Anamika Singh

8 Questions 13 Answers 0 Followers

Questions related from Anamika Singh

What if the sequence from colony PCR is positive but plasmid isolated from the same colony did not get whole sequence? Can I use that plasmid ?

Hi All, I have done pENTR Directional Topo cloning using transfer PCR method. When I was screening the colonies I got positive amplification. I sent the PCR product for the sequencing. Along...

12 December 2018 7,397 6 View

Why is that rev primer of pET28a vector gives different read in sequencer for recombinant gene while there is a full gene seq with T7 promoter?

I have sequenced the different colonies with the forward primer (T7 promoter primer) and getting the same result whole gene with N-terminal His tag..But I was not getting any expression so did...

11 November 2017 7,098 1 View

Why is that rev primer of pET28a vector gives different read in sequencer for recombinant gene while there is a full gene seq with T7 promoter?

I have sequenced the different colonies with the forward primer (T7 promoter primer) and getting the same result whole gene with N-terminal His tag..But I was not getting any expression so did...

11 November 2017 6,835 1 View

Why does a 19.6 kda protein show single band in both the non-reducing & reducing, but different bands in glutaraldehyde cross linking experiment ?

I have done cross linking experiment with 0.1 % glutaraldehyde with native, reduced and oxidized form of protein (~19 Kda). In all case several bands were observed of higher molecular weight. what...

04 April 2015 9,678 7 View

If 20 kda protein shows single band in both the non-reducing, reducing, but showed different mass peak in MS spectrum What could be the reason?

MS spectrum showed peaks at 10kda, 20kda and 40kda. is protein under going oligomerisation or getting degraded ? please do suggest me

03 March 2015 8,121 1 View

Protein precipitates and degradation during dialysis

My 27kda prt(pI 8.2) precipitates during dailysis when left o/n at 20 degree n at RT. The elution buffer 20mM tris ph 7, 100mM NaCl, 250mM imidazole pH 7.

08 August 2012 7,258 16 View

Recombinant Protien purification

hi all actually i am facing problem while purifying rec. protein of plant origin.The protein is coming in pellet. i tried to solubilise it, it works but problem of refolding still persist i.e....

12 December 2010 6,417 1 View

The colony pcr is positive but sequencing failed.

Hello everyone. Recently I faced a situation, and would like to ask how to overcome it. Vector : pENTR Insert : 600 bp I got many colonies and my colony PCR (with Gene-specific primers) came out...

01 January 1970 9,502 6 View