Co-dominance: heterzygote shows phenotypes of both alleles fully/equal Incomplete dominance: two alleles of heterzygote interact resulting in intermediate

phenotype Forward geneticsobserved phenotype then look for causal genetic differences Begins with observed variation in phenotype Looking for patterns of inheritance in descendents of crosses b/t individuals w/different phenotypes Primary purpose: to find the normal pathway of metabolism and development by looking at mutants I) Mutations are individually mapped II) Making crosses b/t strains puts mutations in different combinations III) Characterize DNA of different variants (alleles) of genes and explain from variant DNA sequences the differences in structure, function, amounts, and localization within the organism of the metabolically active molecules Reverse Genetics: normal DNAcreate mutationanalyze function of DNA sequence by observation of phenotypic changes. Advantage: large number of mutations of specific kinds can be made Southern Blotprobing for specific DNA Northern blotprobing for a specific RNA To determine if a gene is being transcribed in a tissue Western blotprobing for a specific protein Usually formed with antibodies Developmental noise--Random events in development that lead to variation in phenotype Mendels law of equal segregation--in meiosis, the members of a gene pair separate equally into eggs and sperm SINGLE GENE INHERITANCE 1:1 for Y/y x y/y 3:1 for Y/Y x Y/y 1:2:1 for Y/y x Y/y PHENOTYPIC RATIONS Monohybrid test-crossed: 1:1 Monohybrid selfed: 3:1 Dihybrid testcrossed (ind. Ass.): 1:1:1:1 Dihybrid selfed (ind. Ass): 9:3:3:1 Trihybrid test-crossed (ind.): 1:1:1:1:1:1:1:1 GENE INTERACTIONS 9:7 ratiogenes in same pathway Double mutant has same phenotypes as two single mutants Each mutant allele controls different step in same pathway Can also come from gene regulation o Regulatory gene produces protein that binds to regulatory stream upstream of target gene o Target gene would be transcribed at low levels Absence of either gene function leads to absence of end product of the pathway 9:3:4 ratiorecessive epistasis Double mutant shows phenotype of one mutation but not other o Epistaticoverriding mutation o Hypostaticoverridden mutation Results from genes in same pathway Epistatic mutation is in gene upstream of overridden one Can be developmentally downstream Typically 3 phenotypes segregate 12:3:1 ratiodominant epistasis Suppressormutant allele that reverses effect of mutation of another gene, results in wild-type or near wild-type phenotype Gene products normally interact Sometimes have no effect in absence of other mutation or can produce own abnormal phenotype Screening Expose mutant to mutation-causing agents Screen for wild-type descendents o Most will by revertants (reversals of org. mutational event) o Some will be pseudorevertantsdouble mutants w/one being suppressor

Mutations Amorph: Null allelesabsence of a normal gene product or absence of normal function. Hypo-morph: Leaky allelemutation that causes reduced function of a gene Neomorph: allele does something new Anti-morph: dominant negative (- affects normal product) Hypermorphincreases in wild type activity/product Haplosufficient--recessive Provides enough gene product to carry out the normal transactions of the cell. In a heterozygote the remaining copy of the + allele provides enough protein product for normal function Haploinsufficientnull allele is dominant b/c heterozygotes single wild type allele cant provide enough product for normal function TYPES OF CROSSES Monohybrid crosstwo heterozygous individuals (i.e. Y/y x Y/y) Test crossto determine if an individual is heterozygous or homozygous dominant Cross individual with tester Tester is homozygous recessive Selfalternative test for homozygosity, if 3:1 ratio in progeny, the individual is a heterozygote Hemizygousin males, the differential region that contains the most genes has no counterpart on other sex chromosomes Pseudoautosomal regions (1 and 2) at the tips of each sex chromosome. One/both regions pair and undergo crossing over. Autosomal-like because they are homologous PEDIGREE PATTERNS (Square=man, circle=woman, dot in middle=carrier of sex-linked recessive) Autosomal recessive Disorder appears in the progeny of unaffected parents Affected progeny are both males and females Pedigrees tend to look bare Shows up in groups of affected siblings People in earlier and later generations not affected Assume people who marry into the family are homozygous normal Includes cystic fibrosis and human albinism

When calculating a probability for an off-spring being affected, and the last effected happens to be a grand grand parent, and the prevalence is given (i.e. 1:10,000). Consider the probability of carriers marrying in: 1/(chance of being affected) Autosomal dominant Phenotype appears in every generation Equal representation of both sexes

Virtually all matings that produce progeny with disorders are A/a x a/a Includes Dwarfism, Huntingtons, and polydactyl Autosomal polymorphisms Co-existence of two or more common phenotypes of a character Assume people who marry into the family are heterozygous (if not affected) Often inherited as alleles of a single gene X-linked recessive Many more males than females show the rare phenotype None of the offspring of affected males show phenotypes o All daughters are carriers o In next generation, half of the sons of the carriers show the phenotype None of the sons of an affected male show the phenotype Assume a normal phenotype of unknown genotype is homozygous X-linked dominant Affected males pass condition to all their daughters, none of their sons Affected heterozygous females married to unaffected males pass condition to their sons and daughters Y-linked--Only affect males Mitochondrial inheritance:

In diploids, show different F2 ratios (13 normal : 3abnormal) Only two phenotypes segregate Modifiersmutation at second locus that changes degree of expression of mutated gene at 1st locus Can down-regulate or up-regulate transcription of gene Appearance of two grades of mutant phenotypes within mutant progeny 9:3:3synthetic lethalsdouble mutants are lethal

Mutant female x wild-type maleprogeny all mutant Wild-type female x mutant maleprogeny all wild type

Polygenic inheritance Show continuous variationtypically in characters that can take a measureable value b/t two extremes Many times has a purely environmental basis Polygenes (aka quanitative trait loci (QTLs))interacting genes underlying hereditary continuous variations

Amount of doses can affect phenotype over spectrum

Mendels law of independent assortment-If genes are on different chromosome pairs they will act independently at meiosis Same chromosomeseparated by /, different chromosomesseparated by ; Dihybrid crosses-- Number of distinct genotypes produced = (Number of genotypes for each pair)number of genes MAPPING BY RECOMBINANT FREQUENCY Three-point testcrossTrihybrid x triply recessive tester (i.e. A/a;B/b;C/c x a/ a;b/b;c/c) Count # of recombinants for two loci at a time Should double-count double-recombinants Two at high frequency Two at intermediate frequency Two at a different intermediate frequency Two raretypically double-recombinant classes If given M.U., M.U.=recombinant frequency, each parent will be 100-RF/2% Multiple crossovers Mapping functionrelates observed recombinantfrequency to map distance corrected for multiple crossovers True determinant of RF is relative sizes of classes with no crossovers in a specific chromosomal region

Interse cross: monohybrid cross (A/axA/a) Phenotypic: 3:1, Genotypic 1:2:1 Test Cross: A/- x a/a Complementation test: crosses two diff. homozygous mutants to check if one rescues normal phenotype Double mutants: test to create double mutant for two diff. mutations to look for new phenotype (if new, interact at some level in a pathway) Case-control test: sample from pop. Individuals w/ and w/o disease, compare allele frequencies at markers, if sign. Diff. from one another, suggest assn. bt/ disease and marker Interference (I)=1-(observed frequency or # of double recombinants/expected frequency or # of double recombinants) Penetrancepercent of individuals with a given allele who exhibit phenotype associated with allele, Percentage of cases in which gene is expressed

Incomplete penetrance can be due to: Environment influence, Other interacting genes, Subtlety of mutant phenotype Expressivitydegree to which a given allele is expressed at phenotypic level (intensity of phenotype)level of expression Linkage disequilibriumalleles at one gene correlated with alleles at second locus on pop. Levelassociated b/t two markers. Broken by recombination. Called association mapping.

RF=1/2 (1-e-m) (m=mean number of successes)convert to map distance by multiplying by 50

GENETIC MARKERS Single-nucleotide polymorphisms (SNPs):a nucleotide difference b/t people Silent SNPs within genestags for particular alleles with no affect on phenotype SNPs in which one SNP allele causes a mutant phenotype showing single-gene inheritance--One allele of SNP alters gene function SNPs in polygenes (QTLs)-help with discovery Intergenic SNPsno measurable phenotypic change, useful milestones to find other genes RFLPs (Restriction fragment length polymorphisms)located at restriction enzymes target site, dont require sequencing to find Mapping by using SNP haploytypes (chromosomal segment defined by specific array of SNP alleles that it carries, inherited down through generations as blocks) Combos of SNPs can be used to find linkage SNP alleles of haplotype have linkage disequilibrium (finite lifetime) Haplotypes can be used to map position of gene of interest (hapmap) Simple sequence length polymorphisms (SSLPs or VNTRs)adjacent multiple repeats of short, simple DNA sequences, tandem repeats, When cut with restriction enzymes, fragment will be proportional to number of repeated elements Minisatellite markersrepeating unit from 15-100 nucleotides long o Total genomic DNA is cut with restriction enzyme with no target sitescuts flanking sites o Southern blot o Used as DNA fingerprints Microsatellite markersbased on variable numbers of tandem repeats of a dinucleotide , Most common is CA (GT) o Revealed with polymerase chain reaction (PCR) Polymerase chain reactionto amplify DNA in vitro If we know sequence of at least some parts of gene

Uses multiple copies of primers (15-20 bases long) that bind to different end of gene to be amplified, Primers bind to opposite DNA strands with their 3 ends pointing at each other, location of primers determines specificity of DNA segment that is amplified, Taq Polymerase adds bases to primers, Polymerization process shuttles back and forth b/t them, Forms exponentially long # of double-stranded DNA molecules

Tools and TESTS Chi-squared (2) testcheck ratios against expectations To make judgment of whether observed #s are close enough to the expected

X2=(Observed-Expected)2/Expected for all classes Degrees of freedom=number of independent variables in the data=number of genes-1 Null hypothesis: no linkageindependent assortment or due to chance, reject if >5%, p<5% maybe linkage df=(number of classes in rows-1)x(number of classes in columns-1)