Empirical population genetics training course for beginners Thierry De Meeûs

1. Empirical population genetics training course for beginners Thierry De Meeûs Interactions hôtes - vecteurs - parasites dans les infections par des trypanosomatidae - (INTERTRYP), UMR IRD/CIRAD 177, Montpellier France. WHO Collaborating Centre for research on host/vector/parasite interactions for surveillance and control of Human African Trypanosomiasis (+226) 20 97 00 94 • (+226) 76 86 40 88 (Burkina-Faso) • (+33) (0)6 19 83 52 60 (France) Fax: (+226) 20-97- 23-20  thierry.demeeus@ird.fr • http://t-de-meeus.fr/TdeMeeus.html

2. Lecture 1

3. Introduction

5. Direct methods Indirect methods

6. ♂ ♀ ♀ ♂ ♂ ♀ ♂ ♀ The detection of genetic variation Cytpolasmic markers

7. The detection of genetic variation Nuclear markers AA Aa aa Dominant markers [A] [a] RAPD (Randomly Amplified Polymorphic DNA) ATGCAC TACGTG TCATGA AGTACT Random PCR primers ATGATC TACTAG AATCTG TTAGTA Presence or absence of amplified product=>Dominant marker Recessive genetic diseases

8. CTCTCTCT AGAGAGAG CTCTCTCTCT AGAGAGAGAG + - The detection of genetic variation: co-dominant markers A1A1 A1A2 A2A2 Microsatellites Primer1 Primer2 mRNA Primer1 AUGCAGCCAUAGGCG Primer2 PCR Enzymes Phe-Pro-Leu-Ileu-Val + - Electrophoresis RFLP, MLST, SNP… Neutrality=an important hypothesis for inferences

9. Population structure Reproductive strategy, size of demographic units and migration (or dispersal)

10. I. Notions of molecular biology and formal genetics

11. DNA, universal molecule for the transmission of genetic information, the molecule of life

12. DNA, universal molecule for the transmission of genetic information, the molecule of life

13. DNA, universal molecule for the transmission of genetic information, the molecule of life

14. DNA, universal molecule for the transmission of genetic information, the molecule of life Purines: Bases, essential nucleotide constituents, themselves basic elements of nucleic acids (RNA and DNA), complementary of pyrimidines. There are two of them: adenine (A) and guanine (G). Pyrimidines: Bases, essential nucleotide constituents, themselves basic elements of nucleic acids (RNA and DNA) , complementary of purines. There are three of them: thymine (T), uracile (U which takes the place of T in RNA) and cytosine (C). A is complementary of T and U G is complementary of C

15. DNA, universal molecule for the transmission of genetic information, the molecule of life The genetic code =>the genetic code is degenerated

16. DNA, universal molecule for the transmission of genetic information, the molecule of life

17. Viruses? 2. The three domains of life 3 to 3.5 Billion years ago Prokaryotes

18. Prokaryote genetics Enveloppe of peptidoglycanes

19. Prokaryote genetics Transformation Conjugation Plasmid exchange Transduction

20. Eukaryote genetics

21. Eukaryote genetics

22. Mutualism/Parasitism Rafflesia Coral Hydra viridis Tridacna gigas Anthopleuraelegantissima Elysiachlorotica Zooxhantella Dinophyceae Zoochlorella HGT: Horizontal genetransfers

23. Eukaryote genetics Plant cell Animal cell A no-plant cell in fact

24. Eukaryote genetics Curling DNA duplication in the nucleus

25. Eukaryote genetics Mitosis a distinguishing feature of eukaryotic cells

26. Eukaryote genetics Meiosis, another distinguishing feature of eukaryotic cells Sexual reproduction has two consequences: segregation and recombination

27. Eukaryote genetics Meiosis is a distinguishing feature of eukaryotic cells that appeared about 850 millions years ago 2N 2N N Anisogamy Repairing the deleterious effects of horizontal gene transfers and/or correcting polyploidy that are thought to have occurred often during the Proterozoic era N Isogamy

28. Eukaryote genetics Isogamy a b d c e Conjugation mechanism in paramecia Association of 2 paramecia by their oral side Degeneration of macronuclei and starting of meiosis Meiosis results in 4 haploid nuclei, 3 of which degenerate The remaining nucleus divides One of the nuclei migrate to the associated cell The 2 nuclei in each associated cells fuse into 1 nucleus The nucleus divides several times and develop into a macronucleus and a micronucleus H. The paramecia divides Sea lettuce (Ulva lactuca)

29. Eukaryote genetics Anisogamy

30. Eukaryote genetics Anisogamy

31. Eukaryote genetics Introns and splicing, a distinguishing feature of eukaryotic DNA transcription (non spliceosomic introns exist in prokaryotes and even in some viruses) Purine Pyrimidine Splice donor site Branch site Splice acceptor site 5' 3' Pyrich A/C A G G U Pu A G U C U Pu APy N C A G G 20-50 bases Exon 1 Intron Exon 2 snRNPs (small nuclear ribonucleoproteinparticles) complexes of snRNAsand proteins Spiceosome Exon 2 Exon 1 5' 3' Excised intron in lariat shape 5' 3' Mature mRNA

32. 3. Mutation, the key to genetic variation and evolution Point mutations Transition: Replacement of a purine by another purine (A<=>G) or of a pyrimidine by another pyrimidine (C<=>T) (antonymic to transversion). Transversion: Replacement of a purine by a pyrimidine or of a pyrimidine by a purine (A<=>T, A<=>C, G<=>C, G<=>T) (antonymic to transition), twice less likely. Insertions and deletions Transposable elements (transposons) and retroviruses Chromosome mutations (translocations, inversions) IAM: Infinite Allele Model (if identical, alleles necessarily are identical by descent) KAM: K allele model (homoplasy: identity not necessarily by descent) SMM: Stepwise Mutation Model (microsatellite loci)(-CACACACA-) TPM: Two Phase Model (SMM+KAM)

33. CTCTCTCT AGAGAGAG CTCTCTCTCT AGAGAGAGAG 4. Genetic variation: genotype and phenotype BbCc=Genotype [B]=Phenotype Phenotype does not necessarily reflect genotype Microsatellites Primer1 Primer2 Primer1 Primer2 + -

34. 5. Genetic variation: mendelian heredity Gregor Mendel (1822-1884)

35. 5. Genetic variation: complex traits Enzyme 1 Enzyme 2 Enzyme 3 Enzyme 4 C A B D E [+] or wild Phenotype Enzyme 1 Enzyme 2 Enzyme 3 Enzyme 4 C A B D E [-] or mutant phenotype Enzyme 1 Enzyme 2 Enzyme 3 Enzyme 4 C A B D E [-] or mutant phenotype Diploid individual Enzyme 1 Enzyme 2 Enzyme 3 Enzyme 4 [+] or wild Phenotype Enzyme 1 Enzyme 2 Enzyme 3 Enzyme 4 Complementation Epistasis Enzyme A_B Character 1 Character 1 Enzyme 1 A Pleiotropy A_b or a_B or a_b Enzyme Character 2 Character 2

36. 5. Genetic variation: recombinaison Thomas Hunt Morgan (1866-1945)

37. 5. Genetic variation: recombinaison Thomas Hunt Morgan (1866-1945) Hybridization AABBaabb B A F1: 100% AaBb r b a Back cross: AABBAaBb (1-r) /2 AB, r/2 Ab, r/2 aB, (1-r)/2 abAB (1-r) /2 AABB, r/2 AABb, r/2 AaBB, (1-r)/2 AaBb F2: AaBb AaBb r100=[f(AABb)+f(AaBB)]100=Genetic distance in centimorgans

38. 6. Inbreeding (F), kinship (φ), relatedness (r) and pedigrees FA=0 FD=(1/2)6×φAGM-AGP A FD=(1/2)7=1/128≈0.00078 FAGP=FAGM=0 AGP AGM 1/2 1/2 FGP=FGM=φAGM-AGP GM GP 1/2 1/2 FP=FM=φGM-GP P M Relatedness rxy=2φxy 1/2 1/2 FD=φM-P D Probability that A gives the same allele to AGM and AGP=φAGM-AGP=P(●● or ○○)+P(●○)FA =(1/2 ×1/2)+(1/2 ×1/2)+1/2×FA=1/2×(1+FA)=1/2 Probability that the same common allele in AGM and AGP is found twice in D=(1/2)6

39. 6. Inbreeding, kinship, relatedness and pedigrees FD=(1/2)(C1+1)×(1+FA)+(1/2)(C2+1)×(1+FAGP) FAGP=FA=0 FD=(1/2)7+(1/2)4=(1/128)+1/16≈0.07

40. 6. Inbreeding, kinship, relatedness and pedigrees FD=(1/2)(CA+1)×(1+FA) +(1/2)(CAGP+1)×(1+FAGP) +(1/2)(CAGM+1)×(1+FAGM) +(1/2)(CGP+1)×(1+FGP) FAGM=FAGP=FA=0 FGP=(1/2)(CA/GP+1)(1+FA)=(1/2)3 FD=(1/2)7+(1/2)5+(1/2)5 +(1/2)3×(1+(1/2)3) FD=(1/2)7+(1/2)4 +(1/2)3+(1/2)6 FD=1/128+1/16+1/8+1/64=0.211

41. Lecture 2

42. II. Population genetics

43. 1. The population the basic unit in ecology, a demographic notion A group of individuals sharing the same demographic parameters Population 1 Population 2 Population 4 Population 3 N1 N2 N4 N3 Multiplication and migration Regulation N1 N2 N4 N3 Populations of constant size

44. 2. The model of Hardy and Weinberg William E. Castle (USA) (1903) Hardy G.H. (GB) (1908) and Weinberg W. (G) (1908) Hermaphrodites or self compatible monoecious populations A single population The size of the population N=∞ Sexual and panmictic reproduction No mutation No migration No selection Non-overlapping generations (discrete)

45. f( ) = f( ) = q f( ) = p² ; f( ) = 2pq ; f( ) = q² Hardy-Weinberg proportions Table of gametes and zygotes formed under the panmictic hypothesis Gametes = p +

46. AA Dt Aa Ht aa Rt Hardy-Weinberg equilibrium ft(A)=pt, ft(a)=qt=1-pt Panmixia (hermaphrodites) Population size N~∞ Migration m=0 Mutation u=0 No selection Discrete generations No hypothesis needed In one generation

47. Hardy-Weinberg equilibrium with three alleles AA At AB Bt AC Ct BB Dt BC Et CC Jt ft(A)=pt, ft(B)=qt, ft(C)=rt=1-pt-qt No hypothesis needed In one generation

48. Hardy-Weinberg equilibrium with dominance AA Aa aa Rt Dt If panmixia p² 2pq q² ft(A)=pt, ft(a)=qt=1-pt Hypothesis: the population matches panmictic proportions: A (very) strong hypothesis

49. 5. Hardy-Weinberg whenNsmall: genetic drift Panmictic population of size N, no mutation, migration or sélection and discretegenerations 4 1 2 3 N ..... Individuals 2N 1 2 4 3 ................................... Alleles Frequency of allelei : ................................... Gametes Frequency of allelei in the pool of gametes: Probability to drawtwiceallelei in the gametes: Piitrue for each of the 2Nalleles Probabilitydrawtwoidenticalallels in the gametes:

50. those already identical at generation t Those that become identical at generation t+1 Genetic diversity 3. Hardy-Weinberg when N small: genetic drift Let a population of small size N be panmictic, without mutation, migration or selection and with non overlaping generations Ft: probability to draw two identical by descent alleles in this population at generation t Alternative demonstration Come from the same allele at t Come from twodifferent alleles at t thatwere identical at t