Garlic Sulphur Biochemistry Workpackage Four Garlic Sulphur Biochemistry Partner 2: Horticulture Research International Laurence Trueman, Brian Thomas, Linda Brown & Brian Smith

To identify developmental control points for CSO synthesis Objective: 2 To identify developmental control points for CSO synthesis Milestones Feb. 2002 - 2003 Analysis of second-year field experiment completed Whole plant labelling studies completed

Hydroponic Printanor A=50% total sulphur (d. 102, end March) B=Start bulbing (d. 130, 20 Apr) C=Start of senescence of root & leaf (d. 170, end May) D=Harvest (d. 230, end Jul) A B C D

Hydroponic Printanor Distribution and remobilization of sulphur taken up early * * * * * * * * * * * every 14d Distribution and remobilization of sulphur taken up late * * * * * * * * * * * every 14d A B C D 34S 32S

What we want to know When does nutrient uptake from the medium stop When does bulb filling start What proportion of S (and N) comes from the roots, leaves and hydroponic medium The form of the S in the root and leaf

2002 - Hydroponic data Bulb Index Fresh weight Dry Weight

To isolate and characterise alliinase cDNA clones Objective: 3b To isolate and characterise alliinase cDNA clones Milestones Feb. 2002 - 2003 Start Expression Analysis

Garlic Alliinase Expression relative to total rRNA content

Comparison of DNA derived polypeptide sequences Peptide 1 = Published, 2 = Al48 & 3 = Al41

Comparison of Peptide sequences Al48 has 98.9% identity to other known garlic cDNA’s Al41 has only the N-terminal 50% of the peptide in common with other alliinases The N-terminal portion of this peptide is likely to direct this peptide to the vacuole - Does it act as a storage protein?

Is it real? Al48 Al41

Clustering relationship of Messidrome alliinase PCR fragment sequences l=Leaf Clone b=Bulb Clone

mRNA Expression - Method