Practical exercises in Plant Biodiversity and Breeding

(BI1103)

Jonas RoosTom Martin Luisa Ghelardini

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

Arabidopsis thaliana

Columbia (Col-0) Landsberg erecta (Ler)

Petiole (short/long)

Leaf (round/elongate)

Largeinflorescence

Longsiliques

Col-0 Ler

inflorescence

All of these features are controlled by a single gene: ERECTA (ER)

- Your task is to find out where this gene is located.

Recombinant Inbred Lines (RIL)

Each RIL is a unique mix of Col-0 and Ler.

8 generations of inbreeding ⇒ homozygous at all loci

- 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

- 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

Genotype- genetic markers

Genetic map of Arabidopsis thaliana

- Five chromosomes, about 50 markers

Lister and Dean (1993) The Plant Journal 4:745-750

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.

m291Ler

m291Col

Indel (insertion/deletion) markers

- Amplified with PCR- Identified on an agarose gel

LerCol-0

m291

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.⇒