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Questions related from Shubhashish Chakraborty
Hello. I am working on a kinase that binds to ATP. I cocrystallized the protein with the ATP analog AMP and solved the structure. However, while performing ITC, I am not getting a complete...
31 August 2022 9,059 3 View
I have done molecular replacement to build a protein structure however even after a lot of refinement and protein chain fixing the difference between rfree and rwork is around 7%. While solving...
28 January 2021 5,956 6 View
Hello, The rmerge for my scaled data is 72% or 0.722 for outershell. I tried to refine using the same data set however the electron density doesn't look good. So what can be done to lower the...
22 December 2020 8,051 6 View
I have diffracted a protein crystal with a resolution of 2.2A. While solving the structure using molecular replacement I am not getting density for 2 residues that form a turn. These two residues...
09 December 2020 8,798 8 View
I am solving a crystal structure using molecular replacement in phenix. The mutated residue is Glycine to gultamate and present in a turn. While refining in the first round I selected rigid body...
18 November 2020 7,791 6 View
Hello, I am working with a kinase domain of a membrane tyrosine kinase receptor. This protein undergoes auto-phosphorylation upon binding of a ligand. So is there any assay available by which I...
23 July 2020 4,487 3 View
I have induced a mutation of cysteine to arginine in a zinc finger motif of a protein. However rather than a missense mutation that was expected the mutation lead to generation of truncated...
18 July 2019 872 3 View
I am trying to express a kinase domain of a protein. The construct is in pET-His10 TEV LIC vector and I am using pLysS cells. The cells were induced with 4mM IPTG and expressed at 22C. When I...
08 May 2019 5,065 8 View
In my construct I have two His tags, one at N-term and another at C-term. There is a TEV site between the N-term His tag and my gene but no cleavage site for the C-term. So, do I need to add...
05 April 2019 9,719 4 View
Is it that vectors with T7 promoter do not grow/transform well in rosetta or codon cells?
01 March 2019 5,429 3 View
I am not getting colonies when I had transformed my construct in rosetta 2 cells whereas transformation worked in Bl21-pLyse cells. I had used the same heat shock protocol for both. The one change...
28 February 2019 4,720 8 View
I am working to purify and determine the structure of a membrane protein with post transnational modification such as glycosylation. So, can anyone suggest any bacterial strain (and/or reference...
06 December 2018 3,516 5 View
Just as lysine acetylation dilutes the positive charge of lysine and leads to histone loosening, what actually drives for transcriptional activation or repression when lysine within histone is...
04 April 2018 3,836 2 View
Can any one provide me appropriate protocol for preparing crude mouse tissue lysate for western blotting. The protein of interest resides both in cytoplasm and nucleus so which lysis buffer would...
05 February 2017 2,048 7 View
How much DNase volume should be used to effectively degrade all of DNA present in a 5ml bacterial cell culture?
09 January 2017 5,514 3 View
Please refer me any protocol for developing colon cancer animal model in particular to mice and zebra fish where I can study metastasis.
14 December 2016 6,615 3 View
Is their any protocol or reagent by which I can store mice colon tumor samples at -20 or 2-4℃ and latter extract intact RNA and protein.
03 October 2016 1,873 3 View
I want to develop mice/rat model for colorectal cancer. The protocols I had studied till now result into tumors after around 4 to 6 months. So is their any other standard protocol where I can...
24 July 2016 1,915 1 View
Hi all. I want to insert an enterokinase site in my plasmid using PCR mediated site directed mutagenesis, so is thier any standard protocol for this. My insert size is 14nt and plasmid 4.2kb
24 June 2016 9,089 4 View
Is their any maximum limit for an insert size in terms of nucleotides for mutagenesis? My insert size is 14nt and plasmid size 4.2kb so will their be a problem?
23 June 2016 1,271 5 View
Can anyone tell me if their is any method or protocol to dry sds page gel after sample has been ran. The gel is to be stored for any further analysis.
07 June 2016 9,927 5 View
Is their any easy method by which I could separate microbial cells from saliva samples of human. It would be really helpfull if someone can share related protocol.
18 May 2016 9,735 3 View
