1. Practical exercises in Plant Biodiversity and Breeding (BI1103) Jonas RoosTom MartinLuisa Ghelardini

2. General aim of these labs: To use recombinant inbred lines (RIL) as an example of genetic variation, and to gain practical experience in working with plants. Specific aim: To map the ERECTA gene in Arabidopsis thaliana. a) Looking at plants ⇒ phenotype b) Looking at genetic markers ⇒ genotype c) Correlating genotype and phenotype ⇒ location of ERECTA gene

3. Arabidopsis thaliana Columbia (Col-0)Landsberg erecta (Ler)

4. Petiole (short/long) Leaf (round/elongate)

5. Large inflorescence Long siliques Col-0Ler inflorescence

6. All of these features are controlled by a single gene: ERECTA (ER) - Your task is to find out where this gene is located.

7. Recombinant Inbred Lines (RIL) Each RIL is a unique mix of Col-0 and Ler. 8 generations of inbreeding ⇒ homozygous at all loci

8. - Each RIL a unique mix of Col-0 and Ler DNA. - Correlate each genotype with the phenotype. - Quantitative Trait Locus (QTL) mapping. QTL mapping – correlate genotype and phenotype

9. - Plants will have either “Col-0” or “Ler” phenotype. - “Col-0” phenotype ⇒ Col-0 version of ERECTA gene - “Ler” phenotype ⇒ Ler version of ERECTA gene QTL mapping – correlate genotype and phenotype

10. Genotype - genetic markers

11. Genetic map of Arabidopsis thaliana - Five chromosomes, about 50 markers Lister and Dean (1993) The Plant Journal 4:745-750

12. Indel markers - The maker order and location is identical in Col-0 and Ler. - However, the markers themselves are different. - Can be either Col-0 version or Ler version. m291 Ler m291 Col

13. Indel (insertion/deletion) markers - Amplified with PCR - Identified on an agarose gel LerCol-0 m291

14. Summary 1.Plant out RIL, compare phenotype with Col-0 and Ler. - “Ler” phenotype ⇒ Ler version of ER - “Col-0” phenotype ⇒ Col-0 version of ER 2.Identify RIL genotype (indel markers) 3.Compare phenotype and genotype (QTL mapping) ⇒ location of ER.