Prokaryotic Genetics in the Lab

Question 1

spontaneous mutations

Answer 1

-relies on natural mutations that arise by random processes
-need large number of bacteria nad powerful methods to isolate mutants of interest

Question 2

gene names and protein names

Answer 2

gene names - first 3 letters decribe function and 4th letter designates specific gene
-gene names are italicized with first 3 letters lower case and last letter upper case
-protein names are the same but start with an upper case letter and are not italizced

Question 3

induced mutations

Answer 3

expose your organism to agents that increase mutation rate
-UV, chemicals

Question 4

transposon insertion mitation

Answer 4

-introduce transposon that inserts randomly into the genome of your organism by transformation or conjugation
-generally disrupts whatefer gene it inserts into
-transposon carries AMR gene to isolate bacteria with Tn insertion

Question 5

selection

Answer 5

-ways to isoalte mutant of interest
-mutant grows but parent doesnt
-selection is highly efficient and you can identify single mutant with a desired phenotype out of millions of cells

Question 6

screening for absence of growth

Answer 6

-easier to identify mutants that grow better than parent by selection
-replica plating (plate same colony on two different plates under two different conditions) you can identify mutants that grow worse than parent

Question 7

auxotroph mutants

Answer 7

-mutants that reuqire specific nutrients to grow

Question 8

transposon insertion site sequencing

Answer 8

-make large library of transposon mutants - lots of different bacteria each with one random Tn insertion
-libaray will be a big pool of bacteria each bacterium has one Tn insertion in a random spot in genome. In library thousands of different Tn sites, many copies and many bacteria for each Tn site
-sequence the Tn insertion sites - the DNA immediately beside when the transposon landed, tells you frequency of each Tn mutant in your librayr, Millions of DNA sequence reads (input population)
-expose mutant library to some challenge, (output population)