In vitro patterning and differentiation of xylem elements in Jerusalem artichoke explants.
Week One

1. Trim Jeruslem artichoke tuber
2. Surface sterilise with sodium hypochlorite solution
3. Transfer under sterile “tent”
4. Wash 3x with sterile water
5. Trim top and bottom of tuber piece with large scalpel on sterile petri dish lid
6. Take cores of pith
7. Slice cores into 1mm explants & transfer to small beaker of sterile water
8. Wash twice with sterile water
9. Transfer to plates: weigh plate immediately before and after addition of explants to obtain fresh weight.
10. Wrap in aluminium foil - incubate for 1 week at 25C
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Week Two

1. Inspect tissue discs and note any differences
2. Weigh the tissue discs in plastic weigh boats
3. Incubate in 5% NaOH solution at 80C for 1 hour
4. Wash several times with water
5. Incubate in 0.03% aqueous safranin O for 0.5 hour at 55C
6. Destain in several changes of 1M HCl over the period of an hour
7. Inspect pattern of differentiated cells within intact discs
8. Transfer discs to small beakers and sonicate
9. Count differentiated cells in haemocytometer slide
10. Calculate numbers of differentiated cells (per g fresh weight) for the different treatments
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