The E. coli MalT protein is a positive regulator ofseveral mal operons, which are induced in the presenceof the sugar maltose. The gene that encodes MalT wasidentified in a screen for mutants causing constitutiveexpression of mal operons; the operons were transcribedeven in the absence of maltose. The screen involved alacZ transcriptional fusion reporter gene in which theregulatory region of a maltose-inducible operon wasfused to the coding sequences of lacZ.a. Bacteria with a lacZ− mutation are transformedwith the reporter gene and spread on petri platescontaining the β-galactosidase substrate X-gal.What color would the colonies be if the plates alsocontained maltose? What if the plates had X-galbut no maltose?b. In the screen, scientists mutagenized the lacZ− bacteria before transforming them with the reportergene, and then spread the transformed bacteria onplates with X-gal and no maltose. All of the colonies were white except for one colony that wasblue. At this stage of the analysis, researcherscould not establish whether the gene mutant in theblue colony encoded a positive or a negative regulator of mal operons.Suppose first that the gene encoded a positiveregulator. (i) How could the wild-type protein respond to maltose? (ii) How would the mutation affect protein function? (iii) Describe the likely natureof the mutation in the gene at the molecular level.Now answer these same three questions for thehypothesis in which the gene encoded a negativeregulator (a repressor) of mal operon expression.c. How do you think the scientists figured out that MalTwas a positive regulator and not a repressor? (Hint:Recall Fig. 14.28. Think about what would happenin each case if the researchers attempted to identifythe malT mutant using a plasmid library made fromthe genome of a wild-type strain versus a plasmidlibrary made from the genome of the mutant strain.)
Bacterial Genomics
The study of the morphological, physiological, and evolutionary aspects of the bacterial genome is referred to as bacterial genomics. This subdisciplinary field aids in understanding how genes are assembled into genomes. Further, bacterial or microbial genomics has helped researchers in understanding the pathogenicity of bacteria and other microbes.
Transformation Experiment in Bacteria
In the discovery of genetic material, the experiment conducted by Frederick Griffith on Streptococcus pneumonia proved to be a stepping stone.
Plasmids and Vectors
The DNA molecule that exists in a circular shape and is smaller in size which is capable of its replication is called Plasmids. In other words, it is called extra-chromosomal plasmid DNA. Vectors are the molecule which is capable of carrying genetic material which can be transferred into another cell and further carry out replication and expression. Plasmids can act as vectors.
The E. coli MalT protein is a positive regulator of
several mal operons, which are induced in the presence
of the sugar maltose. The gene that encodes MalT was
identified in a screen for mutants causing constitutive
expression of mal operons; the operons were transcribed
even in the absence of maltose. The screen involved a
lacZ transcriptional fusion reporter gene in which the
regulatory region of a maltose-inducible operon was
fused to the coding sequences of lacZ.
a. Bacteria with a lacZ− mutation are transformed
with the reporter gene and spread on petri plates
containing the β-galactosidase substrate X-gal.
What color would the colonies be if the plates also
contained maltose? What if the plates had X-gal
but no maltose?
b. In the screen, scientists mutagenized the lacZ− bacteria before transforming them with the reporter
gene, and then spread the transformed bacteria on
plates with X-gal and no maltose. All of the colonies were white except for one colony that was
blue. At this stage of the analysis, researchers
could not establish whether the gene mutant in the
blue colony encoded a positive or a negative regulator of mal operons.
Suppose first that the gene encoded a positive
regulator. (i) How could the wild-type protein respond to maltose? (ii) How would the mutation affect protein function? (iii) Describe the likely nature
of the mutation in the gene at the molecular level.
Now answer these same three questions for the
hypothesis in which the gene encoded a negative
regulator (a repressor) of mal operon expression.
c. How do you think the scientists figured out that MalT
was a positive regulator and not a repressor? (Hint:
Recall Fig. 14.28. Think about what would happen
in each case if the researchers attempted to identify
the malT mutant using a plasmid library made from
the genome of a wild-type strain versus a plasmid
library made from the genome of the mutant strain.)
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