Roses

Summary

Florigene’s inventions disclosed in two granted United States patents, a PCT application and their national phase entries are directed to:

  • a method for producing a transformed somatic rose embryo, which expresses an exogenous gene,
  • a method for transforming embryogenic, friable, granular rose callus cells with Agrobacterium,
  • a method for transforming a rose plant and producing transformed rose plantlets, and
  • culture media for callus cultivation and somatic embryo maintenance.

An additional protocol is disclosed to obtain somatic embryos out of a mature somatic tissue, a stamen filament and a leaf explant. These protocols are not limited to transformed tissues.

A new United States patent application has been filed by Florigene. Unlike the granted United States patents, the application describes Agrobacterium-mediated transformation of rose callus cells, without specifying the origin of the callus.

Florigene was founded as a joint venture between DNA Plant Technology Co. (now called S&G Seeds) and Rabobank Biotech Venture Fund, the former also listed as an assignee in the PCT application.

Roses – Patents granted to Florigene Europe B.V.

Specific Patent Information

Patent Number Title, Independent Claims and Summary of Claims Assignee
US 5480789

  • Earliest priority – 1 April 1991
  • Filed – 18 November 1993
  • Granted – 2 January 1996
  • Expected expiry – 2 January 2013
Title – Genetically transformed rose plants and methods for their production

Claim 1
A method for producing a somatic rose embryo, which expresses an exogenous DNA sequence including a selectable marker gene, said method comprising:

A) culturing tissue from a rose plant on a callus induction medium containing nutrients, an energy source, an auxin, and a cytokinin in amounts effective to induce formation of embryogenic callus, wherein the tissue is cultured until a friable, granular embryogenic callus is produced;
B) combining cells from the embryogenic callus of step (A) with Agrobacterium cells carrying the exogenous DNA sequence in a co-cultivation medium containing nutrients, an energy source, and an induction compound under conditions which allow the Agrobacterium cells to infect the embryogenic callus cells and transfer the exogenous DNA sequence to the embryogenic callus cell chromosomes;
C) culturing embryogenic callus cells from step (B) in a selection medium containing nutrients, an energy source, an auxin, a cytokinin, and an agent which inhibits the growth of embryogenic callus cells which do not express the selectable marker gene; and
D) culturing the cells selected in step (C) in a maintenance medium containing nutrients, an energy source, an antibacterial agent, and a growth regulator, other than an auxin or a cytokinin, present in amounts effective to produce viable somatic embryos capable of being regenerated into transformed plantlets.

Granted US 5480789 is a continuation of abandoned US 07/678846.

Method for the production of a transformed somatic rose embryo expressing a gene of interest by transforming an embryogenic callus withAgrobacterium carrying the exogenous gene.

Florigene Europe B.V.

US 5792927

  • Earliest priority – 1 April 1991
  • Filed – 5 June 1995
  • Granted – 11 August 1998
  • Expected expiry – 2 January 2013
Title – Genetically transformed rose plants and methods for their production

Claim 1
A method for genetically transforming callus cells from a rose plant, said method comprising: A) incubating friable, granular callus cells withAgrobacterium cells carrying an exogenous DNA sequence; and
B) selecting callus cells which express at least a portion of the exogenous DNA sequence.
Claim 5
A method for genetically transforming a rose plant, said method comprising: A) culturing tissue from the rose plant under conditions selected to produce a friable, granular callus;
B) incubating cells from the callus of step (A) with Agrobacterium cells carrying an exogenous DNA sequence;
C) selecting callus cells from step (B) which express at least a portion of the DNA sequence; and
D) producing transformed plantlets from the selected callus cells of step (C).
Claim 15A somatic rose embryo produced by the method comprising: A) culturing tissue from a rose plant on a callus induction medium containing nutrients, an energy source, an auxin, and a cytokinin in amounts effective to induce callus formation;
B) combining cells from the callus of step (A) with Agrobacterium cells carrying the exogenous DNA sequence in a co-cultivation medium containing nutrients, an energy source, and an induction compound under conditions which allow the Agrobacterium cells to infect the callus cells and transfer the exogenous DNA sequence to the callus cell chromosomes;
C) culturing callus cells from step (B) in a selection medium containing nutrients, an energy source, an auxin, a cytokinin, and an agent which inhibits the growth of callus cells which do not express the selectable marker gene; and
D) culturing the cells selected in step (C) in a maintenance medium containing nutrients, an energy source, an antibacterial agent, and a growth regulator, other than an auxin or a cytokinin, present in amounts effective to produce somatic embryos.

Granted US 5792927 is a divisional of now granted US 5480789 (see above).

Method for transforming rose callus cells with Agrobacterium carrying an exogenous gene and selecting callus cells containing that exogenous gene. Production of a transformed somatic rose embryo and a method for transforming a rose plant starting from a transformed granular callus are part of the claimed invention.

Note: Patent information on this page was last updated on 22 March 2006.

Roses – Patent application filed by Florigene B.V.

Specific Patent Information

Patent Number Title, Independent Claims and Summary of Claims Assignee
WO 1992/00371 A1

  • Earliest priority – 1 April 1991
  • Filed – 21 June 1991
  • OPI – 9 January 1992
Title – Rose plants and methods for their production and transformation

Claim 1

A method for controlled regeneration of a rose plantlet from a somatic embryo which comprises:

(a) providing a somatic embryo;
(b) culturing the somatic embryo on a maturation medium capable of inducing differentiation of the embryo to yield a differentiated embryo;
(c) germinating the differentiated embryo on germination medium to yield a germinated embryo; and
(d) propagating the germinated embryo on propagation medium to produce a mature plantlet capable of being transferred to soil conditions.

Claim 8

A method for obtaining at least one somatic embryo from mature somatic tissue of rose plant which comprises:

(a) culturing mature somatic tissue on callus induction medium comprising effective amounts of a nutrient medium, an energy source, an auxin and a cytokinin to obtain at least one induced callus; and
(b) culturing the induced callus in a regeneration media capable of inducing completion of the development of somatic embryos comprising effective amounts of a nutrient medium, an energy source, an auxin and a cytokinin in which the source of the auxin and cytokinin in the regeneration media differs from the source of the auxin and cytokinin in the callus induction medium to obtain at least one somatic embryo.

Claim 17

A method for obtaining a somatic embryo from a stamen filament of rose plant which comprises:

(a) culturing the stamen filament on callus induction medium comprising effective amounts of a nutrient medium, an energy source, an auxin and a cytokinin to obtain at least one induced callus; and
(b) culturing the induced callus in a regeneration media capable of inducing completion of the development of somatic embryos comprising effective amounts of a nutrient medium, an energy source, an auxin and a cytokinin in which the ratio of auxin to cytokinin is decreased by a factor of two to about 15 relative to the ratio of auxin to cytokinin in the callus induction medium to obtain a somatic embryo.

Claim 20

A method for obtaining a somatic embryo from a leaf explant of rose plant which comprises:

(a) culturing the leaf explant on a callus induction medium comprising effective amounts of a nutrient medium, an energy source, an auxin and a cytokinin to obtain at least one induced callus; and
(b) culturing the induced callus in a regeneration media capable of inducing completion of the development of somatic embryos comprising effective amounts of a nutrient medium, an energy source, an auxin and a cytokinin to which the source of the auxin and cytokinin in the regeneration media differs from the source of the auxin and cytokinin in the callus induction medium to obtain a somatic embryo.

Claim 26
A method for genetically transforming callus cells from a rose plant, said method comprising:

  • incubating the callus cells with Agrobacterium cells carrying an exogenous DNA sequence; and
  • selecting callus cells which express at least a portion of the exogenous DNA sequence.
Claim 29
A method for genetically transforming a rose plant, said method comprising:

(a) culturing tissue from the rose plant under conditions selected to produce a callus;
(b) incubating cells from the callus of step (a) with Agrobacterium cells carrying an exogenous DNA sequence;
(c) selecting callus cells from step (b) which express at least a portion of the DNA sequence; and
(d) producing transformed plantlets from the selected callus cells of step (c).

Claim 34
A method for producing a somatic rose embryo which expresses an exogenous DNA sequence, said method comprising:

(a) culturing tissue from a rose plant on a callus induction medium containing nutrients, an energy source, an auxin, and growth regulator, a cytokinin in amounts effective to induce callus formation wherein the tissue is derived from a plant part selected from the group consisting of stamen filaments, leaf explants, stem sections, shoot tips, petal, sepal, petiole, and peduncle;
(b) combining cells from the callus of step (a) with Agrobacterium cells carrying the exogenous DNA sequence in a cocultivation medium containing nutrients, an energy source, and an induction compound under conditions which allow the Agrobacterium cells to infect the callus cells and transfer the exogenous DNA sequence to the callus cell chromosomes;;
(c) culturing callus cells from step (b) in a selection medium containing nutrients, an energy source, an auxin, a cytokinin, and an agent which inhibits the growth of callus cells which do not express the selectable marker gene; and
(d) culturing the cells selected in step (c) in a regeneration medium containing nutrients, an energy source, an antibacterial agent, and a growth regulator selected from abscisic acid and giberellic acid, other than an auxin or a cytokinin, present in amounts effective to produce somatic embryos.

Claim 50
A rose callus cell which expresses an exogenous DNA sequence.
Claim 51
A rose plant having cells which express an exogenous DNA sequence.
Claim 52
A somatic rose embryo which expresses an exogenous DNA sequence.

The PCT application has DNA Plant Technology Co. as a co-assignee listed with Florigene B.V..

The claims of the PCT application recite methods to regenerate a rose plantlet from a somatic embryo and to obtain a somatic embryo from somatic rose tissue, i.e. stamen filament, and leaf. Similar to its related United States patent, methods are also claimed to transform a rose callus with Agrobacterium having an exogenous gene, to produce a transformed somatic embryo and transformed rose plantlets.

Florigene B.V.
US 2001/007157 A1

  • Earliest priority – 18 November 1993
  • Filed – 10 August 1998
  • Abandoned – 15 July 2002
Title – Genetically transformed rose plants and methods for their production

Claim 1
A method for genetically transforming callus cells from a rose plant, said method comprising:

  • incubating the callus cells with Agrobacterium cells carrying an exogenous DNA sequence; and
  • selecting callus cells which express at least a portion of the exogenous DNA sequence.
Claim 6
A method for genetically transforming a rose plant, said method comprising:

(a) culturing tissue from the rose plant under conditions selected to produce a callus;
(b) incubating cells from the callus of step (a) with Agrobacterium cells carrying an exogenous DNA sequence;
(c) selecting callus cells from step (b) which express at least a portion of the DNA sequence; and
(d) producing transformed plantlets from the selected callus cells of step (c).

Claim 13
A method for producing a somatic rose embryo which expresses an exogenous DNA sequence including a selectable marker gene, said method comprising:

(a) culturing tissue from a rose plant on a callus induction medium containing nutrients, an energy source, an auxin, and a cytokinin in amounts effective to induce callus formation;
(b) combining cells from the callus of step (a) with Agrobacterium cells carrying the exogenous DNA sequence in a cocultivation medium containing nutrients, an energy source, and an induction compound under conditions which allow the Agrobacterium cells to infect the callus cells and transfer the exogenous DNA sequence to the callus cell chromosomes;
(c) culturing callus cells from step (b) in a selection medium containing nutrients, an energy source, an auxin, a cytokinin, and an agent which inhibits the growth of callus cells which do not express the selectable marker gene; and
(d) culturing the cells selected in step (c) in a maintenance medium containing nutrients, an energy source, an antibacterial agent, and a growth regulator, other than an auxin or a cytokinin, present in amounts effective to produce somatic embryos.

Claim 38
A rose callus cell which expresses an exogenous DNA sequence.
Claim 39
A rose plant having cells which express an exogenous DNA sequence.
Claim 40
A somatic rose embryo which expresses an exogenous DNA sequence.

Abandoned application US 2001/007157 is a continuation of now granted US 5792927, which is a divisional of now granted US 5480789 (see above).

Methods for transforming a rose callus with Agrobacterium cells carrying an exogenous DNA. Unlike the granted patent US 5792927, the type of callus to be transformed is not specified.
A method for producing a somatic rose embryo expressing an exogenous sequence is also described. Rose callus, somatic embryo and plant expressing an exogenous gene are recited in the filed claims.

Remarks
  1. National phase entry of WO 1992/00371 in Europe (EP 536327) is deemed to be withdrawn on 2 July 2003.
  2. National phase entries of WO 1992/00371 in Japan (JP H05/507415 T2 and its divisional JP 2001/190169 A2) are deemed to be withdrawn (JP H05/507415 T2 was rejected and an appeal was filed, which was later withdrawn on 13 July 2001; JP 2001/190169 A2 was rejected without any further appeal, notice of rejection sent to applicant on 8 April 2003).

Note: Patent information on this page was last updated on 22 March 2006.