This map of rudamentary alleles does not imply anything about where the various mutations might lie on a meiotic map It is simply a schematic representation of a collection of data from a series of complementation tests designed to determine functional allelism Let s consider three r mutant alleles did they complement Y or N a b Yes they don t overlap on the map and the hybrid fly looked wildtype did they complement Y or N a h No they do overlap on the map Consider a new r mutant allele rz3 z3 a z3 h z3 i Which heteroallelic combination hybrid is most likely to have a mutant phenotype z3 i Which heteroallelic combination is most likely to generate a wildtype allele during meiosis no basis for a determination This map tells us next to nothing about the possible molecular basis for the complex complementation pattern but so long as we have reason to believe that group i contains at least some point mutants all the mutants on this map are likely to be in the same thing that we want to call a gene and i is the most frequent class Mutations changes in DNA the lifeblood of genetic analysis 1 What kinds can we make categories 2 How do we make them mutagenesis 3 How do we find them mutant screens selections 4 Why bother Fig 20 2 a fruit fly 9h after fertilization head tail wildtype ftz ftz amorph null mutant homozygote it s recessive lethal something Thomas Hunt Morgan never guessed among other things you can get a huge amount of phenotypic information out of the skin of a dead maggot by 16h after fertilization even if doomed denticle belt pattern on the larval cuticle cuticle skin tho actually skeleton denticles maggot tire treads reveals where fly cells think they are in space Wieschaus and N sslein Volhard Nobel Prize 1995 Fig 20 19 maximally informative mutant phenotypes for understanding metazoan pattern formation Kr ppel wildtype hunchback knirps ftz is in the pair rule family not the gap family of mutant phenotypes Mutations changes in DNA the lifeblood of genetic analysis fact it s easier to mess things up than to make them better hence most mutations with any functional consequence mutant phenotype disrupt normal wildtype gene function Geneticists like to mess DNA up to discover what genes do what so they can tell biochemists where to look to learn how those genes do it and sometimes geneticists can learn a thing or two about how even before the biochemists enter the picture Forward genetics mutant phenotype functional consequences of disruption wildtype molecules to infer wildtype function Reverse genetics mutant phenotype wildtype molecules including transcription units functional consequences of disruption to infer wildtype function Mutations the lifeblood of genetic analysis In the context of our current goal infer wildtype function Not Mendel s goal predict the appearance and breeding behavior of hybrids Not Morgan s goal learn how inherited information is transmitted how it changes neither had any hope of discovering what gene are 1 What kinds can we make 2 How do we make them 3 How do we find them Mutations the lifeblood of genetic analysis In the context off our current goal to infer wildtype function 1 What kinds can we make functional categories Herman Muller 1930s In what ways can mutations affect normal gene function 2 How do we make them mutagenesis Muller spontaneous radiation induced 3 How do we find them mutant screens selections Muller exploited giant polytene chromosomes invented balancer chromosomes Muller categorized mutations with respect to change in gene function relative to wildtype Loss of wildtype function l o f mutant alleles generally recessive a a one functional copy suffices 1 so long as all other genes ok 2 so long as we don t look too hard clear exception l o f mutations in haploinsufficient genes are dominant by definition Minute mutations in flies Df M M lof alleles causing cancer identify tumor suppressor genes Gain of over wildtype function g o f mutant alleles generally dominant AntpX Antp fly leg where antenna should be often misexpression wrong time place gof alleles causing cancer identify proto oncogenes Loss of wildtype function l o f mutant alleles complete lof amorph ic null Important goal of genetic analysis define the null phenotype partial lof hypomorph ic leaky phenotypic series set of alleles with progressively less function Gain of over wildtype function g o f mutant alleles too much of a good thing hypermorph ic something new different neomorph ic different in kind e g fusion protein wrong time place ectopic expression antagonizes poisons wildtype antimorph ic dominant negative to infer the normal function of a gene LOF alleles simplest to interpret GOF alleles usually more interesting but generally harder to interpret especially if don t know amorph null phene Can use GOF alleles to generate LOF alleles relatively easily revert suppress the dominance of a GOF allele LOF allele and thereby establish allelism between the GOF and LOF alleles consider the fly gene ovo as a good example ovoD ominant 1 dominant female specific sterile antimorph ovoe8K recessive female specific sterile hypomorph normal ovo function establish female identity of fly germ cells how do we even know ovoD1 and ovoe8K are alleles ovoD1 ovoe8K ovoe8K tumorous female germline wildtype female germline wildtype female germline ovoe8K ovoe8K tumorous female germline ovoD1 Df ovo complementation test ovoD1 ovoe8K wildtype or mutant neither would tell us anything cis trans test if in same gene cis phenotype trans ovoD1 ovoe8K ovoD1 ovoe8K tumorous less tumorous or wt if in different genes cis phenotype trans ovoD1 ovoe8K ovoD1 ovoe8K not haploinsufficient tumorous tumorous if in same gene cis phenotype trans tumorous less tumorous or wildtype ovoD1 ovoe8K ovoD1 ovoe8K but how construct Consider a slightly different cis trans tumorous ovoD1 ovo null wildtype ovoD1 ovo null ovoD1 what we have what we want to generate but what if true reversion ovoD1 ovo null if homozyote is tumorous 2 now do complementation test to distinguish the alternatives tumorous mutagenize to revert the dominance wildtype 1 look at homozyogote ovoD1 ovo null ovoe8K vs ovoe8K tumorous wildtype ovoD1 ovoD1 ovo null tumorous mutagenize to revert the dominance wildtype Followed by a complementation test ovoD1 ovo null ovoe8K vs ovoe8K tumorous wildtype We have to lose the gene function that ovoe8K is missing in order to revert suppress the dominance of ovoD1 hence we have established that ovoD1
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