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Selection of a Nicotiana plumbaginifolia universal hybridizer and its use in intergeneric somatic hybrid formation

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Summary

A Nicotiana plumbaginifolia cell strain carrying a positive (dominant) trait, resistance to azetidine-2-carboxylate (A2C), was selected in strain NX1 which lacked nitrate reductase activity (a negative or recessive trait). This universal hybridizer strain, denoted NXAr, was fused with dextran to a Daucus carota strain, PR, which carried glyphosate (GLP) resistance. A large number of hybrids were selected in a medium with NO -3 as the sole nitrogen source and A2C as inhibitor, conditions which prevent the growth of both parents. When the selected colonies were then tested for GLP resistance, 93% carried this trait. In addition the hybrid nature was indicated by additive chromosome numbers, both A2C and GLP resistance in suspension cultures, intermediate nitrate reductase activity and the presence of banding patterns for three isozymes which match those of the parents. Southern hybridization analysis using an enolpyruvylshikimic acid-3-phosphate synthase (EPSPS) probe, pMON 6145, also showed the presence of the gene from both parents in the hybrid strains based on restriction length polymorphisms. The PR strain contains increased levels of EPSPS which confers GLPr due to gene amplification. Since the universal hybridizer can be used as a fusion partner with any wild-type line many protoplast fusion studies can be carried out easily.

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Abbreviations

A2C:

azetidine-2-carboxylate

2,4-D:

2,4-dichlorophenoxyacetic acid

EPSPS:

5-enolpyruvylshikimic acid-3-phosphate synthase

GLP:

glyphosate

HAT:

hypoxanthine, aminopterin, glycine and thymidine medium

IDH:

isocitrate dehydrogenase

MDH:

malate dehydrogenase

5MT:

5-methyltryptophan

NBT:

nitroblue tetrazolium

PGI:

phosphoglucoisomerase

SDS:

sodium dodecylsulfate

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Author notes

  1. Jingsong Ye

    Present address: Department of Botany, Ohio State University, 1735 Neil Avenue, 43210, Columbus, OH, USA

  2. Randal M. Hauptmann

    Present address: Department of Botany, University of Florida, 220 Bartram Hall East, 32611, Gainesville, FL, USA

Authors and Affiliations

  1. Department of Agronomy, University of Illinois, 1102 South Goodwin Avenue, 61801, Urbana, IL, USA

    Jingsong Ye & Jack M. Widholm

  2. Monsanto, 700 Chesterfield Village Parkway, 63198, Chesterfield, MO, USA

    Randal M. Hauptmann & Alan G. Smith

Authors
  1. Jingsong Ye
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  2. Randal M. Hauptmann
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  3. Alan G. Smith
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  4. Jack M. Widholm
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Additional information

Communicated by G. Melchers

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Ye, J., Hauptmann, R.M., Smith, A.G. et al. Selection of a Nicotiana plumbaginifolia universal hybridizer and its use in intergeneric somatic hybrid formation. Mol Gen Genet 208, 474–480 (1987). https://doi.org/10.1007/BF00328142

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  • DOI: https://doi.org/10.1007/BF00328142

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