What happens to the miRNA-RISC complex after it cleaves on a mRNA molecule?

I'm more focused on the miRNA than RISC. Does the miRNA degrade after cleaving mRNA?

09 October 2019 9,280 1 View

When the miRNA is bound to a mRNA, is it expressed in a miRNA PCR?

I conducted a miRNA PCR and I was wondering if miRNA that is bound to a mRNA is expressed in the results from my PCR. Thank you!

08 September 2019 7,420 7 View

Can pseudogenes affect miRNA gene expression or do they just competitively bind to miRNA?

The pseuodgene I'm researching has affected the miRNA levels in my cell culture (I found out through qPCR). However, I thought that pseudogenes just bind to the miRNA rather than affect its gene...

07 August 2019 1,959 2 View

What is the recommended concentration range of miRNA cDNA to use in a qPCR?

06 July 2019 4,290 5 View

How do you locate the exact spot of the scratch on a scratch assay?

I was wondering if anyone had any tricks to locate the exact spot of the scratch for a scratch assay. For example, I know some people in my lab have drawn a grid on their plates. Thank you!

06 July 2019 1,406 2 View

What transfection reagent should I use for the T98G cell line?

I am trying to do a stable transfection in the T98G cell line. I want a transaction reagent with a high transfection efficiency. A list of reagents would be preferable. Thank you!

08 September 2018 1,123 0 View

How do I get transfection efficiency with a plasmid with an HA tag?

I want to use a plasmid that is HA tagged. My research is in vitro and I am bit confused on what to do. Thank you!

08 September 2018 6,804 4 View

Does anyone have a recommendation for yeast reporter gene plasmid/protocol?

Hi, could anyone recommend a plasmid and/or protocol for reporter gene assay in S. cerevisiae? I want to assess the effect of growth conditions on a transcription factor, so I want to clone it`s...

01 March 2021 210 1 View

Why might my GFP tag be dissociating from my plasmid after transfection?

I have a set of stably transfected cell lines all transfected with plasmids containing GFP tags on the C terminus. During a western blot using anti-GFP antibody, one of my plasmids has dissociated...

01 March 2021 9,310 4 View

Expression cloning by using cDNA,do I need to modify the cDNA first?

I am going to have a expression cloning of mammalian gene by using shuttle plasmid to transforming the E.coli However I don't know I should only inserting the Coding sequence ,or I can...

28 February 2021 5,440 3 View

Problem with shRNA transfection. Why are only two plates getting contaminated instead of all?

I transfected my LNCaP-WT cells with 3 shRNA plus their NTC two weeks ago and split two puromycin selected cell plates on Friday last week(Feb 26). I checked for GFP in the cells, and they all...

28 February 2021 4,949 3 View

PUC19 as a transfer plasmid for lentiviral delivery?

Hello, Is it possible to use pUC19 as a transfer vector to be packed in using the second generation viral particles packaging system( pMD2.G; psPAX2 plasmids)? As far as I understand it there is...

28 February 2021 4,868 2 View

I want to performe MALDI-TOF MS for bacterial identification, I would appreciate it if anyone can tell me the matrix solution preparation protocol ?

based on what I've studied on multiple papers, for peptide and protein identification Alpha-cyano-4-hydroxycinnamic acid (CHCA) is an appropriate matrix to use, I also know that the CHCA powder...

26 February 2021 6,799 3 View

Did anyone ever try using pLVX-TetOne Plasmid with a copGFP as a selection marker instead of Puromycin?

When using a lentiviral vector for inducible expression of genes using the Tet-On system. In the literature, I saw a widely used of pLVX-TetOne-Puro Plasmid, is there a reason why people prefer to...

25 February 2021 1,011 3 View

Why do the PCR bands from the cDNA templates (extraction from the cell after transfection) show different size from the band of the plasmid template?

After transfection with the plasmid ( linearized ) and subcloning of the cell lines, RNA was extracted from the cell and then reverse transcripted to cDNA. When PCR reactions were run to verify...

25 February 2021 5,712 3 View

How do I transfect a large plasmid into PC12 cells with nerve growth factor (NGF) receptor?

I have a large 16 kb plasmid, which I need to transfect into PC12 cells. Lipofectamin 2000 didn't work and with GFP alone the transfection rate is very low. I also tried the neon invitrogen...

25 February 2021 6,635 5 View

Problem with ligating 2.8 kb insert into 8.5 kb plasmid?

I am trying to clone 2 copies of the same mammalian gene into the pSF-CMV-Ub-Puro Ascl (contains HindIII, KpnI and NheI cut sites) plasmid. This is what the final product is supposed to look like...

24 February 2021 6,310 3 View