HELP with SNAPgene - NEBuilder HiFi?

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Can someone have an ideas on why i have 2 bands on gel?

I expressed my protein in cell free with wheat germ, and after purification on streptag with (2,5mM, elution) i observed 2 bands on gels at the same weight. I have already expressed the protein...

17 June 2024 5,742 4 View

What is the outcomes of the special program in the arts curriculum to the completers?

The effect of the special program in the arts (SPA) curriculum to the completers.

11 April 2024 7,297 1 View

Western blot problems: sample malfunction?

Hello everyone, I'm a master student and in my project I'm studying RFP-based constructs (RFP + few amino acid-long tags). I'm now performing a Western blot in SDS-PAGE of my cell-line lysates....

04 April 2024 7,382 3 View

Mplus - moderated mediation - covariances among paths?

Hi! I am trying to analyze a moderated mediation using mplus. I first started with a simple mediation, so X -> M -> Y. Path a (X -> M) is not significant, while Path b (M -> Y) is...

25 January 2024 3,738 2 View

Has anyone performed siRNA silencing on BenD.3 cells?

Hello, I am looking for technical advices. I have never worked on mouse cells before. I plan to use lipofectamine RNAiMAX as transfection reagent. Can anyone tell me where can I find protocols or...

08 January 2024 2,680 0 View

Can you fix the proto-vessels after tube formation assay for IHC and/or TEM?

Hello, I was wondering if it is possible to fix HUVEC cells for IHC and/or TEM analysis after they completed tube formation in Matrigel. I use Corning® Matrigel® Growth Factor Reduced (GFR)...

20 November 2023 5,951 0 View

Why has south sudan failed to prevent child recruitment?

After the peace agreement reached in south sudan in 2005, and the policies designed to prevent child recruitment in armed conflict, south sudan is nowadays the country in africa with the biggest...

19 November 2023 6,524 0 View

How do I purify a protein from cell lysate without affinity chromatography?

I have liters of cell lysate and I need to separate the recombinant protein from the rest. The volume is very large for the affinity chromatography column. Any other technique? I don't need high...

28 September 2023 4,208 4 View

What about the industrial use of BL21? Is it allowed? What is the best bacterial for large scale enzyme production?

I hope someone can help me with this! I am currently working with one enzyme expressed in E. coli. We want to scale up the process to an industrial production (no Academia involved). The...

26 September 2023 3,671 1 View

Any suggestions for RNA extraction protocol from buffy coat in Shield?

Hi! I need to make RNA extractions from mammalian buffy coat RNA stored in Shield (Zymo) and frozen at -80ºC, for immune gene detection. I have tried column extraction kits with very poor results...

22 September 2023 1,047 4 View

Is there a problem with my RNA pellet?

Hello, I am currently having problems with RNA extraction. I am using mouse liver (C57BL6J), and I have extracted RNA from mouse liver before. Before this experiment, my final RNA pellets were...

11 August 2024 7,082 3 View

Can I base on reverse DNA sequences to perform alignment, convert to amino acids and GenBank submission?

I have reverse sequences (AB1 format), can I base on reverse DNA sequences to perform nucleotide alignment, convert nucleotides to amino acids and deposit the sequence in GenBank database?

11 August 2024 5,138 1 View

Strugglling with m6A dot blot any suugesstion ?

I have been doing the m6A dot blot for a while with no improvement, I am extracting the RNA, and I can see the dots although the three biological replicas give a different reading on the memberan...

10 August 2024 8,539 5 View

How to confirm the site-directed mutagenesis result without performing NGS?

I'm cloning a fragment of 3200 nts into plasmid. The cloning was successful, however, 02 amino acids were mutated. Now I want to fix these 02 aa by site-directed mutagenesis technique using...

08 August 2024 4,645 2 View

RNA Extraction Using Hot Borate Method No Longer Working?

I've been performing RNA extraction on cotton petiole tissue for a few months now using the method described in the following paper, a derivative of the typical hot borate method...

08 August 2024 9,882 2 View

I can't see the ssDNA band after performing asymmetric PCR. Is there any way to do this?

After performing symmetric PCR, PCR purification was performed. Afterwards, asymmetric PCR was performed using the PCR purification product as a template, but no ssDNA band was confirmed in the...

08 August 2024 1,668 3 View

Does crude extraction using NaOH and Tris work well with Fungi?

I'm trying to find a DNA extraction method for fungi that does not require equipment and heating. Is there anyone who can suggest an alternative option? Thank you

08 August 2024 4,733 2 View

Does Anyone have expertise in in vitro transcription and RNA pull down assay?

I am currently working on LncRNA; to know the lncRNA-protein interactions I want to do RNA pull down assay, so I need to design primers with T7 promoter. I need assistance in this regard.

07 August 2024 6,622 1 View

E.coli contamination in human RNA seq data ?

Recently, we observed that 99% of the sequences in our RNA-seq data corresponded to the E. coli genome. Despite multiple DNAse treatments after RNA extraction and ribosomal depletion, we were...

06 August 2024 807 3 View

RNA later for the preservation of RNA in fecal samples at room temperature for one day (37°C)?

I am planning to collect human fecal samples for metatranscriptomic analysis using MGI. These samples are from indigenous people living in a region with high temperatures. I will have access to a...

06 August 2024 1,367 3 View

Di Xia

Need more details of the issue in description. You can follow the tutorial of hifi cloning using snapgene here https://youtu.be/6Pic2wPoX7U

Irene Bissoli

Right, sorry.

Basically, I've already solved it with snapgene support. My kit needed to use a ssDNa, but in Snapgene you need to convert to dsDNA, otherwise, it doesn't work. For your future knowledge.

Irene Bissoli I have done similar cloning few month ago. My snapgene working fine with ssDNA in NEB hifi cloning without converting into dsDNA. I am using the latest version of snapgene.