Download Powerpoint

The BioRetis Database
A tool for analysis and sharing of array data
Objective
• analysis of Affymetrix GeneChip data
• automation of group comparisons
– standards of analysis developed in the NGFN
• retrieval of candidate genes
– from single group comparisons
– from multiple group comparisons
– by predefined function
• download of results for further analysis
• integration of public data
• restriction of access with options for sharing
Affymetrix GeneChip data
•
hybridization with one sample and a single
color
•
validation of detection by using
–
–
•
statistical analysis of hybridisation based on
the different oligonucleotides per probeset
to provide
–
–
•
sets of several oligonucleotides per gene
mismatch oligonucleotides
a measure for significance of detection
a signal intensity relative to whole chip
intensity
statistical analysis of pairwise chip
comparison based on the different
oligonucleotides per probeset to provide
–
–
a measure for the significance of difference
a value for the magnitude of difference
GeneChip data needed in BioRetis
data for each
array
data for each
pairwise comparisons
Signal
Signal log ratio
Detection call
Change call
Pairwise comparisons needed for each
group comparison
Example:
Experiment group E: n=3 chips
compared to
Baseline group B: n=3 chips
number of pairwise
comparisons:
this results
for 2x3=6 arrays in
for 2x10=20 arrays in
for 2x20=40 arrays in
Chip E1
Chip B1
Chip E2
Chip B2
Chip E3
Chip B3
E vs E
3
45
190
E vs B
9
100
400
B vs B
3
45
190
total
15
comparisons
190 comparisons
780 comparisons
Description of Array Experiments and
Comparisons
Names for each array reflect information to the following categories:
donor group
donor number
tissue type
stimulus
kinetics
array type
ND
1
WBC
LPS
4h
133A
normal donor
first sample
white blood cells
lipopolysacharide
4 hours
HG-U133 Plus 2.0
Example for a group:
ND_WBC_LPS_4h_133A
(Arrays from GEO dataset GSE3284:
healthy individuals were challenged with LPS and
white blood cells were separated and analyzed
after 4 hours of challenge
on a HG-U133A GeneChip)
Example for a group comparison
SPL_28:ND_WBC_LPS_4h_vs_ND_WBC_LPS_0h_133A
(4h challenge is compared to WBC before challenge)
Gene Annotations
•
•
•
•
•
•
Affymetrix ID
GeneSymbol
Gene Ontology Biological Process
Gene Ontology Cellular Component
Gene Ontology Molecular Function
link to Entrez Gene
Access and Rights
• You may access
– public data
– own data
– data owned by others and shared with you
• You may share
– only based on rights for accessing group
comparisons
– not based on rights for accessing chips
Access and Rights
You
Colleague
 Group A
Group C
Group B
Group D
If you allow access to a single or a group of your chips
() to a colleague, the colleague would be able to
perform comparisons with any of his own chips.
Access and Rights
You
Colleague
You
Colleague

 Group A
Group C
Group A
Group C
Group B
Group D
Group B
Group D
To limit access to only one specific group comparison,
for example group A vs group C, rights have to be
limited to a single group comparison.
Basic structure of the database
rights
array
rights
pairwise comparison
rights
group comparison
description
sample
array
description
group comparison
values
signal
values
pairwise comparison
values
group comparison
values
genes
annotation
probe set ID
probe set ID
array type
rights
samples
& comparisons
Working with BioRetis: Access
URL: http://www.acs-nett.de/bioretis
click on the appropriate card to
•login
•register for a personal account
•access public data anonymously
•get an introduction
•see impressum
when you register,
you define your username, your e-mail, and
you will receive a password to this e-mail address
Change your profile
to change to a
new password,
goto
„account“ „settings“ …
… and enter old
and new password
you may also change the
general settings of access
to public data and
to predefined gene list for retrieval
Starting to work
Select from
the drop down menu
or click directly on
the symbol to perform
a new group comparison …
… or to perform
queries on single …
… or multiple group comparisons
Perform a new group comparison:
select the Experiment group
To perform a
new group comparison
you select the
experiment group E from
the list of all groups that
are owned by you or
that are shared with you
by a colleague
You may narrow in to the
group of interest by specifying
group parameters
Select Experiment chips
Select individual chips for
the experiment group E
Select Baseline group
Select the baseline group B
Select Baseline chips
Select individual chips for
the baseline group B …
enter a description (optional) …
and click „Start“
Group comparison started
The group comparison you
have initiated is shown on the
subsequent page …
…and you may
check the queue for newly
initiated group comparisons still
waiting for calculation
See queue of initiated group
comparisons
The queue of
group comparisons
still waiting for calculation.
The process of calculating
a single group comparison
takes 20 sec to 5 minutes,
depending on the number
and type of arrays.
Querying a single group comparison
Select from
the drop down menu
or click directly on
the symbol to query
a single group comparison
Selecting the group comparison of interest
If
necessary,
Specify
parameters
specify parameters
to reduce the number
to reduce the number
of
groupcomparisons
comparisons
of group
displayed
displayed
Select the group
comparison of interest …
… and click the
Next button
Enter query parameters (1)
You may click on the parameterset area …
… to select default or your own
saved parameters …
… and fill the query form
Enter query parameters (2)
You may click on the columns area …
… to change the number of columns in
the query form…
… and reload the query form
to enter more query parameters
Enter query parameters (3)
You may click on the probesets area …
… to select a list of probesets …
… and Start the query in the
search parameter area
Enter query parameters (4)
You may enter
Affymetrix Probeset ID or
GeneSymbol …
Gene Ontology parameters …
thresholds for
statistical parameters of the
Experiment group (EG) …
or the Baseline group (BG) …
results between
columns are
combined
by „OR“ or
„AND NOT“
Signal Log Ratio (SLR) or
Fold Change (FC) …
the percentage of change call
increased or decreased in all of
the pairwise comparisons
between EG and BG
p-value thresholds for the t-tests
comparing log-transformed signals
or SLREGvsBG compared
to SLREGvsEG & SLRBGvsBG,
SLREGvsBG compared to SLREGvsEG
or SLREGvsBG compared to SLRBGvsBG
percentage of no change
all parameters
in one column
are combined
by „AND“
Enter query parameters (5)
You may click on the
save parametersets area …
… enter a title and description …
… and search and save the query
Display of results
symbols to click for switching to cards that
display the arrays that were compared or
to download results and signals
Affymetrix Probeset IDs,
symbol to link to the gene in Entrez Gene,
the GeneSymbol and
all statistical parameters of the
group comparison
Display details and download results
click on the symbol
to download results, …
… to download
parameters & results …
… activate
to
or to activate
extraction
extraction
of signals
of
and to download
signals
and to download
parameters & results & signals
in one single xls-file
Excel file for single group comparison
query
parameters
results
signals
Querying multiple group comparison
Select from the drop down menu
or click directly on the symbol to
query multiple group comparisons
Selecting multiple group comparisons
specify
(optional)…
specifyprameters
prameters,
select multiple group
… select multiple group
comparison
of interest
comparison of interest…
and click the
Next
…
andbutton
click the Next button
Enter query parameters
modify your selection of group
comparisons if necessary, …
… define Boolean operation between
group comparisons, …
… enter selection criteria and …
… click next
Display of results
symbols to click for switching to cards that
display the arrays that were compared or
to download results and signals
Group comparison,
Affymetrix Probeset IDs,
a symbol linking to the gene in Entrez Gene,
the GeneSymbol and
all statistical parameters of the
group comparison
Excel file for multiple group comparison
query
parameters
results
signals
first
comparison
signals
second
comparison …
Administration of rights to share
click on rights and select user or analysis
Select user to share with
select user
Give rights to access a comparison
deny access (No Rights)
or give access to query
(May Read)
or give access to query and
to perform subanalyses (May Analyse)
or give access to query,
to perform subanalyses and
to give rights for access by
other users (Co-Owner)
and confirm your selection
Select comparison to share
specify to reduce listed
number of comparisons, …
… select a comparison …
and click on Next
Administration of rights to share
deny access (No Rights) …
… or give access to query (May Read) …
… or give access to query and
to perform subanalyses (May Analyse) …
… and confirm your selection
… or give access to query,
to perform subanalyses and
to give rights for access by
other users (Co-Owner) …
Upload of gene lists for querying
enter the title for the gene list …
… and the description, …
… select the txt-file on your
computer (consisting of a single
column of Affymetrix Probeset-IDs) …
and start the upload
Administration of gene lists
goto settings …
click on Probeset Lists …
Display, update or delete gene lists
enter the title …
to display
to display
thethe
gene
gene
list list
…
you may delete …
… or update the list
Show Experiments to prospective collaborators
Goto cooperation …
… and show your experiments
Select the experiments you want to show
Activate the experiments that
you want to show to the public …
… and confirm
This will only show your experiment to the public
(see next slides) but NOT your expression data!
Find Experiments of other users
Goto cooperation …
… and find experiments performed
by others
Find an experiment and contact the owner
Specify the parameters …
…to find and select experiments
of interest
Click Next …
… to see the owner …
(all your own experiments will be listed here
that you have activated in „Show experiments“)
… and to get in contact