Biotechniques | Basics of Making His-Tags & Nickel Affinity Chromatography

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As always, the steps of putting a gene insert into vector/plasmid AND nickel affinity chromatography depend on your specific lab and protocol. Here, we look at the general theory of engineering a poly-His tagged protein and Ni affinity chromatography and what they are used for in the lab.

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29 сен 2024

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Комментарии : 38

@giftymammen1515 3 года назад

Wow, whatta explanation. I've been searching for the literature for hours and have been pounding my brain ever since on this topic. The minute I landed onto this video, all my confusions are gone, crystal clear now. Thankyou so much ❤️😭

@robertstadler9897 2 года назад

Very good explained! Thank you and keep going.

@snobbips5107 5 лет назад

Awesome explanation sir😊👌

@bioscienceswithshahtareenswati 2 года назад

Amazing 👏 🙀

@AA-gl1dr 3 года назад

Thank you so much!

@ying2694 4 месяца назад

How about the antibiotic resistance gene?

@izzatiauni3721 4 года назад

It is really helpful!

@CatalystUniversity 4 года назад

Thank you!

@uzoechisamuel 3 года назад

Vector + Gene = Plasmid

@satinderjit4 3 года назад

4th year bio-chem major here and this was very helpful in understanding the basic idea.

@robertadu-agyei7616 3 года назад

so insightive

@AL-eh3jx 4 года назад

I've been searching for hours and finally got to watch your video. This helped me a lot in my lab! Thank you!

@CatalystUniversity 4 года назад

Glad this was helpful.

@AL-eh3jx 4 года назад

@@CatalystUniversity I have a question and would appreciate your help! If we use Nickle binding buffer containing imidazole( low concentration) before the incubation with resin, would any binding between the His-tagged protein and Ni ions happen at this stage?

@christinamrad7341 Год назад

what if a protein contained histidin in the sample and is not our protein of interest?

@vikingmakesballs7799 2 года назад

Abdala Covid vaccine brought me here

@yourfuturedocburenbeiya 5 лет назад

This is so helpful ; thank you for posting this video doc! We're using it in lab to isolate dmAANATA and I couldn't find the molecular weight but came across this video. :)

@CatalystUniversity 4 года назад

No problem! And thanks.

@qamarhennawi9137 2 года назад

Wow, this is an awesome explanation; I've been seeking for something like this for a decade!!!!, Thank you very much.

@davalcom 2 года назад

Thanks a lot man

@ignacioquijadag.7917 2 года назад

Buen video bro, lastima que entendí la mitad porque no sé ingles jajaja pero estuvo muy bueno

@karl-leopoldkontrus6544 4 года назад

important to say, that the his-tag does not undergo post-transcriptional modifikation , or did I miss something? VERY HELPFUL VIDEO!!

@khurshedakabirov8671 3 года назад

Excellent explanation. It would be great if you added little details about how to make the protein (induction, SDS, etc...).

@faizaarshad705 3 года назад

what would be the procedure, if I want to introduce his tag protein into a cyanobacteria

@pingr.8730 3 года назад

This is very useful! Thanks for uploading this video. It's exactly what I was looking for 😊💙

@carlostettey2889 2 года назад

it will be great if you can show how to design the primers for creating these vectors.

@balasubramanianb2107 3 года назад

When wash with salt solution that nickel binded to NTA is removed or not sir?

@NehaSharma-kp8op 4 года назад

Thank u so much for making this vedio with ausome explanation

@CatalystUniversity 4 года назад

You're very welcome!

@jenna9992 4 года назад

in the gel analysis, what are the weights on the molecular ladder?

@CatalystUniversity 4 года назад

To be perfectly honest, it depends on your ladder. If you know the company and the exact product, the company website will have this available. For example, if I knew I was using the 1 kb DNA Ladder made by New England Biolabs (very good company, by the way), I could type into google search "New england biolabs 1 kb ladder". Long story short, you would need to find the exact product or at least know the size of the ladder (e.g., 1 kb).