Intron Splicing - Does it depend on 5' UTR sequences? (Jan/10/2006 )
Hello!
Does anyone know if the intron slicing is dependent on 5' UTR sequences? I'm trying to clone a genomic sequence (including introns) in a vector with an inducible promoter. My insert starts at the initial ATG and does not contain any 5' UTR region. Will this interfere with intron splicing or is the plicing independent of the UTR region and pormoter region?
Thanks in advance for your help,
Raquel
where did you take your insert from and where do you wanna send it?
are they the same or not?
firstly, are you cloning a eukaryotic gene into a prokaryotic system? if so prokaryotic models do not slice introns.
also why not reverse transcribe the mRNA and clone that instead as it will make transcription of the protien easier and sidestep intron slicing
also why not reverse transcribe the mRNA and clone that instead as it will make transcription of the protien easier and sidestep intron slicing
Prokaroytes and Eukaryotes are two different taxonomy(class or phylum)
. They have different characteristics. In Eukaryotes, the gene structures are predicted by the exons and introns, the size of the genomic sequences are larger compared to Prokaryotes. Prokaryotes have a very small genomic sequences, and they are not predicted by introns. firstly, are you cloning a eukaryotic gene into a prokaryotic system? if so prokaryotic models do not slice introns.
also why not reverse transcribe the mRNA and clone that instead as it will make transcription of the protien easier and sidestep intron slicing
Prokaroytes and Eukaryotes are two different taxonomy(class or phylum)
. They have different characteristics. In Eukaryotes, the gene structures are predicted by the exons and introns, the size of the genomic sequences are larger compared to Prokaryotes. Prokaryotes have a very small genomic sequences, and they are not predicted by introns. gene structures? I think you might be a little confused
actually prokaryotes and eukaryotes fall udner two completely different domain. In eukaryotes the RNA is spliced to remove the introns and join the exons to form mRNA from which it is translated into a functional protein. also there are no introns in almost all prokaryotes.
the size of the gene is irrelevant for cloning
what you can do however if you want to clone a gene (and which most people do) is clone a gene from a eukaryote into a prokaryotic EXPRESSION system, and that is where my question comes in, if you are cloning a eukaryotic gene from genomic DNA you will clone all the introns as well, if you insert this into a prokaryotic expression system it will not recognise the cleavage sites and thus you will not get a functional protein
should you however reverse transcribe the mRNA which you will have a DNA sequence that lacks the introns and can be inserted into a prokaryotic expression system and expressed