Mendelian_Genetics

Question 1

complementation test

Answer 1

used to distinguish whether mutations are allelic variants of a single gene, or of different genes.

Question 2

what types of genes must you use for complementation tests? why?

Answer 2

recessive, since in practice you will need to overcome (compensate for) the mutant allele with the wild type

Question 3

what happens during a complementation test if both mutants are in the same gene?

Answer 3

they are both recessive, so no complementation, and the organism has mutant phenotype

Question 4

what happens during a complementation test if mutants are in different genes?

Answer 4

Gene complement each other (heterozygotes in trans), and you get wild type phenotype

Question 5

how do you tell how many complementation groups are in your set of mutants?

Answer 5

make series of punnett squares, mark a + for complement, and - for noncomplement, then advance out until you’ve assigned groups

Question 6

what are two broad ways you can become an exception to a complementation test?

Answer 6

intragenic complementation, and non-allelic non-complementation (second site)

Question 7

what are three ways you can mechanistically achieve intragenic complementation?

Answer 7

different domains in a multi-domain protein could interact, different alleles could have reduced dosage affects, and stabilization of products created by multiple complementary changes in the protein structure (suppression)

Question 8

what is non-allelic non-complementation?

Answer 8

also called haploinsufficiency, happens if one wild-type copy of a gene is not sufficient to rescue a system to WT phenotype

Question 9

what are two mechanistic ways you can get haploinsuffiency?

Answer 9

if your protein has a dosage threshold and needs to make a certain amount to function at all (vesicles fusing at synaptic cleft) or mutant alleles poison the system

Question 10

give me some yeast life cycle/mating facts! (5)

Answer 10

1. can be haploid or diploid
2. haploid state = 16N, and are called mating type alpha
3. Haploids can make other haploids or mater with another alpha to produce a diploid
4. Diploid can stay diploid or undergo meiosis
5. Both haploids and diploids do cell cycle in 90 min

Question 11

what is reverse genetics?

Answer 11

start with a gene mutation, examine resulting phenotype

Question 12

how is a wild type gene denoted?

Answer 12

italicized, all upper case

Question 13

how is a mutant gene denoted?

Answer 13

italicized, all lower case

Question 14

how is a protein denoted?

Answer 14

no italics, first letter capitalized and the rest lower case

Question 15

how is a mutant protein denoted?

Answer 15

no italics, all lower case

Question 16

what are two types of selectable markers?

Answer 16

auxotrophic, antibiotic

Question 17

how does an auxotrophic marker work?

Answer 17

it provides an essential gene that a particular organism cannot live without. commonly used are biosynthetic pathways (Ura3 or HIS3). so these experiments have to be carried out in organisms with these essential genes deleted.

Question 18

how are selectable markers incorporated?

Answer 18

you position them right beside the gene of interest’s ORF (close enough to link), target mutant allele to the locus using 50-500bp of endogenous locus (homology arms, everything introduced to cell by transformation), then homology arms find the locus and take advantage of recombination machinery

Question 19

how often does recombination happen in mammalian vs yeast cells?

Answer 19

1/20,000 in mammalian, 1/100 in yeast

Question 20

advantages to working with yeast?

Answer 20

haploid or dipoid, quick cell cycle, highly recombinagenic, can see cell cycle under light microscope

Question 21

what is sporulation?

Answer 21

production of haploid spores post meiosis of diploid yeast

Question 22

amorph

Answer 22

no phenotype

Question 23

T/F. Null allele behaves the same way as a deletion

Answer 23

True

Question 24

hypomorph (loss of function)

Answer 24

recessive allele with less function than a WT, but more gene function than a null

Question 25

Classic way to distinguish allelic classes (phenotypic analysis)

Answer 25

mutant/deletion, and mutant/2X WT overexpression, then see if phenotype gets “better” or “worse”

Question 26

what is a neomorph?

Answer 26

gain of function allele - protein misexpression or novel ligand binding

Question 27

what if a mutation is lethal?

Answer 27

can use conditional alleles

Question 28

what are some triggers for a conditional mutation?

Answer 28

heat, cold, mechanical stimulation, addition of a specific molecule

Question 29

reverse genetics examples?

Answer 29

delete a specific gene to look at its null phenotype, make a particular mutation to see how it affects protein function

Question 30

common tools for reverse genetics?

Answer 30

crispr, RNAi, TALENs, selectable markers

Question 31

forward genetics examples?

Answer 31

ID mutants with interesting phenotype, random mutations

Question 32

common tools for forward genetics?

Answer 32

Drosophila P-elements, chemical mutagens, Tn-seq

Question 33

why does percent cell survival tank with increasing concentration of a mutagen?

Answer 33

likelihood of hitting an essential gene increases

Question 34

caveats to mutagenesis

Answer 34

can have multiple mutations in one cell, you must back-cross to wildtype, you will miss some genes (essential, short, redundant)

Question 35

T/F. All phenotypes are selectable

Answer 35

False, this is a drawback to selection experiments

Question 36

T/F. Screens are more biased than selection

Answer 36

False