Can anyone explain me or join to me a reference about abclonal seamless cloning?

More Abire Sigbessia's questions See All

Why fluorescent tag lost after cell harvest?

I used pet16b vector to clone my gene of interest to further purify the protein .I added the fluorescent tag at the c-terminal for further microscopy. After cloning ,samples were sent to sequence...

15 July 2020 3,919 2 View

Protein doesn't elute?

I used the pet16b containing an histagg to purify my protein of interest. Ni-NTA beads were used to purify the protein. after washing, the protein was still present when runned on the sds-gel...

19 May 2020 8,903 4 View

Is there any particular site to insert the fragment in histag vector for further protein purification?

Do we need to place the insert in a specific site in his tagg vector for further protein purification or can we just use any restriction site present in the vector?

18 September 2019 6,816 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

Does anyone have a recommendation for yeast reporter gene plasmid/protocol?

Hi, could anyone recommend a plasmid and/or protocol for reporter gene assay in S. cerevisiae? I want to assess the effect of growth conditions on a transcription factor, so I want to clone it`s...

01 March 2021 210 1 View

Expression cloning by using cDNA,do I need to modify the cDNA first?

I am going to have a expression cloning of mammalian gene by using shuttle plasmid to transforming the E.coli However I don't know I should only inserting the Coding sequence ,or I can...

28 February 2021 5,440 3 View

Difficulty in cloning of 1kb gene into vector?

I am facing difficulties in cloning a 1kb gene into a vector (pJIT163). I have my gene on interest (GOI) in pUC57 and want to clone in pJIT163 using SalI and BamHI restriction sites. I am getting...

25 February 2021 3,221 7 View

Why are Sanger Sequencing reads interrupted in some edited clones?

I have a Crispr/Cas9 edited bulk population and I can briefly check the indel ratios in the bulk population using Sanger sequencing. After cloning single cells, I wanted to check the indel types...

18 February 2021 8,198 6 View

Hi, can anybody help me with the suggestions how can I amplify a stress induced gene of 1107 bp?

I am trying to amplify the TGA1 (Transcription factor 1 ) of Arabidopsis ,which is 1107 bp. I have treated the plants with Salycylic solution over night and extracted the RNA and make cDNA. I...

17 February 2021 7,802 2 View

Alternatives to traditional cloning of insert into backbone (dsDNA)?

Hi, I am stuck with a cloning experiment: The insert & backbone(bb) share one compatible end at 3' end(NotI) and one incompatible end at 5' end (XbaI for insert; HindIII for backbone). So i...

16 February 2021 1,664 3 View

Why do I have trouble with cloning duplicate gene in the same plasmid?

Hello! I am trying to generate a construct in this format: seqA - seqB - linker - seqC - seqB into pSF-CMV-Ub-Puro-Ascl plasmid 0.5kb 2kb 66 0.25kb 2kb I have tried many...

15 February 2021 7,118 3 View

Would using an asRNA for repressing transcription of a high expression Listeria monocytogenes non-coding RNA harbored in a plasmid be a viable option?

Hello all, I am interested in deleting a highly expressed non-coding RNA located on a Listeria monocytogenes plasmid. Currently, there are not any optimized gene deletion methods for Listeria...

09 February 2021 4,994 1 View

Quality of antibodies from Creative Biolabs?

It seems Creative Biolabs has everything and so many versions of antibodies not owned or sold by any other company. How reliable are their products? I am looking for a clone where potentially...

08 February 2021 2,167 3 View

Martin Chenal

Seamless cloning integrates PCR technologies into traditional, restriction-ligation cloning, making it a more versatile tool Abclonal is a brand like any other.

Here is a very helpful link about PCR cloning. This website also has many other user-friendly cloning protocols, which are perfect for neophytes: https://www.addgene.org/protocols/pcr-cloning/

To better understand the difference between traditional and seamless cloning : https://international.neb.com/applications/cloning-and-synthetic-biology/seamless-cloning.

Feel free to also check documentation about Gibson Assembly.

Hope it helped

Prof. Gheita

Dear Abire Sigbessia

Check these

https://www.cell.com/trends/biotechnology/fulltext/S0167-7799(05)00054-5

https://www.nature.com/articles/srep44648

https://international.neb.com/applications/cloning-and-synthetic-biology/seamless-cloning

Abire Sigbessia

Martin Chenal thankyou I appreciate