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Improvements in the transformation of Arabidopsis thaliana C24 leaf-discs by Agrobacterium tumefaciens

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Summary

We report here an efficient Arabidopsis leafdisc transformation protocol yielding an average transformation frequency of 1.6 transgenic shoots per leaf explant 4 weeks after the bacterial infection period. Subsequent cultivation in vitro is such that a high percentage (85–90%) of the primary transformants produces seeds with an average seed yield of 100–300 seeds per plant. This improved transformation protocol yields mainly (70%) transformants segregating for a single T-DNA locus of which 68% actually contain one T-DNA insert. The objective is to generate a pool of independent transformants harboring an activator T-DNA construct in a gene tagging approach to isolate genes involved in morphogenesis and auxin signal transduction.

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Abbreviations

2,4-D :

2,4-dichlorophenoxyacetic acid

AGM :

Arabidopsis growth medium

BAP :

benzylaminopurine

CaMV :

Cauliflower Mosaic Virus

CTAB :

Hexadecyltrimethylammoniumbromide

DIG :

digoxigenin

FeNaEDTA :

Iron-sodium-ethylenedinitrilo tetraacetic acid complex

GUS :

ß-Glucuronidase

IBA :

indole-3-butyric acid

LB :

left T-DNA border

MES :

2-(N-morpholino) ethane sulfonic acid

MS :

Murashige and Skoog medium

NAA :

α-naphthaleneacetic acid

RB :

right T-DNA border

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Authors and Affiliations

  1. Institute of Molecular Plant Sciences, Leiden University, Clusius Laboratory, Wassenaarseweg 64, 2333, AL Leiden, The Netherlands

    Eric van der Graaff & Paul J. J. Hooykaas

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  1. Eric van der Graaff
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  2. Paul J. J. Hooykaas
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Communicated by H. Lörz

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van der Graaff, E., Hooykaas, P.J.J. Improvements in the transformation of Arabidopsis thaliana C24 leaf-discs by Agrobacterium tumefaciens . Plant Cell Reports 15, 572–577 (1996). https://doi.org/10.1007/BF00232455

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  • DOI: https://doi.org/10.1007/BF00232455

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