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kaveh's Content

There have been 6 items by kaveh (Search limited from 30-September 19)


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#165922 Can shRNA be picked up by sequencing?

Posted by kaveh on 10 March 2014 - 02:28 PM in Molecular Biology

Imagine you are doing a shRNA experiment. You knock down the gene of interest by shRNA and you study the transcriptome. If part of a shRNA is complementary to the target mRNA, once the next-gen sequencing library is being made, doesn't shRNA become converted to cDNA? Wouldn't this be sequenced and read as the target RNA in sequencing? 

 

Thanks. 

 

KD




#161935 genomic DNA extraction on silica-based membrane

Posted by kaveh on 31 October 2013 - 08:01 AM in Molecular Biology

I see more and more kits that use silica columns for extraction of genomic DNA from tissues, cultured cells and blood. This makes the colony screening faster and easier for mutation analysis studies or other assays. I know that one can buy the silica columns in bulk. I was wondering if anyone has recipes for required buffers (lysis buffer, binding buffer, wash buffer, elution buffer). 

 

Thank you. 




#161546 expressing flag tag by itself

Posted by kaveh on 22 October 2013 - 02:08 PM in Molecular Biology

I was wondering if it's possible to express flag tag all by itself. I was thinking about putting the flag tag (with a kozak sequence in the beginning and a transcription terminator in the end) downstream of a promoter and upstream of a poly-A signal. Would this lead to expression of a short peptide that could be recognized by antibody?

 

thanks.




#160423 How to determine the location of integrated DNA in stable cell lines

Posted by kaveh on 19 September 2013 - 02:39 PM in Molecular Biology

When you make stably transfected cell lines, I assume that the plasmid DNA (or at least part of it) gets randomly integrated into the genome of the cells. How one can determine the exact location of the plasmid DNA in the host genome?

 

Thank you. 




#157114 Transfecting three plasmids into hES cells

Posted by kaveh on 28 June 2013 - 03:52 PM in Cell Biology

Hi,

I would like to transiently transfect 3 different plasmid DNAs into human embryonic stem cells by lipofectamine. If cytotoxicity is not a huge issue, what amount of each plasmid would you use in a well of 6-well plate.

any suggestion is appreciated.

k



#156046 Labeling and tracking RNA

Posted by kaveh on 04 June 2013 - 01:54 PM in Molecular Biology

Hi,

I am trying to find a way to fluorescent-label RNA molecules and track them in the cell by microscopy. My idea is to synthesize the RNA (in-vitro transcription) and transfect them into the cell. I know that you can incorporate fluorescent labeled dNTPs into the RNA (like the way one would synthesize RNA-FISH probes), but is the RNA structure/function would be unaltered? Is there any method for adding a fluorescent label to the end of the RNA, without messing with the main portion of it?

Thank you for your time

K




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