Posted 29 September 2012 - 02:33 AM
im new to molecular cloning. recently i came to know about the gateway technology.. i just was to know how far it is used in gram positive bacteria.
could anyone explain to me what is entry vector, donor vector, destination vector.s????
how will i come to know the compatibilty of plasmid to my bacteria???
more over what is the procedure to gene deletion from chromosomal dna using this technique????
hope somebody can explain in simple way???
Posted 29 September 2012 - 12:27 PM
I havn't used either so I can't comment on the use in gram+ bacteria or their use for gene deletion.
Posted 30 September 2012 - 02:56 AM
me too heard a lot of its good efficiency. when it comes to R&D or PhD what is the main defect if proprietary technology is used???
Posted 30 September 2012 - 04:16 AM
Posted 30 September 2012 - 04:18 AM
So I rather used Origene shuttling system for my vectors. It's normal restriction-ligation system, but with specially selected RE on each side of ORF, that is unique for vast majority of sequences. But otherwise it's normal cloning. They have some commonly used destination vectors linearised and precut, ready to ligate, but unfortunately not the one I needed.
Problem is most of these systems are not compatible to each other, you buy ORF in one company, you can't use destination vector from other. So it's better to look at the vectors available, if they have what you need, because you can't change it.
I never trust anything that can't be doubted.