1. Copyright © The McGraw-Hill Companies, Inc. Permission required to reproduce or display B-1 PowerPoint to accompany Genetics: From Genes to Genomes Fourth Edition Hartwell ● Hood ● Goldberg ● Reynolds ● Silver Reference B Prepared by Malcolm Schug University of North Carolina Greensboro

2. B-2 Copyright © The McGraw-Hill Companies, Inc. Permission required to reproduce or display Arabadopsis thaliana: Genetic Portrait of a Model Plant

3. B-3 Copyright © The McGraw-Hill Companies, Inc. Permission required to reproduce or display Fig. B.1 Fig. B.2 A. thaliana plant Hybrid camellia vs. wild-type

4. B-4 Copyright © The McGraw-Hill Companies, Inc. Permission required to reproduce or display Outline of Reference B Structure and organization of the genome Structure and organization of the genome Plant’s anatomy and life cycle Plant’s anatomy and life cycle Techniques of mutational analysis Techniques of mutational analysis Chemical and radiation procedures Chemical and radiation procedures Insertional mutagenesis Insertional mutagenesis Analysis of mutations to identify gene function Analysis of mutations to identify gene function Genetic analysis applied to various aspects of development Genetic analysis applied to various aspects of development Embryogenesis Embryogenesis Hormonal control systems Hormonal control systems Responses to environmental signals Responses to environmental signals Genetic analysis of flowering: a comprehensive example Genetic analysis of flowering: a comprehensive example

5. B-5 Copyright © The McGraw-Hill Companies, Inc. Permission required to reproduce or display Genome Structure and Organization Genome size: 125 Mb Genome size: 125 Mb Smallest genomes known in plant kingdom Smallest genomes known in plant kingdom Five pairs of small chromosomes Five pairs of small chromosomes Well-defined banding patterns on chromosomes Well-defined banding patterns on chromosomes Complete sequence of genome in 2000 Complete sequence of genome in 2000

6. B-6 Copyright © The McGraw-Hill Companies, Inc. Permission required to reproduce or display Comparison of Genetic and Physical Maps Ecotypes – plant varieties analogous to animal strains with common origin, shape and phenotypes Ecotypes – plant varieties analogous to animal strains with common origin, shape and phenotypes Columbia (Col) Columbia (Col) Landsberg erecta (Ler) Landsberg erecta (Ler) Differ by DNA polymorphisms and phenotypes Differ by DNA polymorphisms and phenotypes Segregation patterns of self-pollinating plants from crosses between ecotypes Segregation patterns of self-pollinating plants from crosses between ecotypes Linkage between DNA markers themselves Linkage between DNA markers themselves Linkage between DNA markers and morphological loci Linkage between DNA markers and morphological loci Create maps of DNA markers and phenotypes and integrate them Create maps of DNA markers and phenotypes and integrate them

7. B-7 Copyright © The McGraw-Hill Companies, Inc. Permission required to reproduce or display Characteristics of A.thaliana Genome Genome has little repetitive DNA and tight arrangement of genes Genome has little repetitive DNA and tight arrangement of genes Protein coding genes 40% of genome Protein coding genes 40% of genome One gene every 4 kb One gene every 4 kb 65% of proteins encoded by > 1 gene 65% of proteins encoded by > 1 gene Derived from ancient tetraploid Derived from ancient tetraploid Multiple copies of transposable elements Multiple copies of transposable elements DNA that originated in mitochondria and chloroplasts DNA that originated in mitochondria and chloroplasts Centromeres are heterochromatic regions Centromeres are heterochromatic regions Genetic and physical distances vary widely Genetic and physical distances vary widely

8. B-8 Copyright © The McGraw-Hill Companies, Inc. Permission required to reproduce or display Functional Analysis of Arabadobsis Genes Figure B.4

9. B-9 Copyright © The McGraw-Hill Companies, Inc. Permission required to reproduce or display The Basic Body Plan Fig. B.5

10. B-10 Copyright © The McGraw-Hill Companies, Inc. Permission required to reproduce or display Life Cycle from Fertilization to Flowering to Senescence Each ovule has six mononucleate cells Each ovule has six mononucleate cells One nucleus and two nuclei One nucleus and two nuclei Pollen triggers fertilization by landing on stigma Pollen triggers fertilization by landing on stigma Fig. B.6 a,b

11. B-11 Copyright © The McGraw-Hill Companies, Inc. Permission required to reproduce or display Fertilization Fig. B.6 c

12. B-12 Copyright © The McGraw-Hill Companies, Inc. Permission required to reproduce or display Stages of Embryonic Development Figure B.7 Insert figure B.7 from Hartwell 3e as shown on slide pages B12 and B13.

13. B-13 Copyright © The McGraw-Hill Companies, Inc. Permission required to reproduce or display From heart stage Final Stages of Embryonic Development

14. B-14 Copyright © The McGraw-Hill Companies, Inc. Permission required to reproduce or display Favorable Environmental Conditions Trigger Seed Germination and Vegetative Growth Gravity sensed by root and shoot Gravity sensed by root and shoot Light hypocotyl develops rapidly in dark and apical meristem and cotyledon growth suppressed Light hypocotyl develops rapidly in dark and apical meristem and cotyledon growth suppressed photomorphogenesis photomorphogenesis Fig. B.8

15. B-15 Copyright © The McGraw-Hill Companies, Inc. Permission required to reproduce or display Reproductive Development Begins when Leaf-producing Apical Meristem Switches to Flower-producing Apical Meristem Environmental signals Environmental signals Photoperiod – day length Photoperiod – day length Vernalization – exposing to cold Vernalization – exposing to cold Scenescence – vegetative plant ages and dies Scenescence – vegetative plant ages and dies Fig. B.9

16. B-16 Copyright © The McGraw-Hill Companies, Inc. Permission required to reproduce or display Mutagenesis by Chemical and Irradiation Produces Different Ratios of Various Mutations Seed contains dormant embryo from which 1-3 cells from apical meristem are destined to form the germ line Seed contains dormant embryo from which 1-3 cells from apical meristem are destined to form the germ line Mutational segregation depends on how many gamete cells are present during mutagenesis Mutational segregation depends on how many gamete cells are present during mutagenesis One cell – ¼ progeny have mutation One cell – ¼ progeny have mutation Two cells – 1/8 progeny Two cells – 1/8 progeny Three cells – 1/12 progeny Three cells – 1/12 progeny

17. B-17 Copyright © The McGraw-Hill Companies, Inc. Permission required to reproduce or display Transformation by T-DNA Bacterium Agrobacterium tumefaciens is agent of transformation Bacterium Agrobacterium tumefaciens is agent of transformation Transfer of plasmid DNA called T-DNA into genome of wounded plant Transfer of plasmid DNA called T-DNA into genome of wounded plant Antibiotic resistance engineered into plasmids provide selectable markers Antibiotic resistance engineered into plasmids provide selectable markers GUS reporter gene with no promoter stain blue when inserted in gene of plant GUS reporter gene with no promoter stain blue when inserted in gene of plant Fig. B.10 a

18. B-18 Copyright © The McGraw-Hill Companies, Inc. Permission required to reproduce or display Transformation by Transposon Tagging Transposon tagging using transposable elements from Corn and Arabidobsis Transposon tagging using transposable elements from Corn and Arabidobsis Insertional mutagenesis not only allows generation of mutations but also facilitates molecular characterization Insertional mutagenesis not only allows generation of mutations but also facilitates molecular characterization Transposon or T-DNA can be used as a probe to identify and clone gene insert disrupts Transposon or T-DNA can be used as a probe to identify and clone gene insert disrupts

19. B-19 Copyright © The McGraw-Hill Companies, Inc. Permission required to reproduce or display Silencing Specific Genes through Antisense RNAs or RNAs Carrying Inverted Duplications of Gene Fragments Activation of defense mechanism evolved to protect plants from invading viruses or transposons Activation of defense mechanism evolved to protect plants from invading viruses or transposons Double-stranded RNAs are targeted by ribonuclease (Dicer) and cleaved into 21 bp fragments Double-stranded RNAs are targeted by ribonuclease (Dicer) and cleaved into 21 bp fragments Fragments of dsRNA recognize homologous molecules in cell and promote degradation Fragments of dsRNA recognize homologous molecules in cell and promote degradation Gene knockout Gene knockout Induction of recombinant gene that carries inverted repeat of gene of interest (endogene) Induction of recombinant gene that carries inverted repeat of gene of interest (endogene) RNA forms hairpin loop and is degraded by Dicer RNA forms hairpin loop and is degraded by Dicer

20. B-20 Copyright © The McGraw-Hill Companies, Inc. Permission required to reproduce or display Fig. B.11

21. B-21 Copyright © The McGraw-Hill Companies, Inc. Permission required to reproduce or display Genetic Analysis of Embryogenesis Researcher have found very few maternal-effect mutations in Arabadopsis Researcher have found very few maternal-effect mutations in Arabadopsis Occur in maternal genes whose products are deposited in egg and alter embryogenesis Occur in maternal genes whose products are deposited in egg and alter embryogenesis Major difference between plants and certain animals such as Drosophila Major difference between plants and certain animals such as Drosophila Lack of maternal-effect mutants is mostly due to cytoplasm that is produced by zygote’s own genome Lack of maternal-effect mutants is mostly due to cytoplasm that is produced by zygote’s own genome Few zygotic mutations affect embryogenesis before globular stage Few zygotic mutations affect embryogenesis before globular stage Zygotic genes encode proteins that are functionally redundant with those of maternal origin Zygotic genes encode proteins that are functionally redundant with those of maternal origin Many genes are members of multigene families Many genes are members of multigene families

22. B-22 Copyright © The McGraw-Hill Companies, Inc. Permission required to reproduce or display Screens for Mutations that Arrest Development at Specific Stages of Embryogenesis Help Identify Regulatory Processes (a) LEC genes help regulate seed maturation (a) LEC genes help regulate seed maturation LEC2 encodes transcription factor controlling differentiating cells into embryo LEC2 encodes transcription factor controlling differentiating cells into embryo (b) TWIN represses embryo development program in suspensor cells (b) TWIN represses embryo development program in suspensor cells Figure B.12 a,b

23. B-23 Copyright © The McGraw-Hill Companies, Inc. Permission required to reproduce or display gal-1 is an x-ray induced deletion that helped identify it as gene in gibberellin biosynthetic pathway gal-1 is an x-ray induced deletion that helped identify it as gene in gibberellin biosynthetic pathway Mutations that Disrupt Hormone Activity Clarify Biological Significance and Biosynthetic Pathway Fig. B.13

24. B-24 Copyright © The McGraw-Hill Companies, Inc. Permission required to reproduce or display Mutations that Render Arabadopsis Insensitive to a Hormone Reveal how Plants Perceive and Transduce Hormone Signals Ethylene exposure inhibits shoot and root elongation and accentuation of apical hook Ethylene exposure inhibits shoot and root elongation and accentuation of apical hook Mutant seedlings grow tall in presence of ethylene Mutant seedlings grow tall in presence of ethylene Figure 14

25. B-25 Copyright © The McGraw-Hill Companies, Inc. Permission required to reproduce or display Ethylene Biosynthetic Pathway Fig. B. 15

26. B-26 Copyright © The McGraw-Hill Companies, Inc. Permission required to reproduce or display Mutational Analysis has Helped Characterize the Photoreceptor Molecules by Which Plants Receive Light Signals hy (hypocotyle) mutants show reduced sensitivity to light hy (hypocotyle) mutants show reduced sensitivity to light hy3 and hy8 coincide with phyA and phyB on linkage maps – members of phytochrome gene family hy3 and hy8 coincide with phyA and phyB on linkage maps – members of phytochrome gene family hy3 altered response to red wavelengths hy3 altered response to red wavelengths hy8 altered response to far-red wavelengths hy8 altered response to far-red wavelengths Same phytochrome family Same phytochrome family hy4 decreased sensitivity to blue light hy4 decreased sensitivity to blue light hy5 encodes transcription factor controls expression of genes contributing to morphogenesis hy5 encodes transcription factor controls expression of genes contributing to morphogenesis

27. B-27 Copyright © The McGraw-Hill Companies, Inc. Permission required to reproduce or display Mutations that Affect Response to Light Fig. B.16 a

28. B-28 Copyright © The McGraw-Hill Companies, Inc. Permission required to reproduce or display Mutational Studies of How Plants Process Light Signals cop (constitutive photomorphogenesis) cop (constitutive photomorphogenesis) det (de-etiolated – released from development without chlorophyll) det (de-etiolated – released from development without chlorophyll) Grow in dark as if they received light Grow in dark as if they received light Recessive Recessive COPI is negative regulator, mediating degradation of transcription factors needed for photomorphogenesis COPI is negative regulator, mediating degradation of transcription factors needed for photomorphogenesis COPI also interacts directly with cytochrome blue-light photoreceptors COPI also interacts directly with cytochrome blue-light photoreceptors det2 mutant shows degradation of light-regulating genes, but does not develop chloroplasts – dwarf adult det2 mutant shows degradation of light-regulating genes, but does not develop chloroplasts – dwarf adult

29. B-29 Copyright © The McGraw-Hill Companies, Inc. Permission required to reproduce or display Genetic Analysis of Flowering: A Comprehensive Example Switch to reproductive growth (flowering) involves reprogramming apical meristem Switch to reproductive growth (flowering) involves reprogramming apical meristem Becomes inflorescence meristem (IM) Becomes inflorescence meristem (IM) Produces smaller leaves, elongated stem, and many side shoots Produces smaller leaves, elongated stem, and many side shoots IM is indeterminate because it produces floral meristems (FMs) indefinitely IM is indeterminate because it produces floral meristems (FMs) indefinitely FMs are determinant because they produce a fixed number of floral organs FMs are determinant because they produce a fixed number of floral organs

30. B-30 Copyright © The McGraw-Hill Companies, Inc. Permission required to reproduce or display Fig. B.17

31. B-31 Copyright © The McGraw-Hill Companies, Inc. Permission required to reproduce or display Analysis of Homeotic Mutations Reveal Three Types of Single-gene Products Influencing Floral Pattern Formation Homeotic gene – plays a role in determining a tissue’s identity during development Homeotic gene – plays a role in determining a tissue’s identity during development Homeotic mutation – cells misinterpret their position and become normal organs in inappropriate tissues Homeotic mutation – cells misinterpret their position and become normal organs in inappropriate tissues Three classes of mutations Three classes of mutations Class A – carpels instead of sepals in first whorl and stamens instead of petals in second whorl Class A – carpels instead of sepals in first whorl and stamens instead of petals in second whorl Class B – sepals in first and second whorl, and carpels in third and fourth whorls Class B – sepals in first and second whorl, and carpels in third and fourth whorls Class C – abnormal radial pattern of sepals, petals, petals, sepals Class C – abnormal radial pattern of sepals, petals, petals, sepals

32. B-32 Copyright © The McGraw-Hill Companies, Inc. Permission required to reproduce or display Fig. B.18 a Insert B.18a again – this image blurry

33. B-33 Copyright © The McGraw-Hill Companies, Inc. Permission required to reproduce or display How Three Classes of Genes Could Determine Identity of Floral Organs Fig. B.18

34. B-34 Copyright © The McGraw-Hill Companies, Inc. Permission required to reproduce or display c) ap2 / ap3, ap2 / ag, and ap3 /ag double mutants ap2/ap3/ag triple mutant AP3 activity Fig. B.18 How Three Classes of Genes Could Determine Identity of Floral Organs

35. B-35 Copyright © The McGraw-Hill Companies, Inc. Permission required to reproduce or display Early Acting Genes Specify Identity of Floral Meristem FM produces flower primordium that differentiates into four whorls of organs, two of which contain gametes FM produces flower primordium that differentiates into four whorls of organs, two of which contain gametes Loss of function single and double mutant analysis Loss of function single and double mutant analysis Gain of function transgenic plants Gain of function transgenic plants Fig. B.19

36. B-36 Copyright © The McGraw-Hill Companies, Inc. Permission required to reproduce or display

37. B-37 Copyright © The McGraw-Hill Companies, Inc. Permission required to reproduce or display Some Genes Control Timing of FM Formation and Flowering

38. B-38 Copyright © The McGraw-Hill Companies, Inc. Permission required to reproduce or display Hundreds of Mutational Snalyses have Generated Preliminary Model-to-Guide Future Research on Flowering  a Preliminary Genetic Model of Process Fig. B.20