How can I measure PKA activity in the yeast cell?

Hi everyone. I would like to measure PKA activity in the yeast cell. Could you give me any suggestions? Thank you!

05 June 2016 3,771 0 View

How to dissolve polylysine?

Hello everyone, I wanna attach my cells on to the slide, so I used poly-lysine to coat the slide. I made poly-lysine solution in ddH2O with concentration was 1mg/ml and stored at -20oC. After...

01 February 2015 1,309 6 View

Express yeast protein in E. coli ?

Hello everyone. I want to express a yeast protein in E. coli. Should I have to optimize codon usage when express in E. coli? In case of a long gene, how can I optimize the codon usage? Thank you...

01 February 2015 1,674 0 View

Is it difficult to express small recombinant protein in E. coli?

31 December 2014 6,670 6 View

Should I insert a simple peptide between GFP and target protein?

Dear everyone, I want to tagged GFP to my target protein. Do I have to insert a simple peptide such as GGGG between GFP and the protein? And how many amino acids is enough? Thanks you so much.

11 December 2014 7,104 8 View

Do you know where to get information about the yeast backgrounds?

Hi everyone, I am working with BY4741 background but it seem to be not working well. So I want to change to another background such as CML128,... But I do not know much about the other backgrounds...

11 December 2014 1,414 0 View

I'm making the trypsin 0.25% solution for cell culture. The recipe is 2.5g trypsin and 0.2g EDTA.4Na in 1l HBSS 1x. My lab only has EDTA.2Na. How can I change EDTA.4Na to EDTA.2Na?

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04 May 2013 6,078 9 View

Why does blocking with my immunogenic peptide cause nuclear fluorescence in ICC?

Hi all, I am validating a polyclonal antibody raised against human ANT4, through WB and ICC. In most of my controls, the antibody shows very little off-target binding, but when I pre-incubate the...

03 March 2021 5,999 3 View

Have someone successfully used the ER alpha (Santa Cruz sc-8002) antibody in immunostaining ?

Santa Cruz now recommends this new antibody (F-10) but I can't find a publication using this reference. I'm working on human et non human primate hypothalamus and so far, it doesn't work. Is...

02 March 2021 10,003 2 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

Does anyone know an alternative product for cardiac Troponin T antibody for Immunofluorescence (Thermo # MA512960)?

We have been using Thermo Fisher anti-cardiac Troponin T antibody for immunofluorescence staining in FFPE mouse heart sections but it was recently discontinued. The literature shows it to be the...

01 March 2021 4,937 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

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

Precipitate after adding laemmli buffer to protein solution containing Potassium Acetate?

Hi, I am running a size exclusion chromatography experiment with a buffer containing Potassium Acetate as a salt. I analyse these fractions through SDS-PAGE. After boiling my SEC fractions in...

01 March 2021 2,622 3 View

What is the most suitable server for predicting protein structures in bacteria?

I have used the i-Tasser several times, however it has been unavailable for several days. I tried the swiss-model, but the output was not very pleasant due to the model used. Is there any other...

28 February 2021 4,521 3 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

Are protein samples in SDS compatible with ELISA?

I have used an AllPrep DNA/RNA/Protein Mini Kit QIAGEN kit to extract RNA and protein from my samples. At the end of the protein purification, I resuspend my protein in 5% SDS. Will these samples...

28 February 2021 7,370 3 View