Do plasmid DNA also get extracted alongwith genomic DNA during the genomic DNA extraction,?

Is there plasmid DNA contamination in genomic DNA preparations?

06 July 2015 5,942 0 View

Is there any chemical difference in Heparin resin used in HiTrap Heparin HP and Heparin sepharose 6 FF from GE healthcare?

Hitrap Heparin HP is coupled to the Sepharose High Performance base matrix via the N-hydroxysuccinamide coupling method and in case of Heparin sepharose 6 FF, Heparin is linked to the Sepharose...

31 December 2014 6,439 1 View

Can we sequence DNA on sequencing gels using fluorescent labels?

Can fluorescent labels be detected with a phosphor imager? We have Fujifilm- FLA 9000 and Biorad Screen Eraser-K set up for detection.

10 November 2014 8,774 3 View

Can we use M13 Reverse and M13 Forward (generally used for sequencing insert in pUC18 MCS) for seuencing/amplifying fragment of M13mp18?

Do these two primers have a sequence in M13mp18 DNA, as the name suggests that these are from M13? Please provide an explanation.

06 July 2014 6,939 1 View

How can I remove DNA from bacterial crude lysates?

06 July 2014 698 12 View

Does anybody know of a plasmid or DNA vector which lacks or has a single TTAA site?

As it is a 4 bp sequence, it frequently occurs in most plasmids. If anybody knows of such plasmid lacking or having single TTAA site?

02 March 2014 7,422 1 View

How do we equilibrate the millipore protein concentrator while using it for the first time?

Can it be reused, and if yes, how many times?

01 February 2014 5,205 8 View

Can coomassie stained gels which have been destained, be stained again? With coomassie stain and silver stain?

I have stained my SDS-PAGE with coomassie blue but bands are very light. Probably, the stain was not good. Can the gel be stained again with new coomassie or silver stain? Will the staining be...

01 February 2014 8,373 12 View

Do proteins retain activity after acetone precipitation?

I need to concentrate my protein samples for running SDS-PAGE as well as checking activity.

01 February 2014 9,457 5 View

What is the binding capacity of DEAE cellulose?

Concentration of sample is 10-12 mg/ml and I am planning to use 4 ml DEAE cellulose resin. I need to know the amount of protein sample required to be loaded on the column to get good resolution?

01 February 2014 4,914 5 View

Does anyone have a recommendation for yeast reporter gene plasmid/protocol?

Hi, could anyone recommend a plasmid and/or protocol for reporter gene assay in S. cerevisiae? I want to assess the effect of growth conditions on a transcription factor, so I want to clone it`s...

01 March 2021 210 1 View

Why might my GFP tag be dissociating from my plasmid after transfection?

I have a set of stably transfected cell lines all transfected with plasmids containing GFP tags on the C terminus. During a western blot using anti-GFP antibody, one of my plasmids has dissociated...

01 March 2021 9,310 4 View

Expression cloning by using cDNA,do I need to modify the cDNA first?

I am going to have a expression cloning of mammalian gene by using shuttle plasmid to transforming the E.coli However I don't know I should only inserting the Coding sequence ,or I can...

28 February 2021 5,440 3 View

Problem with shRNA transfection. Why are only two plates getting contaminated instead of all?

I transfected my LNCaP-WT cells with 3 shRNA plus their NTC two weeks ago and split two puromycin selected cell plates on Friday last week(Feb 26). I checked for GFP in the cells, and they all...

28 February 2021 4,949 3 View

PUC19 as a transfer plasmid for lentiviral delivery?

Hello, Is it possible to use pUC19 as a transfer vector to be packed in using the second generation viral particles packaging system( pMD2.G; psPAX2 plasmids)? As far as I understand it there is...

28 February 2021 4,868 2 View

Did anyone ever try using pLVX-TetOne Plasmid with a copGFP as a selection marker instead of Puromycin?

When using a lentiviral vector for inducible expression of genes using the Tet-On system. In the literature, I saw a widely used of pLVX-TetOne-Puro Plasmid, is there a reason why people prefer to...

25 February 2021 1,011 3 View

Why do the PCR bands from the cDNA templates (extraction from the cell after transfection) show different size from the band of the plasmid template?

After transfection with the plasmid ( linearized ) and subcloning of the cell lines, RNA was extracted from the cell and then reverse transcripted to cDNA. When PCR reactions were run to verify...

25 February 2021 5,712 3 View

How do I transfect a large plasmid into PC12 cells with nerve growth factor (NGF) receptor?

I have a large 16 kb plasmid, which I need to transfect into PC12 cells. Lipofectamin 2000 didn't work and with GFP alone the transfection rate is very low. I also tried the neon invitrogen...

25 February 2021 6,635 5 View

Problem with ligating 2.8 kb insert into 8.5 kb plasmid?

I am trying to clone 2 copies of the same mammalian gene into the pSF-CMV-Ub-Puro Ascl (contains HindIII, KpnI and NheI cut sites) plasmid. This is what the final product is supposed to look like...

24 February 2021 6,310 3 View

Best plasmid could be used as a standard curve for AAV titration?

I have been trying to have a good AAV titration for my production and I have tried different plasmids to set up as a standard curve but each one behaves differently. i am trying to use ITR for...

23 February 2021 5,167 3 View