Silenced gene does not affect downstream target genes?

Hi, I silenced a gene (LSD1, KDM1A) using 2 different shRNA in 3 different cell lines. When I check its mRNA level, it is more than 70-80% silenced. However, if i check its downstream target (at...

03 April 2018 5,872 3 View

Calcium Phosphate Transfection Problem?

Hi, I am trying to transfect 293T cells using 2M CaCl2 and 2X HBS (50 mM HEPES, pH 7.07, 10 mM KCl, 12 mM dextrose, 280 mM NaCl, 1.5 mM Na2PO4) However, some plates, still have particles (probably...

02 March 2018 4,689 6 View

SiRNA Transfection to COLO205, SW620, HT29?

Hi, Is there anyone who has done siRNA Transfection in COLO205, SW620, HT29 especially using either RNAiMAX or Lipofectamine3000? Besides do you have any experience to transfect/transduce plasmid...

11 December 2017 1,759 4 View

Cannot show reduced lncRNA level using siRNAs?

Hi all, I am trying to perform a loss-of-function assay. I silenced some lncRNAs which I found them from a high-throughput screening considering changes in their downstream molecule. In order to...

09 October 2017 3,855 0 View

Alternative to Southern Blot?

08 September 2016 4,045 3 View

Having problem with mutant IDH (Isocitrate dehydrogenase) enzyme overexpressed cells?

Hi, I am having trouble with cells overexpressed mutant IDH enzyme. Problem is that it does not work. In other words, I chose some genes as a positive control from other papers and evaluated their...

02 March 2016 8,254 3 View

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08 September 2015 4,775 2 View

How can I protect RWPE-1 cells from any contamination?

06 July 2014 8,632 11 View

Is there any software or website for matching sequences and diseases?

03 April 2014 8,918 4 View

Can we classify HV and LV lines based on line length?

I have dataset which shows the length of power lines. I need to classify the lines based on the line length. Is there a rule to classify the High voltage (HV) and low voltage (LV) lines based on...

03 March 2021 4,116 4 View

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02 March 2021 3,060 3 View

Issues with Permeabilization during Flow Cytometry?

Hello Everyone. Currently I am working to characterize macrophages in the myocardium after ischemia-reperfusion injury in rats. Due to the low total cell number isolated from rat hearts I can...

01 March 2021 3,867 3 View

Which cell line should we use to perform biological dosimetry experiments?

We are preparing some experiments based on irradiating cells under different conditions in order to evaluate the effects in terms of DNA damage, genetic expression, etc. As our project is...

01 March 2021 3,355 3 View

Is it possible for two human neuronal cell lines to have different gene sequence for the same protein?

I am looking at the ATP1A2 (Sodium/Potassium ATPase alpha subunit 2) in two human neuronal cell lines. Expression levels of this protein seems to be almost equal when detected by one antibody....

01 March 2021 3,607 3 View

Is CD44 degraded in lysosome together with hyaluronan after CD44 mediated endocytosis of hyaluronic acid?

Also when RHAMM binds hyaluronic acid, they get internalized, will RHAMM also be degraded? Or both CD44 and RHAMM will be transferred back to the cell membrane? Asking for breast cancer cell line...

01 March 2021 8,169 2 View

Do you need the CMV enhancer before the CMV promotor for a proper function of CMV on the gene that follows?

I'm a student and I have to produce a cell line with a knockin for the NRF2 gene with GFP. I have to put a promotor in front of the GFP because the gene will be too far away from the promotor of...

28 February 2021 7,127 2 View

What could be some reasons why unstimulated monocytes are getting activated?

Im doing PBMC isolation -> CD14+ enrichment using magnetic beads -> stimulation setup. My negative control is just cells in cRPMI but they seem to get activated over and over again.

28 February 2021 7,883 3 View

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28 February 2021 9,936 2 View

Has anyone had luck using cell lines other than HEK 293 (such as COS1) to package lentivirus?

I am looking to package lentivirus in endothelial cells but I am opposed to using 293 cells due to personal ethics regarding the source. I have done some reading and saw that COS1, COS7, & CHO...

25 February 2021 3,030 1 View