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A novel double T-DNA system for producing stack and marker-free transgenic plants

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Biologia Plantarum

Abstract

This study aimed to develop a new vector system to remove selection genes and to introduce two or more genes of interest into plants in order to express them in a coordinated manner. A multigene expression vector was established based on pCamBIA2300 using a selectable marker gene (SMG)-free system based on the combination of the isocaudamer technique and double T-DNA. The vector DT7, containing six target genes, was constructed and introduced into tobacco using Agrobacterium-mediated transformation. Twenty-one of 27 positive transgenic plants contained both T-DNA regions. The co-transformation frequency was 77.8 %. The frequency of unlinked integration of intact two T-DNAs was 22.22 % (6/27). The frequency of removal of SMG from transgenic T1 plants was 19.10 %. These results suggest that this vector system was functional and effective for multigene expression and SMG-free transgenic plant cultivation. At least six target genes can be co-expressed using this system. Overall, these findings provide a new and highly effective platform for multigene and marker-free transgenic plant production.

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Abbreviations

CF:

co-transformation frequency

GOI:

genes of interest

GUS:

β-glucuronidase

KanR :

kanamycin resistant

KanS :

kanamycin sensitive

LB:

left border

Nos P:

nopaline synthase promoter

Nos T:

Nos terminator

PPTR :

PPT-resistant plant

PPTS :

PPT-susceptible

RB:

right border

SMG:

selectable marker gene

ZFNs:

zinc finger nucleases

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Authors and Affiliations

  1. Biotechnology Research Institute, Chinese Academy of Agricultural Sciences, Beijing, 100081, P.R. China

    X. J. Wang, Y. Y. Su, Y. F. Dong, Q. L. Tang & Z. X. Wang

Authors
  1. X. J. Wang
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  2. Y. Y. Su
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  3. Y. F. Dong
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  4. Q. L. Tang
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  5. Z. X. Wang
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Correspondence to Z. X. Wang.

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Wang, X.J., Su, Y.Y., Dong, Y.F. et al. A novel double T-DNA system for producing stack and marker-free transgenic plants. Biol Plant (2016). https://doi.org/10.1007/s10535-016-0630-6

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  • DOI: https://doi.org/10.1007/s10535-016-0630-6

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