Terminologies (1) and concept
Minimal medium, complete medium
Auxotroph vs. prototroph
Silent, nonsense, missense, neutral mutations
Recessive and dominant mutations
Conditional mutants
Hypomorph (reduction of function), hypermorph (gain-of-function), amorph
(null), antimorph (dominant-negative), neomorph (gaining new function)
Semi-dominant (incomplete dominance, co-dominance), haploidinsufficient,
Reporter (Luciferase)
Ectopic (in time and in space)
Mutant screen vs. mutant selection
Homozygous vs heterozygous
Alleles
Complementation groups
Complementation test
Intragenic, extragenic suppressors (one example of each)
Cloning
Basic tools in cloning (ligase, vector, lacZ, restriction enzymes)
Library construction and screen (will be taught this Thu)
Colony hybridization vs. Southern hybridization
Southern vs Northern
Restriction enzyme vs. Restriction map
Genotype vs. phenotype
Reverse transcriptase vs. transposases vs. Taq polymerase
Denature DNA, annealing, hybridization
Primers
Probes
Competent cells
Transformation
(2) Epistatic, synergistic, additive, suppressive relationship between mutations in
different genes. Learn how to determine epistatic relations between two mutants based on
single and double mutant phenotypes (in biochemical pathway vs regulatory pathway).
(We will practice some questions on epistasis in Thu class).
(3) Technologies and their applications
Northern, Southern, Restriction mapping, library construction, cloning, colony
hybridization, gel electrophoresis
(4) What does a vector need to possess? Different vectors based on insert size
(5) What is the purpose of each step of PCR? Each step of Southern blot?
(6) How to construct a restriction map
(7) Central dogma
(8) Study Homework #1
NO PCR on exam
Unlike humans plants grow outdoors and are under constant sunlight. Why dont plants
get skin cancers? To answer question this Dr. Liu has conducted an EMS mutagenesis
screen and has identified several mutations in genes that control Arabidopsis plants
sensitivity to UV light. Two mutations caused increased sensitivity to UV irradiation,
while two other mutations caused increased resistance to UV irradiation.
a. What is the wild type function of (UV1, 2)same, (3, 4?same) ( hint: simply state if each
wild type gene promotes or inhibits UV resistance) assuming loss of function mutations?
3 and 4 inhibits mutation to UV and may encode regulatory genes, mutate the
repressors can cause plants show more and cause more sensitivity to UV
b. find if UV1 recessive or dominant? Cross with wildtype and if similar to wildtype will
be dominance
c.)how do you find it Uv 1 and uv 2 mutations are actually allelic to each other
cross 1 and 2 f1 all have mutant phenotype more sensitive to uv 1 so uv1 and 2 are
allelic
d) answerUV1 mutant is more sensitive
UV3 and UV4 mutants are more resistant
The double mutant UV1 UV3 more sensitive to UV irradiation
UV1 epistatic to UV3, in pathway UV 1 acts later regulatory pathway downstream gene
occurs later
UV4 epistatic to UV1 meaning that UV4 is downstream of UV 1
ORDER:
UV3
UV1
UV4
In biochemical pathway the earlier acting gene is epistatic to the later acting gene
(opposite)
What is the regulatory relationship among these three genes? Use Arrows and bar to
indicate their positive or negative regulatory activities? Start with UV irradiation and end
with DNA repair
UV 3 and UV4- repressing, more resistant
UV4 inhibiting DNA repair
UV irradiation -----|
UV3-----|
UV1-----|
UV4-----|
DNA repair
If you mutate UV3 the plant is more resistant and nothing is there to stop UV1 and
always active and always allow for DNA repair to occur
UV irradiation cause DNA repair to occur and removes the repressors of DNA UV3 and
UV4