2. ABC for the AEA Basic biological concepts for genetic epidemiology Martin Kennedy Department of Pathology Christchurch School of Medicine

3. Topics n DNA, chromosomes & genes n Transcription and translation n Sex, meiosis, and all that n Mendelian inheritance n Molecular biology methods n Types of DNA variation

4. DNA, chromosomes & genes

6. Chromosome structure

7. Base pairing

8. Eukaryotic genes are split Exons - fused to form mRNA Introns - spliced out of mRNA (intron = intervening sequence)

9. Genome statistics 3 billion basepairs 23 pairs of chromosomes 50-60% repeats 5-10% coding sequences

10. Transcription & translation

11. Typical gene structure promoter AAAAAA nuclear RNA transcription messenger RNA AAAAAA AUG TAG 5’ UTR 3’ UTR splicing

12. Ribonucleic acid (RNA)

13. From gene to protein

14. The genetic code

15. Proteins

16. Protein folding

17. Summary

18. Sex, meiosis, and all that

19. Definitions n Gene A functional unit of inheritance n Genome All of the genetic material in the chromosomes of an organism n Allele Alternative forms of a gene or marker due to changes at the DNA level n Locus The physical location of a gene n Genotype The specific alleles present for a given gene or set of genes n Haplotype A particular combination of alleles in a specific region of a chromosome

20. Cell divisions Mitosis Daughter cells Meiosis Gametes

21. Recombination Meiosis Gametes

22. Mendel’s laws Good for peas, people and anything else that has sex Segregation: The reproductive cells of hybrids randomly transmit either one or the other of paired parental alleles to their offspring. The alleles are unchanged during passage through each generation. Independence: When individuals with different alleles of more than one gene are crossed, alleles of each gene are assorted into the offspring (segregated) independently of the others. This law applies only when there is no linkage between the genes.

23. Mendelian inheritance n Mendelian conditions generally have a monogenic basis - “causative genes” n Complex disease is polygenic or multifactorial - “susceptibility genes”

24. Mendelian inheritance Autosomal dominant

25. Mendelian inheritance Autosomal recessive

26. Mendelian inheritance X-linked recessive

27. Mendelian disorders

28. Molecular biology methods n Nucleic acid collection n Nucleic acid hybridization n Restriction enzymes n Electrophoresis n Southern blots n Polymerase chain reaction n DNA sequencing

29. Nucleic acid collection Buccal swabs Blood spot on paper Tissue sample Blood sample

30. Nucleic acid hybridization The 2 DNA strands can be dissociated by heat or alkali Complementary strands will reanneal during cooling or under neutral conditions Reannealing is highly specific: only complementary sequences will anneal

31. Restriction enzymes ---GATTGAGGATCCTTATAT--- ---CTAACTCCTAGGAATATA--- ---GATTGAGGATCCTTATAT--- ---CTAACTCCTAGGAATATA--- ---GATTGAG GATCCTTATAT--- ---CTAACTCCTAG GAATATA--- EcoRI recognition site

32. Electrophoresis

33. Molecular biology methods (demos) Polymerase chain reaction Southern blot

34. DNA sequencing

35. DNA variation n Chromosomal loss, gain or rearrangements n Deletions n Insertions n Inversions n Point mutations/polymorphisms n Trinucleotide repeats (expansions) -Polymorphisms occur at more than 1% -Mutations occur at less than 1%

36. DNA variants can occur anywhere in the genome AAAAAA nuclear RNA transcription messenger RNA AAAAAA AUG TAG 5’ UTR 3’ UTR splicing * * * * * * *

37. DNA variants n Polymorphisms and mutations are the basis of genetic diversity n Polymorphisms provide markers with which we can track the inheritance of chromosomal regions and genes

38. DNA markers n Restriction fragment length polymorphisms (RFLPs) n Variable number tandem repeats (VNTR) n Microsatellites or short tandem repeats (STRs) n Single nucleotide polymorphism (SNPs)

39. Restriction fragment length polymorphisms (RFLPs) 164bp 527bp 691bp Exon 3 AvaII 164bp527bp Maximum of two alleles Of limited use for linkage Commonly used in association analysis Detected by blot or PCR

40. Variable number tandem repeats (VNTRs) Tandem repeats of between 14 and 100bp in length Multiple alleles Detected by blot or PCR Of limited application

41. Microsatellites or short tandem repeats (STRs) Detected by PCR Multiple alleles Widely used in linkage analysis Detected by PCR

42. Single nucleotide polymorphisms (SNPs)

44. Equipment

45. The genome era