( ures of either 900 or 1300 psi changi...)Optimization of Biolistic Transformation Using the Helium-Driven PDS-1000/He System

ures of either 900 or 1,300 psi, changing the gap width or macrocarrier travel distance had very little effect on transient expression. These experiments indicate that for Biolistic transformation of cauliflower sections using the PDS-1000/He system, optimum transient expression is obtained when the sample is placed at level 2 (6 cm target distance) and bombarded at a helium pressure of 900 psi with either M17 tungsten or 1.6 mm gold microparticles.

Stable transformants can not be isolated from cauliflower tissue sections. However, these transient assays may be used as starting conditions for optimizing stable transformation. A rigorous experiment for resolving the optimal conditions for isolating stable transformants could be determined in a fractional factorial experiment as outlined in Table 4. Such an experiment should be undertaken only if experimental quantitation is relatively simple. A more practical approach would be to vary those parameters which have the greatest effect on transient expression, in this case helium pressure and target level. In optimizing for stable transformation, the most severe Biolistic conditions which are used should be the mildest conditions which produce a high level of transient expression (e.g., helium pressure of 900 psi and target at level 2). Optimization of stable transformation should use lower helium pressures and/or longer target distances than conditions for transient expression. (Also see comments in the Discussion section.) Since similar levels of GUS expression were obtained using M17 tungsten and 1.6 mm gold microparticles, it may be preferable to use gold microparticles for producing stable transformants due to the demonstrated toxicity of tung
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