How to design primers for Gibson assembly

  Рет қаралды 88,837

Oxford iGEM 2014

Oxford iGEM 2014

Күн бұрын

Пікірлер: 31
@roomiemcgee8899
@roomiemcgee8899 4 ай бұрын
Oh my gosh. This was so helpful. Thank you.
@johnoladokun6230
@johnoladokun6230 2 жыл бұрын
What I have been looking for....thank you for the wonderful explanation
@soumyadipXplore
@soumyadipXplore 4 жыл бұрын
Thanks..this is by far the most useful help that I got for my cloning.
@mahasish
@mahasish 3 жыл бұрын
Very good explanation. Highly recommended.
@djdonald2795
@djdonald2795 6 жыл бұрын
Terrific video. I had never heard of Gibson Assembly before, but after seeing this and the NEB videos, I think I could do it.
@ranjit2289
@ranjit2289 9 жыл бұрын
Thanks, that was explained with great clarity. I have a question: After the primer design here, do you still need to check for formation of haripins/primer dimers? Or is this design good enough for a successful ligation using the Gibson Master Mix?
@alexcracker
@alexcracker 7 жыл бұрын
Thank you Corinna, it is the best explanation I have ever seen;-)
@poiuytrew09876
@poiuytrew09876 2 жыл бұрын
Thank you for the video. But I didn't understand why we design both forward and reverse primers on both sides. Isn't it enough if we have one forward primer matching the left side of sfGFP, with the overhang marching the backbone, and a reverse primer for the right side of sfGFP, with the overhang matching the other side of the backbone? Do you use 4 primers for the PCR?
@poiuytrew09876
@poiuytrew09876 2 жыл бұрын
What I was meaning was, do we really need primers for the vector? When we digest them with restriction endonuclease, it's already linear.
@nupur0003
@nupur0003 4 жыл бұрын
thanks! absolutely wonderful explanation.
@polasoliman3955
@polasoliman3955 6 жыл бұрын
Why would retreating to the 19th base pair be prone to the primer misaligning?
@andrevargasaguilar2723
@andrevargasaguilar2723 4 жыл бұрын
because yes.
@noahyi1903
@noahyi1903 6 жыл бұрын
Thank you so much for the video. It is very helpful for me!
@aBirdbyBrancusi
@aBirdbyBrancusi 5 жыл бұрын
It was super useful to START understanding. I need more explanations. Can someone please recommend me places where I can study from? :´(
@cathytang2558
@cathytang2558 5 жыл бұрын
this is 40bp overlap, according to the manual, should 15-25bp overlap be enough?
@skaplan1995
@skaplan1995 10 жыл бұрын
Great tutorial!
@kopolojoyono
@kopolojoyono 8 жыл бұрын
quite clear, quite helpful! thank you!
@strabana5806
@strabana5806 5 жыл бұрын
Thanks, this is very helpful !! :)
@ismailcan5635
@ismailcan5635 7 жыл бұрын
Hi, I am new in this therefore your help would be appreciated. So what is the next step? A PCR where you use both intact plasmid (the plasmid on the left in the video-without the insert) and the sfGFP at the same time?
@noadrow
@noadrow 4 жыл бұрын
Thank you for this great video Where's the music from?
@ArshadPadhiar
@ArshadPadhiar 6 жыл бұрын
The Music is very catchy, where i can find it.
@MrAntihumanism
@MrAntihumanism 9 жыл бұрын
HindIII produces sticky ends, how would this would work with GA?
@rhlsy3209
@rhlsy3209 2 жыл бұрын
I think you would have to switch enzymes to something that produces blunt ends
@Trypanosoma_
@Trypanosoma_ 2 жыл бұрын
HindIII is not being used at all. A site for it was incorporated into the primers but no enzyme is being introduced
@vivekdna1
@vivekdna1 9 жыл бұрын
what software do you use for drawing your plasmids?
@jakehay9406
@jakehay9406 9 жыл бұрын
Vivek Kumar pretty sure its SnapGene
@vivekdna1
@vivekdna1 9 жыл бұрын
Thanks Jake
@Dianax.
@Dianax. 6 жыл бұрын
if I assembled two genes together, when inserting the recombinant DNA into an expression vector, the products wouldn't be altered?
@jamhaslam612
@jamhaslam612 3 жыл бұрын
if you wanted the two genes to be expressed as two separate proteins you would need either an IRES site or T2A in between the two genes.
@zilog-q3u
@zilog-q3u 3 жыл бұрын
11:10 is a mistake
@SohaMokhtariGarakani
@SohaMokhtariGarakani 4 ай бұрын
++++++
Designing cloning primers for classical (restriction) cloning
21:16
Katharine Hubbard
Рет қаралды 55 М.
A Detailed Look at Gibson Assembly
4:55
SnapGene
Рет қаралды 33 М.
REAL 3D brush can draw grass Life Hack #shorts #lifehacks
00:42
MrMaximus
Рет қаралды 12 МЛН
Will A Basketball Boat Hold My Weight?
00:30
MrBeast
Рет қаралды 126 МЛН
Это было очень близко...
00:10
Аришнев
Рет қаралды 6 МЛН
Ouch.. 🤕⚽️
00:25
Celine Dept
Рет қаралды 29 МЛН
Gibson Assembly
5:55
Herbert Sauro
Рет қаралды 51 М.
How to Design Primers for PCR
4:44
Addgene
Рет қаралды 296 М.
Gibson Assembly with SnapGene
5:07
SnapGene
Рет қаралды 18 М.
Primer Design for PCR
16:15
Herbert Sauro
Рет қаралды 655 М.
Isolating Plasmid DNA
10:29
Katie Wolfson
Рет қаралды 212 М.
Biotechniques | Principles of Primer Design for Full Gene Amplification
10:30
Catalyst University
Рет қаралды 123 М.
Primer Design: Important Considerations and Tips for Good Primer Design
6:10
Applied Biological Materials - abm
Рет қаралды 27 М.
How to Design Plasmids: Benchling Tutorial
56:42
Stanford BIOME
Рет қаралды 43 М.
Why white things are white
11:53
Steve Mould
Рет қаралды 982 М.
REAL 3D brush can draw grass Life Hack #shorts #lifehacks
00:42
MrMaximus
Рет қаралды 12 МЛН