Probably, although the efficiency would be lower because of self-ligation and then your parts that are attached to the beads could ligate to each other. Definitely a high risk of side products. Though it'd be nice if you could just take raw PCR product stick it in.
On Wednesday, February 19, 2014 4:48:03 PM UTC-5, Nathan McCorkle wrote:
-- I can't remember what the concentration was for the DNA parts that they supply, maybe that's what gives such a high success rate? High purity from HPLC definitely helps, but I wonder if you could do it with just digested PCR product, maybe purified by a regular kit. That'd be important for using it in a research context.
You could easily assemble expression vectors for your screening conditions when it comes to expressing or co-expressing recombinant protein. Right now, the vectors that my lab uses are completely static. It could have taken me a couple days to assemble all the plasmids that I've made so far vs. the weeks of work that I had put in.
Cheers,
Kevin
On Wednesday, February 19, 2014 4:48:03 PM UTC-5, Nathan McCorkle wrote:
Couldn't you just buy some primers of the same overhang and blunt ligate?
On Wed, Feb 19, 2014 at 1:30 PM, Cory Tobin <cory....@gmail.com> wrote:
>> Dan, what do you mean by 'stuck with what is in the kit'?
>
> The parts aren't easily manufactured. The overhangs are purified via
> HPLC, so unless you have access to that you won't be able to make your
> own parts. Doug (from Genomikon) is on this list I think, so maybe he
> can provide insight about making Genomikon parts.
>
> That being said, it's a great kit especially for educational purposes.
> I've taught a couple classes using the kit and I think we've had 100%
> success rate in terms of the students getting at least one colony
> containing the correct plasmid. It's really foolproof and can be done
> in a couple of hours.
>
> I doubt it will be used in many research settings. If you're making
> one-off DNA constructs you're better off using traditional cloning
> methods. But I could see it being used in a situation where you have
> a small number of parts that you recombine in lots of different
> combinations, in which case it would save you a boat load of time.
>
> -cory
>
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