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Practical exercises in Plant Biodiversity and Breeding (BI1103) Jonas RoosTom MartinLuisa Ghelardini
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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
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Arabidopsis thaliana Columbia (Col-0)Landsberg erecta (Ler)
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Petiole (short/long) Leaf (round/elongate)
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Large inflorescence Long siliques Col-0Ler inflorescence
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All of these features are controlled by a single gene: ERECTA (ER) - Your task is to find out where this gene is located.
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Recombinant Inbred Lines (RIL) Each RIL is a unique mix of Col-0 and Ler. 8 generations of inbreeding ⇒ homozygous at all loci
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- 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
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- 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
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Genotype - genetic markers
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Genetic map of Arabidopsis thaliana - Five chromosomes, about 50 markers Lister and Dean (1993) The Plant Journal 4:745-750
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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
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Indel (insertion/deletion) markers - Amplified with PCR - Identified on an agarose gel LerCol-0 m291
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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.
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