Reference for 0.5 McFarland standard corresponding to 0.08 to 0.13?

Hi, I have a question about the 0.5 McFarland standard. In different websites I read that the for the 0.5 McFarland standard, the absorbance at a wavelength of 625 nm should be 0.08 to 0.13. I...

08 September 2018 4,826 1 View

How do I determine if specific points significantly deviate from the linear regression in a scatter plot?

Hi everyone, I would like to know how I can determine if a point significantly deviates from the linear regression when I plot the data in a scatter plot. I am using excel and I made a scatter...

06 July 2018 5,232 8 View

How to measure density with ImageJ in images that have different magnification?

Hi all, i am new in image processing and using ImageJ. I want to measure the density in areas of pictures that were originally taken at a different magnification. I attached an example that I hope...

05 June 2018 7,206 4 View

Theoretical question about western blot?

Hi, I am new with Western Blot and I need a few clarifications about the use of blocking and incubation solution. I understood that blocking is done to saturate the membrane with proteins...

11 December 2017 4,382 4 View

Preparation of sodium phosphate buffer?

Hi, I am preparing a sodium phosphate buffer solution at 0.1 M pH 7. For the preparation I am following this instruction that requires to mix 1 M Na2HPO4 and 1 M NaH2PO4 like this: 57.7 ml 1 M...

10 November 2017 6,517 3 View

How to calculate the enzyme activity from absorbance?

Hi, I have a question about how to calculate the enzyme activity from absorbance. I have absorbance values for different concentrations of my samples at different times. As far as I understood, I...

08 September 2017 1,623 4 View

Can you help me with understanding how to determine the beta lactamase activity?

08 September 2017 1,180 5 View

Which statistical test is most appropriate for my experiment?

08 September 2017 8,159 15 View

Cutting bands out of the agarose gel?

05 June 2017 3,063 1 View

Optimization of ligation?

05 June 2017 3,601 3 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