rules: cell phones off computers only for class related work no food or drink in lab room
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Biol 423L Laboratories in Genetics. Rules: Cell phones off Computers only for class related work No food or drink in lab room Text Book: Hartwell et al., 2nd Edition 2004 Genetics from Genes to Genomes Mc Graw-Hill, Boston. Web page: www.bio.unc.edu/courses/2006Fall/Biol423L. - PowerPoint PPT PresentationTRANSCRIPT
Rules: Cell phones offComputers only for class related workNo food or drink in lab room
Text Book: Hartwell et al., 2nd Edition 2004
Genetics from Genes to Genomes Mc Graw-Hill, Boston.
Web page: www.bio.unc.edu/courses/2006Fall/Biol423L
Biol 423L Laboratories in Genetics
Goals for course:
Reinforce basic genetic principles
Introduce model organisms commonly used by geneticists
Learn how genetics is used to understandDiseaseBiochemical pathwaysDevelopment
Lab reports:
AbstractIntroductionResultsDiscussion
Course information page has instructions about preparing your lab reports.
Grading:
Lab Reports: 50% of grade5% of that is participation1 day late, 50% offmore than that will only be graded under special circumstances.
Research Paper: 10% of gradeTopics due Oct. 9.Outline due Oct. 23.Paper due Nov. 29
2 quizzes: 10% each of final grade.Oct. 2 and Nov. 13.
Final exam: 20% of final grade comprehensiveDec. 11.
Genes Alleles and Epistasis
Genetics starts with observation
Use genetics to understand the cause of the variability.
What proteins or RNAs are responsible for the variability you can see?
Observe variability
Easy example, flower color
How many genes affect flower color?
How variable are the proteins encoded by those genes?
What is the pathway to make flower color?
List of terms:
Trait: some aspect of an organism that can be observed, measured
Phenotype: the way a trait appears in an individual, the combinationof genotype and environment.
Genotype: the constitution of alleles at any gene in an individual.
Gene: continuous stretch of DNA sufficient to encode a messengerRNA or a functional RNA.
Locus: A region of a chromosome, usually for a single gene.
Messenger RNA: the RNA message for a single protein.
Allele: a variant of the sequence of a given gene.
Diploid: an individual with two copies of each chromosome.Haploid: an individual with one copy of each chromosome.
How many genes affect flower color?
How variable are the proteins encoded by those genes?
What is the pathway to make flower color?
First make sure the types are heritable and true breeding
(homozygous for flower color alleles)
X
purple by purple (self) All uniform
Homozygous: a diploid individual with two copies of the sameallele for a given gene.
Heterozygous: a diploid individual with two different alleles for a given gene.
What are the relationships between color types?
Purple is dominant to Red
X
What are the relationships between color types?
Purple is dominant to white
X
Complementation tests can be madebetween recessive alleles.
A dominant allele cannot be used.
Why?
1. How many genes are required to makepurple pigment in flowers?
Allelism test:Cross different white flowered plants
If the mutations are in the same gene,The progeny will be white
White 1
X
White 1
Allelism test:Cross different white flowered plants
If the mutations are in different genes,The progeny will be pigmented
White 1
X
White 2
Allelism or complementation test
If plants with recessive alleles arecrossed and the progeny also have the recessive trait, The alleles are variants of the same gene
If plants with recessive alleles arecrossed and the progeny have the dominant trait, The alleles are variants of different genes
Precursor 1
Intermediate
White1
White2
Purple
Pathway to purple
White 1
White 2
X
Purple
rrAA RRaa RrAa
Using multiple allelism tests with diverse recessive mutants,
We can identify all the genes specificallyinvolved in making the purple pigment
Genetics can be used to determine the Order of steps in a biological pathway
Epistasis tells which gene productsact earlier or later in a process.
What are the relationships between color types?
Purple is dominant to White 1
X
F2 1 RR, 2Rr and 1rr
X
Purple is dominant to White1
PurpleRR
White1rr
F1X
PurpleRr
Punnet square
r
r rr
Female gametesMale gametes
R
R RR Rr
Rr
What are the relationships between color types?
Purple is dominant to Red
XPP
Pp
pp
Epistasis
Two genes for flower color
Two steps in a pathway to make pigment
Where are the two genes in the pathway?
Purple is either a mixture of blue and red pigments
or
Purple results from modification of the same precursor from a white precursor to a red intermediate and finally a purple pigment.
We can use genetics to distinguish the two possibilities.The effect of variant alleles in multiple genes that affect pigment in combination will answer the question.
Precursor 1 Precursor 2
Blue Red
Precursor 1
RP
Red
R
P
Purple
Pathway 1 Pathway 2
Coexpression of Blue and red pigmentderived from different precursorsMakes purple
Modification of the sameprecursor leads to first a red pigment and then a purple pigment
Relationship between White1 and Red
X
X
White1rrPP
RedRRpp
F1 is all PurpleRrPp
F2
9 43
Punnet Square: two genes with randomly segregating alleles
Male gametes
Female gametes
RP
Rp
rP
rp
rprPRpRP
RRPP
RRPp
RRPp RrPP RrPp
RRpp RrPp Rrpp
RrPP RrPp rrPP
rrppRrPp Rrpp rrPp
rrPp
9R_P_ 3R_pp 3rrP_ 1rrpp
RrPp X RrPp
9R_P_ 3R_pp 3rrP_ 1rrppPhenotypes:
Purple WhiteRed White
Precursor 1
Red
R
P
Purple
No R - get no red precursorNeither purple nor red pigment can be made
White - no pigment
No P – get red pigment but not purple
Precursor 1 Precursor 2
Blue Red
RP
If Pathway 1
Coexpression of Blue and red pigmentderived from different precursorsMakes purple
9R_P_ 3R_pp 3rrP_ 1rrppPhenotypes:
Purple WhiteRed Blue
R no P would make red pigment only
No R but P would make blue pigment,
Fact that loss of R changes phenotype to rr even if Functional P is expressed indicates that P and R affecta common pathway and R is before P in the pathway to make pigment.
Yeast complementation test for next week:
Brewers YeastSaccharomyces cerevisiae:
16 chromosomes12,052 kb DNA6183 ORFsAbout 5800 expected to encode proteins
Yeast is a very useful model for genetics because of its life cycle
Haploid life cycle
Yeast is a very useful model for genetics because of its life cycle
Mating cycleDiploid
We can isolate mutants as haploids
We can test the mutations for allelism by a complementation test
Two haploids are mated. The resulting Diploid has both mutations.
Either the mutations are allelic and do not complement,or they are mutations in two different genes and they do complement.
a1a2
Select mutants that are defective in Adenine synthesis- cannot grow without adenine in medium.
Turn red on media with adenine because an adenine precursor accumulates.
a1
a1
a2
a1
a1
X
X
a1
a1
a2
Which mating results in complementation?
Lab experiment: Corn kernels
R and P
RR or Rr is full color
rr is no color
PP or Pp are full color
pp is weak color
Question is pprr weak color?P epistatic to R or
Is pprr no color? R epistatic to P
End