Biology Forum › Molecular Biology › Plasmid design
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- December 6, 2011 at 11:31 am #15808
labfairyParticipantHi,
I am trying to design a plasmid to be synthesised with two genes in the MCS, and appropriate RE’s. I will then subclone this into a number of other vectors pET and one other.The question I have is – do i have to include a specific sequence between the two genes as a transcription initiation of gene 2? Or can i go straight from the stop codon of gene 1 to the start codon of gene 2?
Thanks for your help!
Labfairy - December 6, 2011 at 1:52 pm #108597JackBeanParticipant
in what organism will you transcribe it? Bacteria (but probably every organism) should be able to do transcriptional fusion, so that two genes would be transcribed from one promoter, but the second gene would have lower expression. If you want the same level of expression, you should probably use two promoters.
- December 7, 2011 at 5:22 pm #108615labfairyParticipant
Thanks for your help – the plasmid will be going into bacterial system. I’m hoping that the level of expression won’t be too different.
Thanks
Labfairy - December 8, 2011 at 1:17 am #108620canalonParticipant
Or use a novagen Duet plasmid. They are designed for co-expression study and each gene has an independant initiation sequence, with two MCS and everything to make sure that it is easy to have your 2 proteins expressed at the same level.
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