38 Questions 29 Answers 0 Followers
Questions related from Robert Adamu Shey
I am purifying a protein following bacterial expression by affinity chromatography. Unfortunately, in addition to the expected band (about 34 kDa), I am also seeing another protein which is about...
16 September 2024 9,492 2 View
I am trying to investigate the thermal stability of a protein I work with using the TSA with SYPRO Orange. However, even though my protein produces a peak at about 43 degrees, it shows very high...
16 September 2024 5,597 4 View
There are many different software tools available to visualize and sometimes to edit protein structure files, eg. in PDB format. I am working on a protein with close to 1900 amino acids. I have...
28 July 2023 6,324 3 View
We have been trying to purify a chimeric of MW, 31kDa in insect and mammalian cells using the pOET5 and pTT5 vectors respectively. Unfortunately, we can detect very low levels of the protein in...
26 April 2022 4,361 3 View
So I am working on 3D structure prediction for some proteins and need to validate the structure using amongst others a tool that does Ramachandran plot. Any ideas about possible tools?
14 October 2021 9,442 4 View
I am interested in determining some protein structures using bioinformatics. There's been a lot of stories about DeepMind AlphaFold? Anyone has any experience working with it? Is there an online...
28 July 2021 2,048 6 View
Working on some phylogenetic trees, and wanted to change the scale of the tree from 0.05 to 0.005. Anyone has experience with this?
16 June 2021 4,050 1 View
We are trying to isolate cf-DNA from urine to no avail. Anyone with an in-house protocol that works?
01 June 2021 3,216 3 View
I am doing some work on a couple of gene SNPs that could be involved in susceptibility and resistance to infectious diseases. Does anyone know any tools that could be used to detect potential...
20 March 2021 4,608 5 View
Hello everyone, so I expressed this protein and I am trying to further purify using SEC and still having the impurities in the fraction I expected to have the protein. How do I eliminate these. I...
28 April 2019 8,242 3 View
I am doing RNA, DNA and protein extraction from the same sample and my protocol recommends this water saturated phenol:chloroform mixture. I am not not very versed with how to go about with the...
30 April 2018 2,263 7 View
Hello everyone, I need some help on assigning ProSA Z-scores to the quality of the structure validated. My modeled structure had a score of -3.93.
18 December 2017 9,789 0 View
Hello everyone, My aim is to isolate surface proteins from a parasitic nematode, however I have looked at protocols on line and most seem to be the same for 'cuticular' and 'surface' proteins,...
16 December 2017 8,550 3 View
Hello everyone, I am about doing an antigen capture ELISA and a little confused about which buffer can be used in absorbing the capture antibody to the microplates. I am using Nunc MaxiSorb 96...
13 November 2017 4,084 3 View
Hello everyone, So I planned this ELISA to do, unfortunately I coated plates and for some reason I am not able to continue with the experiment as planned. So basically, instead of coating...
10 October 2017 3,040 3 View
Hello everyone, I am doing gel filtration chromatography with a protein I purified from Ni++ IMAC. The protein however seems to be coming out before the expected elution volume. However, when I...
17 September 2017 1,318 7 View
Hello every one, My question is based on the definition of memory B cells below: "The immune system can remember a previously experienced pathogen and can evoke an enhanced response to reinfection...
17 September 2017 6,545 5 View
Hello everyone, Some questions I asked recently on this forum have been related to antibody-antigen reactions. Now I wish to find out if " ..Antigen-antibody interaction, or antigen-antibody...
25 August 2017 3,325 4 View
Hello every one, I am interested in doing some analyses on the genome of some organisms but the downloaded format I have have this ...genomic.fa.gz as extension. What can I do to have it in the...
20 August 2017 2,123 4 View
I wish to find out if all secreted proteins have a signal peptide. Also, if yes, are these signal peptides always cut?
19 August 2017 7,436 2 View
I am working with a pET30a+ construct and my protein is C-terminal His-tagged. My protein is forming inclusion bodies and I need to get a protocol for isolation of a pure protein from this. An...
01 August 2017 5,128 3 View
Hello everyone, I want to do ELISA to detect IgE in serum against an antigen I an working with and some one indicated that abundant proteins like IgG and HSA could either block or lead to...
22 July 2017 3,874 3 View
I want to draw a phylogenetics tree using a protein I am working with and its homologs using MEGA7. After the alignment using MUSCLE or CLUSTALW, there are so many options on the MEGA program for...
06 July 2017 4,683 4 View
Good day everyone, I am to construct a phylogenetic tree with proteins but some of the homologs of the protein I am working with are truncated. How do I deal with these?
02 July 2017 5,963 4 View
I am working with a recombinant protein from a parasitic nematode we expressed in bacteria and we have raised antibodies (affinity purified) against this protein. However, we have tried doing...
30 June 2017 9,136 6 View
I need to construct a cartoon structure for a GPCR I am working with.
27 June 2017 4,412 4 View
Hello everyone, I intend to detect genetic mutation using Single-Strand Conformational Polymorphism and the loading dye components from Cold Spring Harbor Protocols is listed below: 95% (v/v)...
01 June 2017 5,612 2 View
I saw a protocol for recombinant protein expression that recommended working in larger volumes (1L cultures) without antibiotics after inoculating overnight in antibiotics. How practical can this be?
06 September 2016 8,269 3 View
I am working with a parasite who total protein contents I have to analyse both by PAGE and Western blot. Can anyone suggest a protocol for going about this. I will be grateful. Thanks
15 October 2015 7,583 1 View
Dear All,I seem to have lost the plasmid DNA of one of my clones for protein expression.However, I do have a glycerol stock of BL21(DE3) cells transformed with the clone. Would it possible to a...
17 August 2015 8,422 6 View
Hello everyone, I am working with a 25 kDa protein which is 6xHis tagged. I have the gene coding for the protein in pET15b and expression is being done in BL 21 DE3. I tried purification and had...
07 July 2015 935 5 View
Hello everyone, I am working on a GPCR which we initially failed to express in HEK 293 cells. Following consultations with some colleagues we switched to CHO K1cells but we detected proteins of...
10 June 2015 6,880 5 View
I have been working with HEK 293T cells so far for expressing a protein. I was however recently asked to try CHO cells since these may a better expression system for my protein which failed to be...
06 June 2015 8,152 3 View
I have a GPCR that I am working on. I have used a number of databases to classify it by bioinformatics but the webservers seem to be "confused" about it's class. I have used GPCR-CA for example...
06 March 2015 4,557 3 View
I am working with a putative protein whose homologues in the databases are predicted to be intimal thickness related receptors. However, I do not know much about such and very few papers are...
24 February 2015 5,602 0 View
I have 3 synthetic peptides that I am using for ELISA: analysis indicate that all of the have good water solubility. They have net charges of -1, -3 and -1.9 at pH 7 with corresponding PIs of...
18 February 2015 7,280 3 View
I have a protein characterising and it happens to be a GPCR from bioinformatic analyses of the Class A. I have the gene coding for it in pET 15b vector and have tried to express it by IPTG...
08 April 2014 8,564 2 View
I am working on amplifying and mass expressing a plasmid in bacteria using a pET 15b vector but I have not been able to achieve this. I have tried many options but the results are still negative....
21 February 2014 5,861 2 View
