Question

Both the lacIS and lacP- mutations result in the inability to induce expression of lacZ of lacY. You have recently isolated a mutant E,col with thie phenotype, and want to determine whether it is a new lacIS or lacP- mutant. Using whatever haploid or dipliod genotypes you want, describe a set of genotypes that would let you determine this. Explain reasoning.

Answer 1

This question deals with the regulation of gene expression in bacteria. This is generally called as a lac operon where a polycistronic RNA is produced which codes for more than one protein. These sequence of genes codes for proteins which are involved in the metabolism of lacotse as an alternate energy source. The structural genes of this lac operon are lacZ, lacY and lacA. in lacP^- mutation the RNA polymerase fails to bind to the promoter which leads to decreased expression of structural genes and are poorly inducible with IPTG. Thus, pseudo diploid genetics method would involve transforming a plasmid having wild type promoter along with the structural genes. Then the cells can be analyzed for mRNA or B-galactosidase activity post induction to ensure that the mutatiin was in the promoter.

Similary lacI^s stands for superrepressor mutation where there is a mutated lactose repressor which does not dislodge from the operator even when lactose is present. Since the represor is a trans acting factor there would be no induction of lac operon genes even in the presence of a wild type lacI+ because the mutant repressor can still bind to the plasmid's wild type operator.Thus, a plasmid containing lacI⁺ along with the wild type structural genes would fail to induce the gene expression in the presence of lactose or an inducer.

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