Table 2. Comparison of the cloning procedures of transgenic tobacco BY-2 cells.

Cloning procedure	Nocarova and Fischer, 2009	This paper
Transformation method	Agrobacterium tumefaciens	Agrobacterium tumefaciens LB4404
culture preparation from primary calli	1 ml of fresh calli in 30 ml medium (+AB) by pipeting	small calli (diameter ~5 mm) in 10 ml medium (+AB) with a spatula
subculturing of primary suspensions	after 7 days: 1,5 ml in 30 ml medium	after 7 days: 1–2 ml in 40 ml medium
cloning of secondary calli	7-day-old transgenic cells are diluted 1:3 and mixed with 4ml of similarly prepared WT cells in a ratio 1:1000	7-day-old WT cells are diluted 1:10 (3 ml + 27 ml of medium) and mixed with 7 day old transgenic cells (60 µl) in a ratio 1:500
dilution factors and ratios	7-day-old wild-type: 1:3 7-day-old transgenic cells: 1:3 transgenic:wild-type: 1:1000	7-day-old wild-type: 1:10 7-day-old transgenic cells: - transgenic:wild-type: 1:500
Spread on solid medium (+AB)	500 µl on Ø 6 cm Petri dish	7 ml on Ø 12 cm Petri dish (sufficient to cover the surface)
calli appearing (after 3–6 weeks)	~25	~25 – 100