Quantitative Genetic Perspectives on Loss of Diversity in Elite Maize Breeding Germplasm Jode W. Edwards USDA ARS CICG

Outline Diversity Population genetics of maize Quantitative genetic processes –Bottlenecks –Selection Implications

What is diversity? D = 1 –  p i 2 –p i = allele frequency At Hardy-Weinberg equilibrium D is an estimator of heterozygosity, H With population subdivision, heterozygosity is related to F st: –H t = (1- 1 / 2N ) t H 0 = (1-F st )H 0 Sources: Nei, M, 1973, PNAS, 70: ; Wright, S., 1943, Genetics, 28:

Diversity in Maize Inbreds and Landraces Tenaillon, Sawkins, Long, Gaut, Doebley, and Gaut, 2001 Estimated SNP diversity by sequencing –7 known genes, –6 cDNA clones –8 RFLP clones –All chromosome 1 Germplasm –16 exotic landraces (1 inbred per landrace) –9 U.S. inbreds (B73, Mo24W, Mo17, W153R, Ky21, NC258, Oh43, Tx601, T8) Inbreds contained 77% as much diversity as the landraces (D I /D L ) Source: Tenaillon et al., 2001, PNAS, 98:

‘the U.S. inbred sample retains a high proportion of diversity, which is difficult to explain given that U.S. elite germplasm has a narrow origin based largely on two open-pollinated varieties, Reid yellow dent and Lancaster (14)’ –[“(14)” is Major Goodman’s paper in Heredity] Tenaillon et al. Conclusion

Is 77% Hard to Explain? 1 - F st = For F st of 0.23, N=2.2 If inbreds were: Sampled randomly, E[1-F st ] = 0.89 Subpopulation with Fst = 0.87, E[1-Fst] = 0.77

How should we measure diversity? Heterozygosity (formally)? Number of alleles? Number of polymorphic loci? Number of rare alleles? JE thoughts: Diversity is important, but we don’t know how to measure it (or what it is) Something else may be more important

Sustainable Selection Response Plant breeders’ main goal is selection –Short term: Maximum response –Long term: Sustainable response In order to address sustainability of selection response, we need to understand phenotype –Population genetics of maize –Quantitative genetics of population bottlenecks –Quantitative genetics of selection with finite size

Maize Population Genetics: BSSS Started with maize land races (O.P.) and develop ‘first cycle’ inbreds 16 lines intermated to form BSSSC0 –Expected diversity = 87.5% of ancestor B14, B37 emerge from Cycle zero –Expected diversity =.875 x.5 = 43.75% B73, B84 emerge from C5, C7

Corn Belt Maize Land Races Outcrossing, monoecious populations Large N e (?) Mass selected for visual characteristics (low h 2 ?) Corn belt dents existed 100+ generations, longer for other groups Corn belt dents (Labate et al., 2003) –Accessions: F st = 0.15 –Varieties: F st = 0.04 “Almost” one large randomly mated population Source: Labate, J.A. et al., 2003, Crop Science, 43:80-91

Maize Land Races Hardy-Weinberg equilibrium Linkage equilibrium Mutation-selection equilibrium

Haldane (1937) Principle Mutation frequencies determined by equilibrium –New mutations are constantly added to the population –Mutations removed by selection (and drift) –Mutation rates estimated to be 0.4 – 1.0 per diploid individual per generation At equilibrium –Individuals carry many mutations –Reduction in fitness due to mutations = “genetic mutation load” (Muller) Source: Haldane, J.B.S., 1937, The American Naturalist, 71: ; Crow, J.F., 1993, Oxford Surveys in Evolutionary Biology, 9:3-42

Does Mutation Load Apply to Maize? Inbreeding depression –Severe in first cycle inbreds –Less in germplasm with inbreeding history (purging of recessives) –If many loci carry mutations, complete purging takes many generations Observation of major “lethal” mutations Empirical work in maize is needed!

Significance of Haldane Principle Mutation load provides a model of quantitative genetic variation more realistic than ‘infinitessimal theory’ Provides a basis for understanding quantitative genetic variation, and thus, Basis for predicting effects of bottlenecks and artificial selection

Bottlenecks Population is formed from small number of individuals –Change allele frequencies –Hardy-Weinberg and linkage disequilibria Under additive model –‘within subpoplation variance’, V w = (1-F st )  2 A –‘among subpopulation variance’, V b =2F st  2 A Non-additive model: effects of bottlenecks are complex Source: Wang, J., et al., 1998, Genetics, 150:

Edwards and Lamkey (2003) Source: Edwards and Lamkey, 2003, Crop Science, 43:

Garcia, Lopez-Fanjul, and Garcia-Dorado, 1994 D. melanogaster, Full-sib lines Source: Garcia, N., et al., 1994, Evolution, 48:

Gene Effect Sizes Wang, Caballero, Keightley, and Hill, 1998 Source: Wang, J., et al., 1998, Genetics, 150:

Gene Effects and Bottlenecks Genes of all sizes important in the base After a bottleneck: large recessives become much more important (and hence large increase in dominance) Explanation: Nonlinear relationship between frequency and variance: small increase in frequency = large increase in variance

Limits to Selection Response Robertson, 1960 Max response = 2 N e times initial response Half-life occurs at 1.4 N e generations Total response is maximized at 50% intensity (greater with linkage) Based on ‘infinitessimal’ theory –Many genes of ‘infinitely’ small effect –Can we understand ‘side effects’ of selection under more realistic conditions? Source: Robertson, A., 1960, Proc. Roy. Soc. London, Ser. B, 153:

Selection Effects Loss of heterozygosity (diversity) Linkage disequilibrium –Bulmer –Hill-Robertson Epistasis

Linkage and Selection Bulmer effect –Correlation between alleles induced by selection –Causes excess of coupling phase linkages and reduced genetic variance Hill-Robertson effect –Effect of repulsion phase linkages –Unfavorable alleles become fixed because of selection for favorable alleles linked in repulsion phase Sources: Bulmer, M.G., 1971, American Naturalist, 105: ; Hill, W.G. and Robertson, A., 1968, Theor. Appl. Genet., 38:

Zhang and Hill, 2005 Simulated selection in cage populations derived from ‘equilibrium natural populations’ of D. melanogaster Conditions –Genetic model: mutation-selection balance under joint pleiotropic and stabilizing selection –40% intensity –Recombine 40 individuals –V G0 = 0.5 V E –3 chromosomes of varying length Source: Zhang, X.S., and Hill, W.G., 2005, Genetics, 169:

Selection and Linkage Zhang and Hill, 2005 Source: Zhang, X.S., and Hill, W.G., 2005, Genetics, 169:

Gene Numbers and Effects Zhang and Hill, 2005 Distribution of gene effects –90% of genes have a<0.1  p and account for 27% of genetic variance –10% of genes have a>0.1  p and account for the rest of the genetic variance Estimated that 10 3 – 10 4 loci are polymorphic in a cage population Source: Zhang, X.S., and Hill, W.G., 2005, Genetics, 169:

Evidence of Linkage in Maize Degree of dominance, d, can be estimated as a ratio,  D 2 /  A 2, in F 2 -derived populations Linkage disequilibrium causes a bias called ‘associative overdominance’ Random mating breaks up linkage and reduces bias AAAa -> d=1 aa Aa -> d=2 Aa -> d=0

Maize NCIII Experiments Lonnquist, J.H., 1980, Anal. Acad. Nac. Cs. Ex. Fis. Nat., 32: ; Gardner, C. O., Personal communication to E.T. Bingham

Epistasis Favorable epistatic interactions are increased by selection Lamkey, Schnicker, and Melchinger, 1995 –Began with BSSS lines B73 (cycle 5) and B84 (cycle 7) –Formed the F 1, F 2, BC 1 (to both parents) and intermated F 2 –Testcrossed all generations onto Mo17 –With additive model (no epistasis) there is a linear relationship among generations Source: Lamkey, K.R., et al., 1995, Crop Science, 35:

Epistasis in B73 and B84 Lamkey, Schnicker, and Melchinger, 1995 Source: Lamkey, K.R., et al., 1995, Crop Science, 35:

How did we get here? Bottleneck followed by 5 and 7 cycles of selection During selection –Linkage disequilibrium increases –Epistatic combinations become more important –Selection may be dominated by genes of large effect Slow increase in frequency of many small favorable alleles is not a good model –For positive effects, i.e., response –For negative effects

Sustainable Response is a Function of More than Diversity Loss of alleles (diversity) Increase in linkage disequilibrium (reduced variance) Increased dependence on specific epistatic combinations Shift in size of genes that contribute to genetic variance (small to big)

Implications for Elite x Exotic Crosses Genetic variance within a single population is due mostly to genes of large effect Linkage disequilibrium within the cross may reduce genetic variance Any new alleles from the exotic parent are preferentially lost if: –Linked to negative alleles at physiologically selected loci, e.g., photoperiod –There are favorable epistatic interactions among elite alleles

What can be done? Map major genes (especially photoperiod) and use markers to break linkages Recycle lines from different crosses Enhance or improve land races directly to maintain more variation and reduce disequilibrium –If major genes were identified, could speed up with markers –Preserve more variation due to genes of small effect Random mate individual crosses

Basic Research Questions How differentiated are maize land races from each other and from elite lines? –At neutral loci –At selected loci Can we identify major genes that –Differentiate elite lines from ancestral varieties –Corn belt dent from tropical races Genetic architecture –Can we estimate mutation load parameters? –Can we distinguish purging of recessive load from selection for physiological effects

We can succeed doing what we are already doing However, can we be more successful?