Chapter 12

Question 1

transposons

Answer 1

• can Inactivate and Tag Genes

- For example, identifying genes required for survival in acid

Question 2

finding the location of a transposon insertion

Answer 2

• determined by sequencing from the end of the transposon across the insertion junction and into the neighboring chromosome
• Using a sequencing primer specific for the end of the transposon

Question 3

how does the transposon work

Answer 3

• The transposon can “hop,” or transpose, out of the bacteriophage DNA and into the host chromosome at random sites.
• These cells become tetracycline resistant and form a colony on medium containing tetracycline; all cells without a transposon die
• The collection of colonies, each with a transposon in a site on the chromosome, is a mutant library
• mutant libraries can be tested for altered phenotypes

Question 4

Operon fusion

Answer 4

• or transcriptional fusion
• Only the reporter protein is made
• Reflects only transcriptional control of subject gene

Question 5

Gene fusion

Answer 5

• or translational fusion
• Reflects transcriptional and translational control of subject gene, and can be used for localization of the tagged protein

Question 6

Exploring Gene Regulation

Answer 6

-Regulation of a gene under different conditions can be determined by fusing the promoter of the gene of interest to a reporter gene

Question 7

reporter gene examlpes

Answer 7

• lacZ encodes beta-galactosidase, which has easy to measure enzymatic activity, good for quantitation of gene expression
• gfp encodes green fluorescent protein, which is easily seen by fluorescent microscopy, good for localization in cells