How to get going smRNA FISH on fresh frozen breast tissue?

Hello, I am having difficulties to get smRNA FISH going on fresh frozen breast tissue. The technique worked well on MCF7 cells. We tried the same probes on tissue sample, and we haven't had...

05 June 2019 7,841 0 View

Kit or protocol for Bisulphite conversion of DNA from breast FFPE?

Does anyone have experience with bisuphite conversion of DNA from Breast FFPE sample. I have been using Qiagen direct Bisu FFPE kit with deparafinisation buffer . I am facing difficulties in...

02 March 2018 524 0 View

How can one ensure ligation of a 300bp fragment into 5369bp host?

09 October 2014 3,484 16 View

How to troubleshoot Site Directed Mutagenesis?

I did SDM with double primers PCR. This was followed by dpn1 digestion.The gels showed amplicons but when I transformed there were no colonies. The polymerase used was pfu and Tm was 60 degrees...

31 December 2012 1,081 17 View

Docking

I would like to dock an enzyme with its substrate. Can anyone suggest any free software which I can use and which has a tutorial provided?

06 July 2012 6,756 22 View

How to perform MSA for two domains of a single enzyme by clustal X?

I have been trying to do clustalx for my sequences of Polyketide synthase. Many of my sequences have two Acyl Carrier protein (ACP) domains. The problem is that clustal x is not accepting the...

05 June 2012 6,283 17 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

Can two EC50 values with different hill slopes be compared?

Hello, I fit a classical dose response curve Y=Bottom + (Top-Bottom)/(1+10^((LogEC-X)*HillSlope)) to my data (not related to inhibitors etc). The model I use is purely qualitative but it fits...

23 February 2021 8,155 5 View

Will SARS-CoV-2 Spike be broken into S1 and S2 by denaturing conditions?

Hello, I am performing a western blot of my SARS-CoV-2 Spike-pseudotyped lentiviral particles. I denature my virus by adding concentrated virus to 2X Laemmli buffer at 1:1, then boiling for...

17 February 2021 4,844 4 View

What is the best temperature to coat an ELISA plate?

Hello, I want to control the temperature of my ELISA plate during incubations to get consistent results. In general, what is the best temperature to coat the plate with antibody? I have been...

16 February 2021 2,377 7 View

Is there a way to tell based on appearance which protein crystal should diffract the best?

I have had unheard of success with protein crystallography lately from a super successful protein expression and purification batch. I have attained a lately reproducible vast amount of crystals...

09 February 2021 5,155 5 View

How to measure the concentration of a labeled protein with no tryptophan or tyrosines?

Hey I'm trying to measure the concentration of a protein I've purified and then labeled with Atto 647 flourescent dye. The protein has no tryptophans or tyrosines (although does have...

04 February 2021 4,482 4 View

How to solve error "Cannot find position restraint file restraint.gro (option -r)." on GROMACS?

Hi, I am about to do energy minimization but the above error occurred, I checked my .mdp file and .top file but could not find the problem. TOP file ; Include forcefield parameters #include...

01 February 2021 780 3 View

Why is Fermented Plant Juice (FPJ) always diluted with water? Could it be diluted with other solvents such as rice water?

I am currently making a research paper with my classmates in Philippine Science High School and we are trying to find references that mixed Fermented Plant Juice with other solvents instead of...

17 January 2021 5,099 4 View

Any stable dye to color acrylamide hydrogel beads?

Hi! We are generating hydrogel beads(50-60um) using microfluidics as single-cell beads. To simply all the experiments (especially the centrifuge), I am looking for a dye. The dye should be stable...

14 January 2021 4,383 8 View

How to phosphorylate specific residues on pymol?

Hi everyone, I am using the pyTMs plug in on Pymol to phosphorylate a particular threonine in my protein. I am having difficulty in selecting only one or two residues - it seems I can only...

05 January 2021 230 3 View