Must a recombinent gene be cloned at the site of lacZ for it to be deactivated?

Answer 1

Your question doesn't make a lot of sense (how about adding more details) but let's see.

Many plasmid vectors have a lacZ gene under an inducible promoter and a multiple cloning site in the begining of the lacZ gene. When a recombinant gene is cloned there, the lacZ gene is disrupted and thus you can screen for the colonies that have an insert by looking at the lack of beta-galactosidase activity with a proper assay (like growing on plates with X-gal; colonies with the insert will be white while colonies with the intact plasmid will be blue).

However not all plasmid vectors have this system and you also need an appropriate host strain (with deleted chromosomal lacZ).

If you formulate your question better I could give you a better answer

Answer 2

If you are wondering whether the color selection process for screening colonies for cloned inserts requires cloning into the sequence of the lacZ gene, then generally speaking you are correct. The most commonly used method for determining if a cDNA or other DNA fragment has been cloned into a specific vector is to engineer the vector to have the specific lacZ complementing gene fragment that is missing in the host bacterial genome, and to couple it to a promoter that is sensitive to the presence of gratuitous inducers such as IPTG.

When a sequence of interest has been cloned into the complementing lacZ fragment then the bug can not produce active beta-galactosidase and when grown in the presence of X-gal or any of a number of other chromogenic substrates there will be no color produced by the recombinant colonies.