Vishal Srivastava

9 Questions 27 Answers 0 Followers

Questions related from Vishal Srivastava

Is it possible to perform biophysical studies (CD, fluoroscence, DLS etc.) using such purified fractions?

Hi fellow researchers, I am trying to purify a protein of 50KDa for structure function analysis and folding unfolding kinetics using chemical denaturant. I have purified my protein from Inclusion...

03 March 2018 9,802 3 View

How to get pure protein from Ni-NTA which gives protein with lot of contaminants?

I am trying to purify one of my protein of interest having molecular weight around 52 Kda. Upon over expression it comes in inclusion body. So I isolated and washed inclusion body and then...

16 August 2017 8,775 8 View

How do i perform pH optimization of a protease as well as Stability at different pH?

I am working on a protease enzyme and recently i have decided to perform a pH optimization as well as temperature optimization for maximal activity of the protein using azocasein as...

03 May 2017 4,241 4 View

Is it normal getting mutation in Plasmid backbone?

Dear all, I am planning to clone a gene in pET24a(+) Series vector but before going on for cloning i tried to sequence pEt24a(+) null vector from our lab repositiory, but in...

01 February 2017 6,992 3 View

How can I stop the protein coming out in flow through while doing purification?

I am trying to purify protein having molecular weight ~50Kda under denaturing condition through Ni-NTA chromatography and got following profile. Any suggestion to stop protein coming in flow...

20 June 2015 7,597 15 View

How do you solve No expression of a protein problem in E.coli?

My cloned gene in pET23b contains a C-terminal his tag and mature form of protease. Its not getting expressed in e.coli cells. Even after transformation very little colony forms ~1-5 colonies...

21 February 2015 2,809 7 View

What should you do if you get no colonies after transformation in E.coli BL21 de3 cells?

I successfully cloned a gene and checked it by colony pcr and restriction digestion. There after I transformed the construct in E.coli DH5alpha for plasmid prep and isolated sufficient amount of...

18 September 2014 10,056 9 View

How can I determine the restriction digestion of an isolated vector and Insert?

Hello everyone, I prepared my vector (pET23b) and got 10.5ug of Vector dissolved in 150ul of nuclease free water and the insert is about 11 ug in 100ul. i wanted to perform a double digestion on...

29 July 2014 395 4 View

How do I further proceed for cloning?

I had done PCR amplification of my gene of interest using bacterial genomic dna as template. Further gel extraction (thermo scientific) of 200ul pcr reaction volume had given pcr product having...

26 June 2014 4,391 16 View