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Hybrid genes in the analysis of transformation conditions
Authors:I. Negrutiu  R. Shillito  I. Potrykus  G. Biasini  F. Sala
Affiliation:(1) Friedrich Miescher-Institut, P.O. Box 2543, CH-4002 Basel, Switzerland;(2) Dipartimento di genetica e microbiologia, Università di Pavia, Via S. Epifanio, 14-27100 Pavia, Italy;(3) Present address: Instituut voor Moleculaire Biologie, 65 Paardenstraat, B-1640 Sint-Genesius-Rode, Belgium
Abstract:
Direct gene transfer into plant protoplasts has been recently developed, and conditions for high frequency transformation of SR1 tobacco protoplasts established. In this paper we analyse numerous transformation parameters in a comparative study on SR1Nicotiana tabacum and N. plumbaginifolia, and report on a simple chemical technique for very efficient protoplast transformation. It is based on the synergistic interaction of MgCl2 and PEG. The technique yielded up to 1400 transformants per 3×105 treated N. tabacum protoplasts (up to 4.8% of the survivors, late selected clones). Using N. plumbaginifolia, the frequencies were 10-fold lower, indicating that the lsquocompetencersquo for transformation has a species-specific component.Abbreviations PEG polyethyleneglycol - RTF relative transformation frequency - ATF absolute transformation frequency
Keywords:transformation  MgCl2-PEG-electroporation  competence  protoplasts
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