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BIOL 202 5.6 Phenotypes and gene interactions In humans, look at families that are similar; look for traits as they are distributed in the family—how the phenotype moves LOD scores - Log Of Difference - Look for polymorphisms (e.g. 2 different alleles) in a family - Let M be a molecular marker o M has 2 forms, M1 and M2 Differ by 1 bp M1 is GAATTC (restriction enzyme sequence for EcoR1 M2 doesn’t have this sequence - Let D represent the dominant allele responsible for the disease o Dad, e.g. is heterozygous o Test if disease locus linked to molecular marker o Mated with “test cross”—homozygous recessive at both alleles Look at each indiv born and see probability of getting that indiv Look at probability at different RF o See the odds ratio; then find LOD o Not significant, but do the same for more families with the trait, and see their LODs; then add the LODs together o Get the probability of getting the end locus and the disease o As get closer to the gene, see higher probability Look at different markers; the ones with highest probability of linkage show the highest LOD scores LOD of 3 is statistically significant to p=0.1 Phenotypes and gene interactions - different kinds of dominance o complete dominance (recessive mutations) o incomplete dominance o codominance - Haplosufficiency o 1 (recessive) allele that codes a gene LOF, the other normal gene allele will suffice for completing the function if homozygous for recessive allele, disease/otherwise different phenotype will appear; if heterozygous, will have normal phenotype - Haploinsufficiency BIOL 202 - - - o Dominant mutation Even if 1 normal allele, 1 bad allele (usually just as) bad phenotype If you have t/t, e.g., will have function, but not if have T/t or T/T LOF for all but homozygous recessive o Sometimes need proteins to work as dimers Homo/heterodimers If have protein that functions as dimer, and mutation alters one of the proteins’ shape, could screw up the function Being heterozygous makes proteins completely nonfunctional though have both normal and bad (?) proteins—is dominant Incomplete dominance o Gene encoding an enzyme or protein that, e.g. makes colour If have 0 WT, will have white; 1 WT, will get a lighter colour; if have 2 WT, will get dark colour o E.g. blood type Based on the I gene Encodes different forms of a surface protein that have 3 major alleles o A, B, nothing (i) o People with B blood would be IB/IB for B allele o i makes no protein can see co-dominance: can recognize protein B and A using different antibodies AB has both A and B proteins How characterize phenotype can influence type of dominance seen o Consequence of pleiotropy o E.g. sickle cell allele Encodes less water-soluble protein that keeps blood cells from puffing up Red blood cells have most efficient surface areas for oxygen transfer Heterozygous has both normal and sickle-cell blood cells Wrt anemia, HbS (sickle cells) is recessive to normal HbA But wrt blood cell shape, see incomplete dominance o At molecular level, HbA and HbS are codominant Recessive lethals o E.g. yellow mice Normal mice have dark pigmentation; yellow mice have lighter coats Never get pure yellow mice Couldn’t get purebred BIOL 202 Got 2:1 ratio o One of the genotypes is lethal in utero Mating yellow with normal mice gives 1:1 ratio o Also occurs in other organisms E.g. cats with no tails: end up with 2:1 ratio again; can’t get truebreeders o Some mutations present at different conditions E.g. lethal at higher temperatures; normal at lower temp (e.g. because protein is on verge of instability) Useful for determining mutations E.g. auxotrophs Can screen for these by putting them in minimal media o Just has basic salts and sugars o Supplement media with vitamins and amino acids E.g. find auxotroph of arginine (needs arginine to survive) o Systematically put in medias with and without arginine o If can produce arg from precursor, have 3 proteins in the pathway Look for single genes encoding the enzymes in the pathway (1 gene, 1 enzyme theory) Bombard WT with X-rays to mutagenize o Process with WT neurospora o Put ascospores into individual tubes with media (complete media—permissive conditions for growth) Transferred into minimal media Look for a lack of growth; go back to the original stock of that which didn’t grow in minimal media See if grows if amino acids or vitamins added to minimal media o Grew in presence of amino acids o what’s missing is ability to make one of the amino acids o took same strain out and put in different conditions added single amino acids found that grew in presence of arginine (was deficient in arg) BIOL 202 then looked to see if 1 gene affected all the reactions, or if needed different genes o (found several mutants that didn’t grow; picked out the arg mutants) o found 3 types of arg mutants mated them together arg-1 defect was in enzyme X that formed ornithine: grew if added ornithine; citrulline, arg arg-2 defect was in enzyme y, so grew if added citrulline and arg arg-3 grew only if added arg mutation was in enzyme z Interactions between genes in pathways - Several varieties of, e.g., true-breeding white-flowered mutants can be found; are they mutations of the same or different alleles? o E.g. find in Japan a white version of nightshade, as well as in Nova Scotia find another white nightshade—probably different mutations How prove mutations are/aren’t in the same gene? Using complementation o Only works for recessive mutations o Always start with true-breeding strain o 2 genes control colour of flower, e.g. have different areas with white flowers; know that they’re all recessive. Are they mutations in the same genes? if they’re mutations in the same gene: interbreed and see if end up with white flowers or blue flowers if end up with blue flowers, the 2 complement each other (mutations are in different genes) o get 1 good copy from the other gene—presuming complete dominance if end up with white flowers, mutations don’t complement each other (same gene mutation) o get 2 bad copies of the gene o see that the 2 genes interact genetically to produce blue phenotype (by producing different enzymes that process precursors) - Independence of actions of each gene o If took truebreeding black snake and mated with truebreeding orange snake, end up with camouflage snake BIOL 202 - - - End up with a 9:3:3:1 ratio in the next cross o Back to flowers from before: crossing, e.g. british with Japanese strains, have a 9:3:3:1 genotypic ratio, but not a 9:3:3:1 phenotypic ratio Get blueness if have 1 dominant allele encoding proper enzyme 2 and 1 dominant allele encoding proper enzyme 1 But if homozygous for british mutation (is on W1 gene), end up with homozygous WT for W2 gene, but enzyme 1 blocked (LOF) white flower Same idea if W2 gene is blocked/LOF: end up with white flower again Get 9:7 ratio—still adds up to 16 Interaction b/w regulator genes and target genes o Mutations in regulatory gene, recessive homozygous mutation: gene for protein won’t make protein o But if mutation in protein gene making non-functional protein, same phenotype o Mating dihybrid ends up with 9:3:3:1 genotype ratio but not phenotype ratio because a few genotypes cause the same phenotype Recessive epistasis o If, e.g. enzyme 1 makes pink flower, and enzyme 2 makes blue; enzyme 2 block makes pink; if enzyme 1 blocked, remains white (colourless) o 1 gene affects ability of other gene to make phenotype = epistasis o e.g. Laborador retrievers black, chocolate, golden B = dominant, black ; b = recessive, chocolate Truebreeding chocolate mated to truebreeding golden classic heterozygous; then end up with 9:3:3:1 in selfed cross Dihybrid cross 9 black, 3 chocolate, 4 golden phenotypic ratio B/--; E/-- black b/b; E/-- chocolate B/--; e/e golden b/b; e/e golden o recessive phenotype of one gene affects what you see from the other gene Dominant epistasis o E.g. flowers: D gene affects intensity of redness of pigment D = red; d = pink W gene affects whether pigment can be made w/w = pigment production; W/-- = no pigment production W/-- ; D/-- white Bred to a truebreeding red (w/w; D/--) white BIOL 202 - If have W allele, doesn’t matter whether have D or d, or 1 w will be white because of dominant epistasis