How to validate sgRNA for cripsra?

14 October 2023 0 7K Report

hi i'm gonna use crispra system for my study, how can i validate sgRNA not having off-target effect?

Badges
Science topic

Similar topics
Microbiology
Assays

More Yr Hwang's questions See All

Lipopolysaccharide (LPS) extraction problem?

Hello, all I am currently extracting LPS from E. coli MG1655 and its mutant strains. I followed several different protocols (phenol methods or phenol+diethy ether methods), but I couldn't see...

25 June 2024 7,568 1 View

Different results from western blot and icc?

hi have you ever experienced of coming different results from western blot and icc? I treated toxin (a-syn PFF) to control ES-derived midbrain doparminergic neuron and experimental group that I...

25 May 2024 3,162 0 View

In Materials Studio, I can't select an atom or atoms by clicking and dragging. What should I do?

In Materials Studio, I can't select an atom or atoms by clicking and dragging. I can select only bonds in 3D model by clicking or dragging. What's the problem?

30 January 2024 1,259 1 View

Can you explain me about the principle of Spatial transcriptomics?

I'm trying to study spatial transcriptomics by myself, but it's not easy to understand. So, spatial GEX is to cut the tissue into slides, attach it to the slide where the probes are located, and...

24 December 2023 6,069 1 View

Which one is best protocl for Thp-1 differentiation?

I found two papers to establish a Thp-1 differentiation protocol (Thp-1 -(PMA)-> M0 -(LPS+IFN or IL-4+IL-13)->M1 or M2). I want to compare both protocols, but the PMA priming times and the types...

12 September 2023 1,149 0 View

In DFT, Which is the most accurate method to calculate band alignment?

As I know, there are 3 reference energy to find or compare band level when we need to band alignment data 1. fermi energy set to zero 2. Vacuum energy (from slab calculation) set to zero 3. Align...

03 August 2023 2,187 0 View

Which of the direction in measuring Uv-vis spectrometer is correct?

When measuring the reflectance or transmittance of a sample, light can be incident on a) the sample surface / or b) the substrate surface. While a) is correct from the point of view of measuring...

29 June 2023 4,282 0 View

Is there anyone to have a trouble in cell culture using cells purchased from Celprogen?

For my reseach, I had purchased several frozen vials of HFSC(Hair follicle stem cell) from Celprogen, but I could not characteriz the Celprogen-HFSC with several anti-human antibodies for...

19 April 2023 3,801 2 View

Before preforming principal component analysis, can I transform variables into first-differenced form to make them stationary?

Before preforming principal component analysis to construct multidimensional index, can I transform variables into first-differenced form to make them stationary if some of these variables present...

30 March 2023 1,313 2 View

How to get work function of semiconductor by DFT?

As I know, by DFT, work fucntion is calculated from Ev(Vacuum level) - Ef(fermi energy) = WF And Vacuum level can be calculated from vacuum region of slab model fermi energy can get from OUTCAR or...

19 March 2023 6,627 0 View

Where to find a gene list for CRISPRa/i library screening of regulatory factors that affect pathogenic Th17 differentiation in PBMC?

I want to perform a CRISPRa/i library screening for candidate regulatory factors that affect pathogenic Th17 differentiation in PBMC. But I am wondering how to define the size of gene library, so...

31 July 2024 2,150 0 View

SgRNA Oligos Annealing / Ligation issues with LentiCRISPRv2?

I am trying to clone my target sequence for CRISPR/Cas9 work. I have performed digestion and gel purificatoon of the lentiCRISPRv2 plasmid with a good yield and purity. However, when I performed...

26 March 2024 6,376 2 View

How can I improve the efficiency of single gRNA to hybridize with Cas9 to get the RNP complex?

We use synthetic gRNA from IDT but we think have some problems for sgRNA hybridization with Cas9. any suggestion is welcomed!

25 March 2024 1,536 3 View

Accidentally included extra 5' GG on gRNA sequence?

I'm doing CRISPRi with dead spCas9 on mouse cells, and normally we do the following for gRNA design: 1. 20 nt spacer 2. Extra G on the 5' to enhance expression from hU6 promoter. Final product:...

25 February 2024 4,498 3 View

Advice for TIDE Primers?

Hi Everyone! I am currently designing primers to validate my CRISPR-KO mammalian cell line using TIDE. The primer is designed about 250bp upstream of the PAM site. Then, Sanger sequencing is...

27 January 2024 2,219 1 View

Does anyone know if there is sgRNA library designed for MDCK cell line?

I am interested in knowing if there is any sgRNA library designed against MDCK cell line which is a canine cell line. i believe it shares about 84% homology with human cell line. let me know if...

03 January 2024 5,148 0 View

Do DNA fragments expressing sgRNA have to be integrated into the host cell genome when doing a crispr screen?

Lentiviruses are often used as delivery vectors for in vivo crispr screening because the lentiviral genome is able to integrate into the cellular genome, so is it okay to use AAV, a virus that...

11 December 2023 7,742 0 View

Has anyone got experience with low knockout efficiency using Crispr?

I work in a project where we need to knockout a gene at primary T cells targeting an exon that is shared between all variants. However, while the sgRNA design has been optimised ,we still see low...

24 October 2023 5,722 1 View

What transfection reagent should I use transfecting HEK293T cells with IVT transcribed sgRNA and plasmid carrying dCas9 backbone?

I am trying to transfect HEK293 cells with dCas9-KRAB-DNMT3A carrying plasmid and multiple IVT sgRNAs. RNAi max invitrogen and Hiperfect Qiagen are only good for small RNAs but not plasmids. Does...

04 October 2023 3,360 1 View

What's a good online sgRNA designer for dCas9?

Hello! I am trying to design sgRNAs for dCas9 fluorescent tagging, but all of the tools I find online are for standard CRISPR knocking in/out. Anyone have a recommended tool for dCas9?

27 August 2023 1,966 0 View