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Questions related to Cloning
I am generating brain organoids with two different iPSCs cell lines. I have made many times brain organoids with one of these. However, when I start an experiment with other iPSCs cell line, some...
26 November 2023 7,365 7 View
Respected expert, as I have done antioxidant assay for zinc oxide nanoparticle using medicinal plant Serriphidium kurramense. In this assay I have used Ascorbic acid as standard and DPPH as free...
24 November 2023 2,538 2 View
What is the best way of metagenomic analysis using multiple rDNA amplicons (e.g. multiple 18S and ITS for each sample)? What methodology would you use to reliably normalize and integrate the data...
22 November 2023 8,842 3 View
I want to synthesize about 2.5kb cDNA and I need a suitable kit.
21 November 2023 5,170 4 View
My Western Blot shows LANES OF PROTEINS instead of the band I'm looking for. I tried stripping it with RESTORE WESTERN BLOT STRIPPING BUFFER from Thermofisher but it still shows up like this. Any...
21 November 2023 1,837 2 View
Since one of the proteins contains Leucine-rich repeats (LRRs), this region may exhibit structural flexibility that could impact the binding interactions. Do I do molecular dynamic simulations on...
19 November 2023 7,954 0 View
Dear I performed Western blot on full mouse heart lysates of MHC-cre mice. I tested 2 different genes, and in both Western blots I had an unexpected band at 70kDa. Does someone has an explanation...
17 November 2023 7,841 0 View
I am doing cloning of a big bacterial insert (3705bp) into a vectors of varying sizes ranging from 3017bp to 3469bp for my bacterial two hybrid experiment. Among other problems with my cloning I...
16 November 2023 7,077 4 View
I need to get one single band from my sample to sequence the target . what should I change? I changed annealing and DNA concentration, time of each cycle and used different PCR master mixes.
06 November 2023 1,452 4 View
Hi all, I have a naive question and would like some opinions. I performed 2 flow cytometry runs with different samples, and I found more dead cells in my second run. My question is will the...
06 November 2023 3,098 2 View
I have an adenovector but the titration is very low and I need more of it.
06 November 2023 3,574 0 View
I have one experiment need flow cytometry, but we only have 1 non-conjugated antibody. Apart from buying a new antibody for flow cytometry. Could I use secondary antibodies in flow? How can I do...
03 November 2023 6,787 1 View
Dear Community, we have been struggling this year with purification of some antibodies from serum samples and or HEK293T/Recombinant produced antibodies. We have attempted ProteinG,...
02 November 2023 6,926 3 View
We want to complement our indels study in primary cells after CRISPR/CAS9. We are looking for a cheaper way to identify large deletions in our targeted loci. Normal Tide/ice analyses only...
02 November 2023 1,679 1 View
I am trying to use BODIPY on cultured B cells and then analyze using flow cytometry. However, I will have close to 80 samples in a 96 well plate and am afraid that the fluorescence shift will be...
31 October 2023 9,572 1 View
Pull down assay showed a discrepancy of band size between endogenous and over-expressed stable clones. Left: Endogenous Right: OE
31 October 2023 624 6 View
Hi, I’m doing transwell migration with B16F10 cells recently. But I ran into a problem after staining with 1% crystal violet, some dirty print showed in background (shown as figures). What could...
27 October 2023 8,650 1 View
How much do neutrophils, microglia, and other immune cell activation and infiltration play a role after craniocerebral trauma?
26 October 2023 6,581 3 View
I am preforming WB on mice cortical samples and receive double band on the blot- no matter the method of blocking. Im using a monoclonal primary antibody. Has anyone encountered? or Knows if it is...
26 October 2023 4,386 0 View
Hello, I am trying to construct phylogenetic tree of HIV-1. I downloaded sequences from few neighbor countries from Los Alamos HIV database. After aligning and trimming the length of sequences is...
25 October 2023 5,959 4 View
I make the BP reaction with gBLOCKS (correctly sequenced previously) and the results is a pDONR with lot of miss sequence. I confirmed by plasmid sequencing that the gBLOCK is missing a big part...
24 October 2023 582 0 View
I'm trying to generate ligand topology files through CHARMM FF but i'm getting this error. I installed the respective python version but error is still same python cgenff_charmm2gmx.py UNK...
24 October 2023 1,561 4 View
I've read that template-switching can create duplications, but those duplications are inverted (aka TIDs - tandem inverted duplications). Can template-switching result in a non-inverted...
23 October 2023 9,380 7 View
Whenever I do a western blot, when the comb comes out of the stacking gel, there are little bits of gel stuck in the wells and some bubbles. What is the best way to get rid of the debris and bubbles?
20 October 2023 2,384 2 View