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Alternative start codon in plasmid

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#1 tapi



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Posted 28 October 2013 - 07:24 AM

Hi all,


just a question of concept:


We made a mistake in the designing of a construct but we realized once the experiment was finished.


We wanted to overexpress a gene. We cloned the desired gene substituting a LacZ gene but we accidentally left a start codon (with kozak sequence) upstream of the ATG of our gene (which also have its own Kozac sequence). The ATG left by mistake was from a hp68/70 promoter-LacZgene


This not desired ATG codon was in frame with the other and there was a stop codon between both of them.


Surprisingly everything worked perfectly and the protein seems to give the effect desired.


I was wondering what has happened here. Possible explanations could be:


1- Translation started in alternative ATG and ended and later started again with the correct ATG?: I though that the mRNA deattach from ribosome after stop sequence is found


2- Translation stated in the alternative ATG but continued because the stop codon was not enough to stop translation, resulting in a protein with some more aminoacids at the beginning but perfectly functional?: Maybe there is any other more sequence required to stop translation apart from the stop codon?


I am extremely puzzled by this and I would really appreciate some comments


Many thanks!!!

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