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Questions related from Tias Saha
If a bacteria bearing the resistance gene for an antibiotic is grown in presence of the antibiotic, how slow will it be compared to its native form (where it does not have the resistance gene...
07 July 2017 801 8 View
I'm using RPMI 1640 media from Himedia. Within 24 hours the colour of the media is changing to yellow even with very low density of cells. I have checked for any kind of contamination, but that is...
05 May 2017 1,495 14 View
I have to purify a bacterial protein from E. coli. My protein's yield is quite low. I have read that batch binding gives better yield, if so, for how much time do I have to incubate my protein...
02 February 2017 4,052 6 View
Isolated plasmid when mostly in supercoiled form eases most of the downstream processes, but is it essential for restriction digestion also? If there is less supercoiled and more of nicked...
12 December 2016 8,627 6 View
I know that beta lactamase produced by the colonies will degrade the ampicillin in vicinity where spurious satellite colonies can grow but how long can unused plates of Luria Bertani Agar media...
11 November 2016 7,961 9 View
I have cloned my gene of interest (WT) in a vector and in the same vector with same restriction sites I have cloned the point mutated gene. I am not getting any expression for the WT whereas the...
10 October 2016 4,091 7 View
In a plasmid maxi prep, many a times, nicked circular species are isolated apart from supercoiled. Can this nick be rejoined by incubating the prep with ligase? I know it won't increase...
09 September 2016 5,623 6 View
If I have a sequence of amino acids. Can its tertiary structure be predicted using any in silico tool? There are proteins which have not been crystallised yet and hence we do not know the...
08 August 2016 4,893 6 View
I feel it contradictory to evolutionary advantage that the two strands of DNA as evolved are anti-parallel but the polymerases that synthesize/ replicate them can do so in only one direction....
07 July 2016 3,705 16 View
a. As protein folding generally starts from N terminus. If hypothetically I delete some parts of its N-terminus, will it still be stable and able to fold properly? b. If I delete 30 amino acids...
07 July 2016 6,010 8 View
I have isolated plasmid DNA by maxipreparation from 100 mL bacterial culture. I have dissolved the DNA pellet in 50 microlitres. It has given me the concentration of 17-20 microgm/microlitre. Is...
06 June 2016 823 8 View
I need a quite a large number of cells to check the expression of certain constructs. Right now I do not have that many cells growing. Can I thaw a frozen vial and without passaging, use them for...
06 June 2016 543 6 View
Ligation is carried out at varied temperatures like 16, 22, 25, 37 degrees and for different time like 16 hrs, overnight, 4-6 hours, 2 hrs, 10 mins. What among all these would work best for...
06 June 2016 4,115 22 View
I have digested my insert (gel purified PCR amplicon) with 2 restriction enzymes and kept for overnight incubation. After which I have put it in 4 degree without doing anything (phenol-chloroform...
06 June 2016 4,526 6 View
I am facing some problem regarding the residual phenol contamination in my DNA samples. I use phenol-chloroform-isoamyl alcohol mix to remove proteins. The 260/280 ratio suggests the presence of...
06 June 2016 8,727 9 View
I have subjected the same DNA with single digestion and double digestion using different buffers to compare the efficiency of the buffers. I have loaded the uncut DNA (maxiprep DNA containing,...
05 May 2016 7,732 12 View
The buffer contains Tris-HCl ph 8.8, which is to maintain the pH for the polymerase enzyme to be active optimally. I understand the role of MgCl2 or other Mg salts, for interactions with the dNTPs...
05 May 2016 1,022 5 View
For plasmid vector, linearization is the indication of complete digestion, but how to check whether the insert which is a linear PCR amplified fragment is completely digested?
05 May 2016 5,666 3 View
I need to know the differences between these 2 vectors. If anyone has the detailed plasmid map of these vectors, please share it. thanks!
05 May 2016 3,447 3 View
I get quite sharp bands of my amplicon, when I loaded some of it to check the PCR product. But when I electrophoresed the samples all together to elute them, although I got my band but it appeared...
04 April 2016 9,583 4 View
I eluted my amplicon from gel and checked its concentration after elution (column based kit) on 0.8% agarose gel. In the eluted product I found another band above my amplicon, which is very...
04 April 2016 4,261 14 View
I checked the pH of a solution using pH strip and it always showed it to be pH 10. Whereas the same solution's pH when measured with a pH meter (calibrated using freshly prepared standard buffers)...
04 April 2016 7,970 8 View
I want to store an agarose gel (with EtBr) for using it in later time. How should it be stored (at 4 degree or simply in buffer) so that it can be successfully used. What is the maximum time that...
04 April 2016 5,052 11 View
I want to study truncated versions of a protein to analyse its domain specific function. So, for detection I need to put some tag at the C terminal as I have N terminal deletions. I have used HA...
03 March 2016 4,111 7 View
I loaded approximately 20ug protein in each lane. After electrophoresis, the outermost lanes appear to have been spread outward rather than following a linear path. What could be the reasons...
03 March 2016 5,364 22 View
I set up a small scale PCR of 20uL to check a set of new reverse primer (RP). I have previously used the same template, buffer, enzyme and other reagents (including same forward primer, FP) under...
03 March 2016 9,009 10 View
How does it vary with different cell types? If I keep the cells longer in trypsin, what damage can be caused to the cells? What is the minimum and maximum time one can safely keep the cells in...
01 January 2016 3,268 12 View
I use electroporation method for transfection. I want to know the factors on which the transfection efficiency depends and what measures can be taken to improve the efficiency?
01 January 2016 9,230 4 View
I transfected HeLa cells and U2OS cells with same DNA (pCDNA GFP) in same amount under similar transfection condition and at the same time. When I checked the transfection efficiency by flow...
01 January 2016 3,167 11 View
I wanted to check the expression of a protein of 14Kda MW by Western Blot technique. I used 15% and 12.5% SDS PAGE gels and semi dry transfer method. No bands were visible even after loading 100...
09 September 2015 10,055 10 View
Why does the freshly prepared media seems to be yellowish red, while the media bottle opened many a times turns purplish red? Even used media (yellow colour indicating acidic pH) when kept outside...
07 July 2015 3,205 1 View
