RF-Cloning.org Forum

Anything and everything cloning: Go...

You are not logged in.

#1 2016-01-27 13:08:54

Registered: 2016-01-26
Posts: 1

trouble with 9 kbp plasmid

Hi all,

I've been doing RF cloning for quite a while. I always worked with vectors of an final length of 6 kbp without any major issues. With 9 kbp I run into troubles. I end up without colonies. I'm pretty much sticking to the literature in my protocol. In the primer design, I must stick to a certain spot. So, the overhangs of the megaprimer a directly adjacent on the parental vector. I cannot leave out any bp in between. Could this also be an issue?
Any comment appreciated. Thx



#2 2016-01-27 14:18:25

Steve Bond
Registered: 2014-01-23
Posts: 129

Re: trouble with 9 kbp plasmid

Hi Chris,
The fact that your 6 Kb constructs are working is great news, because it means you are doing things right from a technical sense. So there is probably something going wonky on the design side of the 9 Kb construct. Did you use the website to design it? It's much easier for me to help troubleshoot if I can look at it directly, instead of trying to work from your description. I can search the backend for past projects if you did design it on the site, I just need something to search for.


#3 2021-05-31 12:23:32

Registered: 2021-05-31
Posts: 1

Re: trouble with 9 kbp plasmid

thanks for the awesome information.
https://ometv.onl https://chatroulette.top https://omegle.wtf https://bazoocam.cam

Last edited by alexsunny123 (2021-08-14 14:16:33)


Board footer

Powered by FluxBB