What's the longest insert you put into a plasmid via quickchange-like methods?

We would like to put a 12-20 aminoacid linker into a recombinant protein expressed in E coli. We've had good experience with Quickchange-like protocols; but never tried with 36-60 nucleotide...

02 March 2019 3,185 4 View

Is Lithium Lauryl Sulfate OK to run K+-containing samples on SDS-PAGE?

Hi all, Can one use Li Lauril Sulfate (LDS) in loading buffer to run protein samples containing high concentrations of KCl, on SDS-PAGE? Does one need to replace SDS in the running buffer and in...

01 February 2019 1,614 6 View

Can you recommend a tabletop front-loading autoclave?

Can anyone recommend a front-loading tabletop autoclave which could be used to sterilize up to six 2L flasks at once with steam? Looking for something that is robust, simple, and ages well (i.e....

07 August 2018 6,653 0 View

Is there a chemically defined, FBS-free media formulation for HCT-116 and U2OS cells (adherent human cells)?

FBS-free medium is what I am looking for. If defined growth factors are required, I can express them (if they are available in recombinant form). Artificial meat companies seem to have figured it...

06 July 2018 7,101 4 View

Suggest a method for harvesting of adherent cells from hollow fiber modules? Does scraping work? Would using hydrophobic fibers help?

We are looking for a cost-effective way to grow adherent cells (HCT116, US2OS), in amounts of 1*10^10, for subsequent isolation of nuclei and preparation nuclear extracts per Dignam 1980....

05 June 2018 7,168 5 View

Has anyone tried using standard hollow fiber PES hemo-dialysis cartridges to culture adherent mammalian cells?

The surface area (1-2 sq meters) and the basic specs (excluded volume, PES memberane chemistry, pressure drop, non-fowling, sterile, QC, Mw rejection) of hollow fiber dialysis filters appears...

05 June 2018 9,437 0 View

Does anyone Happen to know what BioRad's iProof PCR enzyme is (beyond what's in the catalog description)?

09 October 2016 1,608 2 View

Does anyone have experience with the mini8 PCR cycler?

09 October 2016 3,488 0 View

How to control pH and composition of PEG solutions?

Hi all, I am trying to concentrate a DNA sample using dialysis against 50% PEG, through a 10K cut-off membrane. I made a 50% solution of PEG20K supplemented with 10mM Tris (target pH=7.8), and...

08 September 2016 3,873 10 View

An affordable and reliable thermal cycler?

02 March 2016 3,750 5 View

Protein-aptamer gel electrophoresis help, please??

Hi, I have problems with running gel electrophoresis. I have tried agarose gel electrophoresis and native PAGE. I have two proteins, which have molecular weights of ~30kDa and ~180kDa and two...

03 March 2021 4,275 4 View

Buffer to disrupt platelet alpha granules for PF4 quantification?

Hello! I want to quantify by ELISA the secreted (from platelets poor plasma) and the non-secreted (from platelet lysate) PF4 before and after TRAP stimulation. I will use the ELISA from R&D...

03 March 2021 1,499 2 View

Buffer Exchange using column packed with sephadex G-25 effect on protein concentration?

Hello, If i am doing a buffer exchange for an antibody of 1mg/mL, does the elution lose protein in the process of buffer exchange? For example, if i flow through 500 uL of 1mg/mL sample, and...

03 March 2021 6,299 3 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

No title

03 March 2021 5,360 2 View

Gel electrophoresis result for total RNA samples, 1.5 Agarose gel was used, How to improve the integrity of RNA?

Gel electrophoresis, RNA degradation, RNA extraction from fresh tissue

02 March 2021 5,433 5 View

How to regulate SSCP temperature in simple PAGE?

I'd like to perform single-strand conformation polymorphism (SSCP) in my thesis, however I cannot control the temperature of the vertical PAGE since we are using the conventional tanks. Is there a...

02 March 2021 9,157 1 View

How optimize the Size Exclusion Chromatography?

Hello! I'll do a size exclusion chromatography, but I only have an open column, and I'll perform the peptide extraction from yeast, using buffer lysis (sodium phosphate 50 mM/NaCl 30 mM/DNAse and...

01 March 2021 2,215 2 View

Why are my bands weird in this Agarose gel?

Can someone please give me some possible things that could go wrong? Here is my recipe: 0.5g Agarose 50 mL of TAE 1x 1 uL ethyl bromide. Gel was run at 100V for 1 hour. The buffer used is also TAE.

01 March 2021 9,952 3 View

What do I do with DLS peaks that are from the buffer?

I'm looking at the aggregation of my protein sample using DLS. Unfortunately, my buffer (20mM HEPES) also results in a set of peaks. These are at approximately 1 and 1000 d.nm. The lowest peak...

01 March 2021 9,015 2 View