Jump to content

  • Log in with Facebook Log in with Twitter Log in with Windows Live Log In with Google      Sign In   
  • Create Account

Submit your paper to J Biol Methods today!
Photo

workflow - start from microarray


  • Please log in to reply
4 replies to this topic

#1 jiro_killua

jiro_killua

    Enthusiast

  • Active Members
  • PipPipPipPipPip
  • 55 posts
0
Neutral

Posted 07 February 2009 - 01:58 PM

I think many ppl who are new to this field will have similar situation as me

We did microRNA microarray and got some data

But what's next?


I know if you want to prove the relationship between a microRNA and a protein, you can do transfection, then Western, then probably 3'UTR mutation and luciferase assay

But what's the best and most efficient way from microarray data to narrow down one microRNA-protein pair?

Like....what assay should be done to verify the change in microRNA? Northern or realtime? what algorithm to predict target is the best? etc....


Thanks

#2 miRNA man

miRNA man

    Enthusiast

  • Active Members
  • PipPipPipPipPip
  • 73 posts
1
Neutral

Posted 09 February 2009 - 06:40 AM

I did qRT-PCR (TaqMan) to confirm some of the interesting miRNAs on our array and it worked out pretty well and was concordant with the array results. After that, I guess it's up to you and your ideas and what your hypothesis is. My impression is that finding an up/down regulated miRNA and then validating one mRNA target isn't enough to get into a nice journal anymore. I think you need to do some functional studies and see how the miRNA fits into a pathway. But luciferase assay is the way to go to validate targets.

#3 Fizban

Fizban

    Enthusiast

  • Active Members
  • PipPipPipPipPip
  • 63 posts
0
Neutral

Posted 10 February 2009 - 05:34 AM

I think many ppl who are new to this field will have similar situation as me

We did microRNA microarray and got some data

But what's next?


I know if you want to prove the relationship between a microRNA and a protein, you can do transfection, then Western, then probably 3'UTR mutation and luciferase assay

But what's the best and most efficient way from microarray data to narrow down one microRNA-protein pair?

Like....what assay should be done to verify the change in microRNA? Northern or realtime? what algorithm to predict target is the best? etc....


Thanks


I agree with miRNA man.
qPCR should be your choice instead of northern, it's much more sensitive and quick.
aside from the "standard" assays such as luciferase and so on you should try to understand or to think what is the real function of your miRNA. having a confirmed target could help, but up to now it is still the hard part of the job!
good luck
Fizban

#4 Jon Moulton

Jon Moulton

    Veteran

  • Active Members
  • PipPipPipPipPipPipPipPipPipPip
  • 137 posts
3
Neutral

Posted 10 February 2009 - 10:14 AM

You can confirm a target MRE with a Morpholino oligo.

Choi WY, Giraldez AJ, Schier AF. Target Protectors Reveal Dampening and Balancing of Nodal Agonist and Antagonist by miR-430. Science. 2007 Oct 12;318(5848):271-4. Epub 2007 Aug 30.
Jon D. Moulton
Gene Tools, LLC
www.gene-tools.com

#5 Fizban

Fizban

    Enthusiast

  • Active Members
  • PipPipPipPipPip
  • 63 posts
0
Neutral

Posted 26 February 2009 - 12:43 AM

You can confirm a target MRE with a Morpholino oligo.

Choi WY, Giraldez AJ, Schier AF. Target Protectors Reveal Dampening and Balancing of Nodal Agonist and Antagonist by miR-430. Science. 2007 Oct 12;318(5848):271-4. Epub 2007 Aug 30.


not an easy task when your predicted targets are 200/300! B)
prediction is SO hard!!
fizban




Home - About - Terms of Service - Privacy - Contact Us

©1999-2013 Protocol Online, All rights reserved.