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How do i validate a cancer gene?


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#1 Tszki

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Posted 08 January 2015 - 12:01 AM

I am now doing exome sequencing to identify candidate genes which may responsible  for the parthenogenesis of lung cancer. After I have some candidates, I need to do some downstream functional validation for the genes. I would like to ask whether there is any standard experiments or techniques i can use to do the initial functional screening of cancer genes?



#2 bob1

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Posted 08 January 2015 - 01:45 PM

You could test to see if knockdown/knockout affects growth rates in the cells.



#3 CPRES

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Posted 13 January 2015 - 02:25 PM

Of course, sky is the limit with characterizing any new gene. Even from cancer point of view you can do hundreds of things, here is a quick and brief list:

 

1. Try to identify from literature/bioinformatics what this gene might do based on its class, structure etc. This is important because sets of experiments you would do are different for each class: transcription factor, signal transduction, transport, growth factor, structural protein etc.

 

2. As above, do stable/transient knockdown/knockout and overexpression of the gene in right cells.

 

3. Characterize the modified cells: proliferation, apoptosis, shape change, migration change, changes in global/select gene expression (with focus on known oncogene, tumor suppressors and EMT) genes etc.

 

4. Inject the modified cells in mouse (xenograft assay) to document all of the above in in-vivo condition. Can do 3D culture too.

 

5. If transcription factor, do luciferase assays once targets are know. Do ChIP-Chip to identify global targets (again look for known cancer-related genes).

 

6. Make a transgenic or knockout mouse.

 

7. Look at human tumor gene expression registries, and identify which human cancers show consistent change in expression of your gene. Focus on that tumor. Get human tumor tissue and validate the data analysis findings.

 

8. Good luck, you will need it.


So. Now that you have your first ever question on bioforum answered (or not), mail yourself your username and password so you don't forget them, and then come back soon to update us on how it all worked out. That's how you build Karma in science.


#4 Tabaluga

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Posted 15 January 2015 - 12:39 PM

Excellent post, Cellcounter.


Il dort. Quoique le sort fût pour lui bien étrange,
Il vivait. Il mourut quand il n'eut plus son ange;
La chose simplement d'elle-même arriva,
Comme la nuit se fait lorsque le jour s'en va.

 





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