What it is the best method to transfect mirna mimics and UTR plasmids to make a luciferase assay?

I would like to know a goog transfection reagents to use with plasmids and mirna mimics together, looking the different lipofectamines I wonder if it is necessary to use for example the lipo 3000...

04 May 2017 7,067 2 View

What is the best and easy software to analyze promoters?

I used genomatyx to analyze transcription factors in promoters sequences, but now I have many problems to login, so does anyone knows another facility to do this?

09 October 2016 5,956 5 View

Too much beta lactamase in protein expression BL21 cultures?

06 July 2016 9,412 5 View

Why is recombinant mutant protein expressed less than wild-type?

09 October 2015 1,153 7 View

Bad resolved band in EMSA?

Sometimes it is not possible to see single bands in EMSA, and complexes remain as a big spot not resolved. Could anyone explain to me how to find a solution?

09 October 2013 3,564 5 View

Bands in EMSA gel remain as a big spot, not well resolved, is it impossible to see single bands?

Could anyone bring me a solution?

09 October 2013 1,197 0 View

Protein is not visible on western blot of transfected cells by confocal microscopy

I made cDNA constructions to produce proteins with GFP or dsRED (clontech), and one construction with dsRED after transfection in HeLa cells. I obtained RNA by qPCR, red cells by confocal...

31 December 2012 866 14 View

How can I increase release of my GST-fused protein after digestion with Prescission?

I bind my fused GST-protein to glutathion beads and I cut with Prescission at pH 7.5 to eluate overnight in batch method, sometimes my protein (always the same clone) remains attached to beads...

01 January 1970 6,346 5 View

Enhanced Yellow Fluorescent Protein (Aequorea victoria) DNA sequence?

I am on the lookout for the Enhanced Yellow Fluorescent Protein (Aequorea victoria) DNA sequence. Does anyone know where I can find it? Thank you in advance

03 March 2021 3,568 1 View

Can linear DNA be used as template for site-directed mutagenesis?

Is it possible to induce site-directed substitution mutation by quick-change method on linear dsDNA? or it has to be cloned in some vector? If yes, should it be treated with the Dpn1 enzyme...

03 March 2021 401 4 View

How do I increase the yield of my PCR product?

Hello, We would like to increase the yield of our PCR product. We are running a series of PCR reactions that is targeting ~1.1kb sequence. We begin each reaction with ~400pg of template DNA...

02 March 2021 4,029 3 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

Does anyone know HRM channel in a 2 plex QIAGEN Rotor- Gene Q qPCR can be used as a separate channel?

I am going to have 3 different probes in my qPCR work that I am going to do. But I realized that the machine we have in the lab is a Rotor-Gene Q 2plex HRM Platform, saying it has green, yellow,...

01 March 2021 8,544 1 View

How to address this issue in the analysis of Real-Time PCR results?

To dear Researchers, I was analyzing a series of concentration for estimation of Real-Time PCR efficiency. The concentration was 1:10. I used MS-excel to evaluate Slope. The result of slope was -8...

01 March 2021 8,683 4 View

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Does anyone have the experience of using Taq Man probes in the QIAGEN Rotar- Gene qPCR machine?

01 March 2021 5,311 1 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

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

MiRNA qpcr problem?

Dear All, mirna primer showing some problem in the melting curve? any idea why? As attached is the melting curve. The forward sequence is obtained from miRBase and reverse primer is universal.

28 February 2021 5,008 4 View