For Hela cell,which microRNA is highly expressed?

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mei wang
mei wang's picture
For Hela cell,which microRNA is highly expressed?

I need to set up the microRNA analytical assay.But the most important things are to isolating the microRNA from some cells and validate the microRNA by electrophresis or QRT-PCR!
However,for many tumor cells the microRNA is downregulated,such as let-7、mir-15 and so on! So it is not benefitful for me to use these classic microRNA to set up methods!
I need help! what I need to do !
In my lab,nobody was engaged in this field!
I need directions!

ryan_m
ryan_m's picture
Why not try a microRNA array

Why not try a microRNA array to determine which miRNAs are present and highly expressed?

Ryan

saswati1
saswati1's picture
Read the following paper

Read the following paper
* Tables

Abstract

Eukaryotic cells express small noncoding RNAs to silence target genes or regulate chromatin domains. MicroRNAs (miRNA) are endogenous small RNAs that are thought to each regulate multiple mRNA targets. To gain a deeper understanding of processes regulated by small noncoding RNAs, techniques are required to identify and detect them. This unit describes standard laboratory methods for and comments about high-throughput technologies for the identification and detection of small RNAs. This unit also outlines a cell-based reporter gene assay for assessing the regulatory potential of a microRNA on a candidate mRNA target. Curr. Protoc. Mol. Biol. 81:26.7.1-26.7.15. 2008 by John Wiley & Sons, Inc.

mei wang
mei wang's picture
ryan_m wrote:Why not try a

ryan_m wrote:

Why not try a microRNA array to determine which miRNAs are present and highly expressed?

Ryan

Thank you for reading my topic and giving me an advice! However, My boss wouldn't spend much money in setting up a method! He asked me to select a cell line and a target microRNA which has been verified! Finally,I am determined to select A549(a kind of lung cancer cell lines) and hsa-miR-21 (highly expressed) !Then isolating microRNA and performing real-time PCR to check it!
I don't know whether such design is OK!
So could you give me some better advices?
Another question :if I want to use SYBER GREEN to perform QPCR,which kit is suitable to select? The SYBER GREEN I type or II type?

mei wang
mei wang's picture
saswati1 wrote:Read the

saswati1 wrote:

Read the following paper
* Tables

Abstract

Eukaryotic cells express small noncoding RNAs to silence target genes or regulate chromatin domains. MicroRNAs (miRNA) are endogenous small RNAs that are thought to each regulate multiple mRNA targets. To gain a deeper understanding of processes regulated by small noncoding RNAs, techniques are required to identify and detect them. This unit describes standard laboratory methods for and comments about high-throughput technologies for the identification and detection of small RNAs. This unit also outlines a cell-based reporter gene assay for assessing the regulatory potential of a microRNA on a candidate mRNA target. Curr. Protoc. Mol. Biol. 81:26.7.1-26.7.15. 2008 by John Wiley & Sons, Inc.

Thank you very much for reading my topic!But this paper is not available! Would you please send this article to my email address? My email address is eval(unescape('%64%6f%63%75%6d%65%6e%74%2e%77%72%69%74%65%28%27%3c%61%20%68%72%65%66%3d%22%6d%61%69%6c%74%6f%3a%77%61%6e%67%6d%65%69%38%34%31%37%40%31%36%33%2e%63%6f%6d%22%3e%77%61%6e%67%6d%65%69%38%34%31%37%40%31%36%33%2e%63%6f%6d%3c%2f%61%3e%27%29%3b'))!