Cisgenic barley with improved phytase activity

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Cisgenic barley with improved phytase activity. / Holme, Inger; Dionisio, Giuseppe; Brinch-Pedersen, Henrik; Wendt, Toni; Madsen, Claus Krogh; Vincze, Éva; Holm, Preben Bach.

In: Plant Biotechnology Journal, Vol. 10, No. 2, 2012, p. 237-247.

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Holme, Inger ; Dionisio, Giuseppe ; Brinch-Pedersen, Henrik ; Wendt, Toni ; Madsen, Claus Krogh ; Vincze, Éva ; Holm, Preben Bach. / Cisgenic barley with improved phytase activity. In: Plant Biotechnology Journal. 2012 ; Vol. 10, No. 2. pp. 237-247.

Bibtex

@article{ff1dec64d57a4b959a890f505083a4ed,
title = "Cisgenic barley with improved phytase activity",
abstract = "The cisgenesis concept implies that plants are transformed only with their own genetic materials or genetic materials from closely related species capable of sexual hybridization. Furthermore, foreign sequences such as selection genes and vector-backbone sequences should be absent. We used a barley phytase gene (HvPAPhy_a) expressed during grain filling to evaluate the cisgenesis concept in barley. The marker gene elimination method was used to obtain marker-free plant lines. Here, the gene of interest and the selection gene are flanked by their own T-DNA borders to allow unlinked integration of the two genes. We analysed the transformants for co-transformation efficiency, increased phytase activities in the grain, integration of the kanamycin resistance gene of the vector-backbone and segregation between the HvPAPhy_a insert and the hygromycin resistance gene. The frequencies of the four parameters imply that it should be possible to select 11 potentially cisgenic T1-lines out of the 72 T0-lines obtained, indicating that the generation of cisgenic barley is possible at reasonable frequencies with present methods. We selected two potential cisgenic lines with a single extra copy of the HvPAPhy_a insert for further analysis. Seeds from plants homozygous for the insert showed 2.6- and 2.8-fold increases in phytase activities and the activity levels were stable over the three generations analysed. In one of the selected lines, the flanking sequences from both the left and right T-DNA borders were analysed. These sequences confirmed the absence of truncated vector-backbone sequences linked to the borders. The described line should therefore be classified as cisgenic.",
keywords = "Agrobacterium, barley, cisgenesis, HvPAPhy_a, grain phytase activity, marker-free transformants",
author = "Inger Holme and Giuseppe Dionisio and Henrik Brinch-Pedersen and Toni Wendt and Madsen, {Claus Krogh} and {\'E}va Vincze and Holm, {Preben Bach}",
year = "2012",
doi = "10.1111/j.1467-7652.2011.00660.x",
language = "English",
volume = "10",
pages = "237--247",
journal = "Plant Biotechnology Journal",
issn = "1467-7644",
publisher = "Wiley-Blackwell Publishing Ltd.",
number = "2",

}

RIS

TY - JOUR

T1 - Cisgenic barley with improved phytase activity

AU - Holme, Inger

AU - Dionisio, Giuseppe

AU - Brinch-Pedersen, Henrik

AU - Wendt, Toni

AU - Madsen, Claus Krogh

AU - Vincze, Éva

AU - Holm, Preben Bach

PY - 2012

Y1 - 2012

N2 - The cisgenesis concept implies that plants are transformed only with their own genetic materials or genetic materials from closely related species capable of sexual hybridization. Furthermore, foreign sequences such as selection genes and vector-backbone sequences should be absent. We used a barley phytase gene (HvPAPhy_a) expressed during grain filling to evaluate the cisgenesis concept in barley. The marker gene elimination method was used to obtain marker-free plant lines. Here, the gene of interest and the selection gene are flanked by their own T-DNA borders to allow unlinked integration of the two genes. We analysed the transformants for co-transformation efficiency, increased phytase activities in the grain, integration of the kanamycin resistance gene of the vector-backbone and segregation between the HvPAPhy_a insert and the hygromycin resistance gene. The frequencies of the four parameters imply that it should be possible to select 11 potentially cisgenic T1-lines out of the 72 T0-lines obtained, indicating that the generation of cisgenic barley is possible at reasonable frequencies with present methods. We selected two potential cisgenic lines with a single extra copy of the HvPAPhy_a insert for further analysis. Seeds from plants homozygous for the insert showed 2.6- and 2.8-fold increases in phytase activities and the activity levels were stable over the three generations analysed. In one of the selected lines, the flanking sequences from both the left and right T-DNA borders were analysed. These sequences confirmed the absence of truncated vector-backbone sequences linked to the borders. The described line should therefore be classified as cisgenic.

AB - The cisgenesis concept implies that plants are transformed only with their own genetic materials or genetic materials from closely related species capable of sexual hybridization. Furthermore, foreign sequences such as selection genes and vector-backbone sequences should be absent. We used a barley phytase gene (HvPAPhy_a) expressed during grain filling to evaluate the cisgenesis concept in barley. The marker gene elimination method was used to obtain marker-free plant lines. Here, the gene of interest and the selection gene are flanked by their own T-DNA borders to allow unlinked integration of the two genes. We analysed the transformants for co-transformation efficiency, increased phytase activities in the grain, integration of the kanamycin resistance gene of the vector-backbone and segregation between the HvPAPhy_a insert and the hygromycin resistance gene. The frequencies of the four parameters imply that it should be possible to select 11 potentially cisgenic T1-lines out of the 72 T0-lines obtained, indicating that the generation of cisgenic barley is possible at reasonable frequencies with present methods. We selected two potential cisgenic lines with a single extra copy of the HvPAPhy_a insert for further analysis. Seeds from plants homozygous for the insert showed 2.6- and 2.8-fold increases in phytase activities and the activity levels were stable over the three generations analysed. In one of the selected lines, the flanking sequences from both the left and right T-DNA borders were analysed. These sequences confirmed the absence of truncated vector-backbone sequences linked to the borders. The described line should therefore be classified as cisgenic.

KW - Agrobacterium

KW - barley

KW - cisgenesis

KW - HvPAPhy_a

KW - grain phytase activity

KW - marker-free transformants

U2 - 10.1111/j.1467-7652.2011.00660.x

DO - 10.1111/j.1467-7652.2011.00660.x

M3 - Journal article

C2 - 21955685

VL - 10

SP - 237

EP - 247

JO - Plant Biotechnology Journal

JF - Plant Biotechnology Journal

SN - 1467-7644

IS - 2

ER -