought as the summary records of all experimental results describing
the respective IRES element (i.e. summary of the data stored in
plasmid_with_promoter_and_putative_IRES_translationally_characterized records). Obviously, the natural_transcript records
lack the attributes describing actual values measured during the
experiments.
The plasmid_with_promoter_and_putative_IRES_translationally_characterized records are the real experimental
results characterizing new IRESs.
The plasmid_with_promoter_and_putative_IRES_translationally_characterized records on the other hand require you to
enter detailed information about translatability of the mRNA/+RNA
molecule containing the putative IRES. Information include yields of
reporter proteins synthesized during translation or other
quantification of IRES activity in relative scale to controls;
description of experimental controls used in the experiment and
conclusion whether the IRES element is functional or not. To simplify
any future analyses of promoter existence in original constructs
plasmid_with_promoter_and_putative_IRES_translationally_characterized records must be accommodated with full-length
sequence of the plasmid with inserted IRES and reporter
genes.
Which information is optional?
There are many optional fields spread through every record but
there are three completely optional sections:
regions of the mRNA/+RNA complementary to rRNA
proteins physically interacting with any part of the mRNA/+RNA
IRES trans-acting factors (ITAFs) which somehow modulate IRES
activity and might not interact directly with the mRNA/+RNA
regions of the mRNA/+RNA being experimentally characterized in
term of its secondary structure
Why should I submit my own data?
Extracting all the data from original publications looks
straightforward but involves a lot of manual and creative
work. Deducing the primary sequence of the plasmid containing inserted
IRES is just the most difficult part and in several cases we were
blocked by mistakes in published materials and had to resolve the
issues with original authors anyway. Therefore we hope scientists will
submit their own primary data freely and we will be happy to assist
anyone who asks for assistance.
Thank you very much for your interest!
Your IRESite team
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