I am constructing a DNA stretch with many genes under a single promoter by overlap extension PCR. The genes are joined in such a way such that the stop codon of one genes is followed by the start codon of next gene. Kindly suggest if this method is fine.
Madam, since you are putting different genes under a single promoter, then your expressed protein is expected to be terminated at the first occurrence of the stop codon. That is your construct will translate only the first gene. To the best of my belief, you have to remove the stop codons in-between the gene. Stop codon should appear only after the last coding region. This will translate a fused protein comprising all the genes.
Revise your construct design with some expert.
SD
As answered by Dr. Data, your method produce only the first protein. However, even the first protein may have difficulties in translation due to lengthy 3' untranslated region. If you are working with mammalian expression system, there is a possibility of expressing multiple proteins using 2A sites between the coding regions without stop cordons which produce a fused protein at first and eventually cleaved to individual proteins during translation. You can find more details about this in the attached article. Good luck.
remove the stop codon in between for each gene and place it to the end while the one promoter you use at the begining will be able to synthesize a whole protien
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