ANNOTATION OF EXPRESSION : ANNOTATION OF EXPRESSION Mani Grover
Institute Of Bioinformatics,
International Technology Park, Bangalore Date: 16-09-09
What is Expression? : What is Expression? Expression is defined as synthesis of functional protein from genetic material DNA/RNA.
Pipeline For Annotation of Expression : Pipeline For Annotation of Expression 1. Expressions are considered from NORMAL HUMAN TISSUES (HPRD) and
NORMAL MOUSE TISSUE(MPRD).
2. Expression from DISEASED TISSUES or CELL LINES is not considered
3. Standardizations for the tissue terms, hierarchy of tissues in which expression is
considered has been listed in the Master List for expression.
4. ALWAYS CITE THE ORIGINAL PAPER FOR EACH EXPRESSION. Not any back refrence Continue.........
Slide 4 : 5 . Expression refers to 2 things:
1.The tissue which has been shown to produce the protein.
2.The part of the body from which the protein has been isolated.
For instance, if the protein is found in the blood or milk, then we can add these to the expression, though technically they are not synthesized there .
6. We should not annotate expression of protein in transgenic mouse tissue and only normal wild type mouse tissue (MPRD),Normal human tissue(HPRD) should be considered. Pipeline continue...... Continue...............
Slide 5 : 7. If we find papers talking about 'Ubiquitous' expression, and other papers talk about tissues, we can put both.
8. If a paper has performed expression analysis (mRNA expression or immuno-detection of protein), those articles should be preferred over cDNA library sources for the same tissue. cDNA library can be cited if there is no experiment done for the expression analysis for that particular tissue.
For e.g. we have a paper talking about the protein source from a human placental cDNA library we have another paper detected expression in brain, heart, skeletal muscle and Kidney we can cite placenta expression from former paper, along with brain, heart, skeletal muscle and Kidney from the latter.
In other words, if latter also mentions Placenta expression, we will not cite the former paper talking about cDNA source. Only expression analysis paper will be cited for placenta expression along with other expression there in. Continue........
Slide 6 : 9. If we have a paper talking about expression in "Peripheral blood leukocytes" and then the same paper goes on to specify the PBL worked on - like monocytes, lymphocytes etc, then put expression in these specific cells and do not include Peripheral Blood leukocytes again.
But, if we have a paper talking about expression in Peripheral blood leukocytes, and another paper talking about expression in monocytes and lymphocytes, we will include all three - Peripheral blood leukocytes, monocytes and lymphocytes.
10. We put as expressions what the protein gets secreted into (E.g. Tears, Sweat etc). While putting these, it will be better to put the tissues in which they are actually expressed also. (E.g., Lacrimal glands, Sebaceous glands etc). So we will have both Tears and Lacrimal glands under expressions for that protein.
If many papers give the expression of the protein being annotated as ubiquitous, and we get a recent paper that has done an extensive analysis of expression, then put the expression in tissues specified and include ubiquitous expression also.
Slide 7 : MASTER LIST Developmental stages and fetus :
+Zygote
++Blastomere
+++Morula
++++Blastocyst
+++++Trophoblast
++++++Syncytiotrophoblast
++++++Cytotrophoblast If expression in parts having same number of plus sign is proved we should include both but write it in two different rows
Slide 8 : Methods For Detection Of Expression In Situ Hybridization
RT-PCR
Western blotting
Immunofluorescence
Immunoblotting
Peptide pull down followed by MS
Immunoelectron microscopy
Fluorescence immunostaining
Immunofluoresence microscopy (intra vital microscopy) Continue....
Slide 9 : Northern blotting
Immunohistochemistry
Mass Spectrometry
RNase protection assay
GFP fluorescence, Microscopy
Two-dimensional nonreducing/reducing diagonal gel.
MALDI-TOF-MS followed by Western blotting. Methods continue....
Example of Expression : Example of Expression ++++Cerebellum ++Spinal cord ++++++Cerebral cortex +++++++Striatum Brain stem is not mentioned in master list
Medulla ++++
Pons +++++
Mid brain +++
: 1. Annotation of Expression of m-RNA
2. Annotation of Expression of protein in transgenic mouse tissue.
Problems ................................ 1. Annotation of author statement “a protein is ubiquitously expressed in Brain”.
2. For house keeping gene, we can put the expression as ubiquitous.(pipeline statement) Doubts Faced
Challenge we are facing : Challenge we are facing Whether we should annotate expression in cell lines or not especially in primary cell lines................................................. “Innovation is our journey and not our destination” Thank you!