I am to design sgRNA for Cas9 fluorescent tagging, but all of the tools I find online are for standard CRISPR knocking in/out. Anyone have a recommended tool for dCas9?
The source of classification bias in marker gene metagenome sequencing? a variability in the taxonomy classification of microbial communities when using different primer pairs (e.g. for 16S rDNA)...
29 June 2024 1,048 1 View
How to store SDS-PAGE for fluorescent imaging without fixatives? I am running protein samples on SDS-PAGE that are tagged with fluorophores. If I place the gel in a destain solution of...
29 June 2024 8,720 1 View
Is a 5` phosphate needed for homologous recombination? I am a bit confused whether phosphorylated DNA is needed to achieve efficient homologous recombination in e coli. I believe there are methods...
06 June 2024 7,065 1 View
What is the reason or importance of diluting an enzyme before an experiment? I'm writing a report on enzyme activity and characterisation and i need help with understanding this: we were told to...
06 June 2024 8,714 2 View
So, here is what I am dealing with: The Tm of the primers is 55 for the forward one and 56 for the reverse one. In -silico PCR produces 750 BP band, but in reality I get 3 bands. The third one...
17 March 2024 9,914 5 View
I am amplifying target sequence 450 bp. I get single sharp band in the control and faint in sample with another sharp nonspecific product. why? I need to get one single band from my sample to...
17 March 2024 804 3 View
I am doing a CRISPR targeting to knock out one intron of my gene of interest in a cancer cell line and replace that intron with GFP. I am wondering if I should pick single cell clone with possible...
17 March 2024 9,713 1 View
For example, If a PCR primer can detect ~100 fg of DNA, how we will calculate it to Bacterial copy number?
17 March 2024 1,816 2 View
How can the design and synthesis of guide RNA molecules be optimized for improved CRISPR-Cas9 gene editing?
05 August 2023 1,375 2 View
I could not find a study that used CRISPR-mediated HDR for stable integration of large genes. I would be grateful if anyone could point me in the right direction.
05 August 2023 486 2 View
I have been running a MAGeCK test command on the terminal for a CRISPR screen to rank sgRNAs and genes based on the read count tables. However, I get only the plot of the top-ranked genes...
28 July 2024 9,811 1 View
Why knockouts for same genes from two different gRNA are showing different result? I followed CRISPR-CAS9 gene editing technology.
25 July 2024 8,313 1 View
Hello everyone! Someone working with Crispr Cas9. I have a question. Is it possible to edit a gene that is present on a plasmid (in this case with a low copy number)? In this case, is it possible...
22 July 2024 7,073 2 View
It was assumed that 4E8 T cells should be isolated and activated using plate-bound anti-human CD3, after which would be used for a genome-wide CRISPR screen. (Ref: Genome-wide CRISPR Screens in...
17 July 2024 3,370 0 View
I have attempted various methods, including homozygous gene disruption and CRISPR-Cas9, but have not achieved the desired results. It appears that the gene of interest is situated in a...
15 July 2024 3,977 1 View
Studying a specific monogenetic disease based on mutations in different individual genes. I try to recreate the disease phenotypes of donors in vitro. I knocked out a gene x with above 90%...
12 July 2024 6,313 0 View
I am looking for a CRISPR protocol for application in leishmania strains
08 July 2024 9,732 1 View
I was wondering if it is possible to form a permanent open "ssDNA bubble" similar to a transcription bubble (>13 nucleotides) within E. coli. These criteria are important: 1. Open ssDNA...
07 July 2024 8,275 5 View
I am currently investigating the CRISPR 9 case to target my gene of interest for knockout. The vector being used is lentiCRISPR v2puro. I digested and dephosphorylated 5ug of the lentiviral...
03 June 2024 2,877 7 View
Now, I am using CRISPR/Cas 9 system to make a point mutation at THBD gene. I am wondering how can I isolate the desired cell population after making a point mutation by using CRISPR/Cas 9? Are...
30 May 2024 9,993 0 View