Patent EP0270356A2 successfully refreshed from Google Patents.
EP0270356A2
Expired / Lapsed
EP
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Plant-cell transformation by accelerated particles coated with DNA and apparatus therefor.
Assignee: Powderject Vaccines Inc
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Filing: 1987-12-02
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Granted: —
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Expires: 2007-12-02
Abstract
Abstract A method and apparatus is disclosed for the genetic transformation of plants and plant lines by pollen mediated transformation. Foreign genes are introduced into pollen by coating on carrier particles which are physically accelerated into plant pollen. The treated plant pollen is then hand pollinated and the progeny are recovered, a portion of which will contain in their genome the foreign gene. The procedure may be used to create genetically engineered maize plant and lines.
Inventors
- 👤 Dennis E. Mccabe
- 👤 William F. Swain
- 👤 Brian J. Martinell
Bibliographic Details
Application Number
—
Jurisdiction
EP
Data Source
google_patents
Last Synchronized
2026-10-23 11:32
CPC Classifications
C; C12; C12N; C12N15/00; C12N15/09; C12N15/87; C12N15/63; C12N15/79; C12N15/82; C12N15/8201; C12N15/8206; C12N15/8207
Claims Analysis
Total Claims: 33
Claim 1
Independent Claim
104 words
A method of making a genetically transformed line of plants characterised in that it comprises the steps of: preparing copies of a foreign gene including a coding region and flanking regulatory sequences effective to express the coding region in the plant cells; joining copies of the foreign gene to biologically inert carrier particles; physically accelerating the particles carrying the gene copies at pollen of the plant in such a fashion that some particles lodge in the interior of some of the pollen; pollinating female organs of the plant with the pollen so treated; and screening among the progeny from this pollination for transformed progeny.
Claim 2
Independent Claim
104 words
A method of making a genetically transformed line of plants characterised in that it comprises the steps of: preparing copies of a foreign gene including a coding region and flanking regulatory sequences effective to express the coding region in the plant cells; joining copies of the foreign gene to biologically inert carrier particles; physically accelerating the particles carrying the gene copies at pollen of the plant in such a fashion that some particles lodge in the interior of some of the pollen; pollinating female organs of the plant with the pollen so treated; and screening among the progeny from this pollination for transformed progeny.
Claim 3
Dependent Claim
16 words
A method as claimed in claim 1, characterised in that the biologically inert particles are metallic.
Claim 4
Dependent Claim
16 words
A method as claimed in claim 2, characterised in that the metallic particles are gold spheres.
Claim 5
Dependent Claim
35 words
A method as claimed in either claim 2 or claim 3, characterised in that it includes the step of coating the metallic particles with an encapsulating agent before the gene copies are joined to them.
Claim 6
Dependent Claim
15 words
A method as claimed in claim 4, characterised in that the encapsulating agent is polylysine.
Claim 7
Dependent Claim
37 words
A method as claimed in either claim 4 or claim 5, characterised in that the step of joining the foreign gene copies to the metallic particles includes drying a solution of the gene copies onto the particles.
Claim 8
Dependent Claim
29 words
A method as claimed in claim 6, characterised in that a solution of the gene copies includes CaHPO₄ which is precipitated with the gene copies onto the metallic particles.
Claim 9
Independent Claim
60 words
A method as claimed in any one of the preceding claims, characterised in that the step of physically accelerating the particles includes layering the particles on a planar carrier sheet (18), accelerating the carrier sheet by impacting it with a shock wave, and restraining the carrier sheet such that the momentum of the carrier particles carries them from the sheet.
Claim 10
Dependent Claim
36 words
A method as claimed in claim 8, characterised in that the shock wave generated to impact the carrier sheet (18) is generated by high-voltage electric discharge through a spark gap bridged by a water droplet (24).
Claim 11
Independent Claim
46 words
A method as claimed in any one of the preceding claims, characterised in that the step of accelerating the particles at plant pollen includes placing a monolayer of pollen on a target surface and placing the target surface in the acceleration path of the carrier particles.
Claim 12
Dependent Claim
30 words
A method as claimed in claim 10, characterised by the steps of applying a layer of mineral oil to the target surface, dusting pollen thereon, and removing the excess therefrom.
Claim 13
Dependent Claim
55 words
A method as claimed in either claim 10 or claim 11, characterised in that the step of pollinating female organs of the plant is accomplished by brushing pollen into which particles have been accelerated off the target surface with a brush and dusting that pollen onto the female organs of a selected female parent plant.
Claim 14
Independent Claim
28 words
A method as claimed in any one of the preceding claims, characterised in that the screening among the progeny is done on the basis of a biochemical assay.
Claim 15
Dependent Claim
16 words
A method as claimed in claim 13, characterised in that the assay is for antibiotic resistance.
Claim 16
Independent Claim
30 words
A method as claimed in any one of claims 1 to 12, characterised in that the screening among the progeny is done on the basis of a morphological plant trait.
Claim 17
Independent Claim
18 words
A method as claimed in any one of the preceding claims, characterised in that the plants are maize.
Claim 18
Dependent Claim
15 words
Plants genetically transformed by a method as claimed in any one of the preceding claims.
Claim 19
Independent Claim
70 words
A method of making a genetically transformed maize plant characterised in that it comprises the steps of: physically injecting a foreign gene including a coding region and appropriate regulatory sequences into maize pollen from a male parent plant; hand pollinating the pollen onto the mature maize silks of a female parent plant and segregating the silks from other pollen sources; growing up progeny produced from the fertilized female parent plant.
Claim 20
Independent Claim
70 words
A method of making a genetically transformed maize plant characterised in that it comprises the steps of: physically injecting a foreign gene including a coding region and appropriate regulatory sequences into maize pollen from a male parent plant; hand pollinating the pollen onto the mature maize silks of a female parent plant and segregating the silks from other pollen sources; growing up progeny produced from the fertilized female parent plant.
Claim 21
Dependent Claim
33 words
A method as claimed in claim 18, characterised in that the step of physical injection includes joining copies of the foreign gene to carrier particles and accelerating the particles at the maize pollen.
Claim 22
Dependent Claim
23 words
A method as claimed in claim 19, characterised in that the carrier particles are gold beads coated with an encapsulating agent, preferably polylysine.
Claim 23
Dependent Claim
16 words
Maize plants produced by a method as claimed in any one of claims 18 to 20.
Claim 24
Dependent Claim
17 words
A maize plant which has a parent plant as claimed in either claim 17 or claim 21.
Claim 25
Dependent Claim
24 words
Maize pollen comprising in it a foreign chimeric gene and at least one carrier particle which physically carried the foreign gene into the pollen.
Claim 26
Dependent Claim
24 words
Maize pollen comprising in it a foreign chimeric gene and at least one carrier particle which physically carried the foreign gene into the pollen.
Claim 27
Dependent Claim
34 words
Maize pollen as claimed in claim 23, characterised in that the foreign gene includes a coding sequence and appropriate flanking regulatory sequences so that the coding sequence is expressed in progeny of the pollen.
Claim 28
Dependent Claim
136 words
Apparatus for injecting carrier particles carrying DNA into living cells characterised in that it comprises: a spark discharge chamber (12); two electrodes (14) extending into the spark discharge chamber and spaced apart by a spark gap, the electrodes being adapted for attachment to an external source of high voltage discharge; a carrier sheet (18) held spaced above the spark discharge chamber, the carrier sheet receiving the carrier particles thereon; a retaining screen (20) fixed in place above the carrier sheet; and a target surface (22) held spaced above the retaining screen (20) and carrying the cells so that a spark discharge generating a shock wave in the discharge chamber (12) will accelerate the carrier sheet (18) into the retaining screen (20) so that the carrier particles are accelerated into the cells on the target surface (22).
Claim 29
Dependent Claim
136 words
Apparatus for injecting carrier particles carrying DNA into living cells characterised in that it comprises: a spark discharge chamber (12); two electrodes (14) extending into the spark discharge chamber and spaced apart by a spark gap, the electrodes being adapted for attachment to an external source of high voltage discharge; a carrier sheet (18) held spaced above the spark discharge chamber, the carrier sheet receiving the carrier particles thereon; a retaining screen (20) fixed in place above the carrier sheet; and a target surface (22) held spaced above the retaining screen (20) and carrying the cells so that a spark discharge generating a shock wave in the discharge chamber (12) will accelerate the carrier sheet (18) into the retaining screen (20) so that the carrier particles are accelerated into the cells on the target surface (22).
Claim 30
Dependent Claim
20 words
Apparatus as claimed in claim 25, characterised in that a water droplet (24) is disposed bridging between the electrodes (14).
Claim 31
Dependent Claim
29 words
Apparatus as claimed in either claim 25 or claim 26, characterised in that it includes a vacuum chamber to allow the apparatus to be operated in a partial vacuum.
Claim 32
Dependent Claim
26 words
Apparatus as claimed in any one of claims 25 to 27, characterised in that the spark gap between the electrodes is between 1 and 1.5 millimeters.
Claim 33
Dependent Claim
42 words
Apparatus as claimed in any one of claims 25 to 28, characterised in that a spacer ring (16) is positioned above the spark discharge chamber (12) but below the carrier sheet (18) and with at least a partial opening at its top.
References Cited (Backward Citations)
No citation records indexed.
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