How can I transform 50 kb plasmid into ecoli?

I must put 50 kb plasmid into ecoli strain. I think 50 kb is too big for heat shock with ecoli. Is that correct? Will electroporation work? Or do I need to use cos sites and lambda phage...

22 January 2021 8,612 3 View

Bacteria strain received has fungal contamination -- how remove?

I received several strains of bacteria from another lab far away. However, one of the samples appears to have high fungal contamination. When I grow it in broth, I see lots of fungal...

03 January 2021 2,749 16 View

New knockout bacterial strain: Can't electroporate it?

I made a new bacterial strain with an unmarked deletion using 2 step homologous recombination (with sacB counterselection). The full protocol is given in the link at the bottom. The protocol...

01 January 2021 1,580 4 View

Western blot: what causes patchy, uneven background?

Sometimes my western blots have very uniform/even background, other times they're very uneven and patchy like in the attached image. Why is this? I thought my blots were drying out during...

05 December 2020 8,656 5 View

Licor Intercept Blocking Buffer -- sterility and reusing antibody dilutions?

In the past, I've always used Licor's old blocking buffer that included sodium azide. I would dilute my antibodies in the buffer, stain, re-collect the dilution, and re-use it a few times. The...

28 November 2020 5,795 3 View

What mutations in oriE increase the plasmid copy number?

I have a low copy plasmid (ori=pBR322) and I wanted change it to high copy (like ori=pUC). This paper seemed to say that only one mutation was required to increase the copy number: High copy...

13 November 2020 7,292 4 View

Western blot: adjusting volume of buffer for blocking/staining?

For western blots, I normally stain with 10 mL buffer with 1:1,000 dilution of my specific antibody. Recently, there is an equipment change in my lab and 10 mL isn't enough volume to cover the...

27 October 2020 2,749 7 View

Is this correct molecular docking?

Hello, buds I need your help to interpret the attached data. R group in serine is CH2OH, but the data show the two O in the group with hydrogen bonds with a molecule. Is this correct?...

02 October 2020 7,787 6 View

Concentrating protein: comparison of methods?

I must concentrate and purify a lot of dirty lysate and secreted protein samples. I just want to run them on protein gels for western blots. I'm trying to decide the best method of...

10 September 2020 2,911 8 View

Oxygen detection using Ru(bpy)?

I am trying to detect if my nanoparticles generate oxygen. Is there a way I can use Ru(bpy) and obtain an emission spectra? (Any other "quick and fast" method I can use to detect oxygen generation?)

26 August 2020 1,942 3 View

Enhanced Yellow Fluorescent Protein (Aequorea victoria) DNA sequence?

I am on the lookout for the Enhanced Yellow Fluorescent Protein (Aequorea victoria) DNA sequence. Does anyone know where I can find it? Thank you in advance

03 March 2021 3,568 1 View

Can linear DNA be used as template for site-directed mutagenesis?

Is it possible to induce site-directed substitution mutation by quick-change method on linear dsDNA? or it has to be cloned in some vector? If yes, should it be treated with the Dpn1 enzyme...

03 March 2021 401 4 View

How to increase a model accuracy ?

dear community, my model is based feature extraction from non stationary signals using discrete Wavelet Transform and then using statistical features then machine learning classifiers in order to...

03 March 2021 6,994 5 View

How to normalise data on OPM package from R Studio ?

Good afternoon, I recently used OmniLog from BIOLOG for my experimentations : I tested the metabolism of different strains on 2 types of plates. I have 16 strains of 3 different groups...

02 March 2021 3,584 1 View

New Journal, "Integrative Psychological and Behavioral Science" -- do you not know, and have you not seen, this done before?

Question to you and THEM, the New Journal, "Integrative Psychological and Behavioral Science" -- do you not know, and have you not seen, this done before? There appears to be a core problem for...

02 March 2021 3,024 2 View

Which is a lucid standard comprehensive book to guide on psychological scale construction and its standardization?

02 March 2021 7,082 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

Does anyone know HRM channel in a 2 plex QIAGEN Rotor- Gene Q qPCR can be used as a separate channel?

I am going to have 3 different probes in my qPCR work that I am going to do. But I realized that the machine we have in the lab is a Rotor-Gene Q 2plex HRM Platform, saying it has green, yellow,...

01 March 2021 8,544 1 View

Which cell line should we use to perform biological dosimetry experiments?

We are preparing some experiments based on irradiating cells under different conditions in order to evaluate the effects in terms of DNA damage, genetic expression, etc. As our project is...

01 March 2021 3,355 3 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