Production of plants using somatic embryogenesis

Patent metadata

Field	Value
Publication number	US-2016286749-A1
Application number	US-201415037972-A
Country	US
Kind code	A1
Filing date	Nov 19, 2014
Priority date	Nov 19, 2013
Publication date	Oct 6, 2016
Grant date	—

Abstract

A process for propagating Theobroma cacao L. is described. The process uses direct or indirect somatic embryogenesis to produce primary embryos from explant material, and then applies direct somatic embryogenesis to the resulting primary embryos to produce secondary embryos. These may be matured, pre-germinated, and germinated to form plantlets. Plants, plants bearing fruit, and plant materials are also provided, as well as methods for processing the fruit of the plants to generate cocoa products.

First claim

What is claimed:

1 . A process for propagating Theobroma cacao comprising:
(a-1) obtaining Theobroma cacao L. explant material, followed by
(i) placing the explant material into an induction medium with exposure to light for a period sufficient to obtain primary embryos; and
(ii) optionally transferring the primary embryos from (i) into a development medium and culturing them with exposure to light for a period sufficient to produce primary embryos by direct somatic embryogenesis;
(b-1) removing tissue of primary embryos, followed by
(i) placing the tissue into an induction medium with exposure to light for a period sufficient to obtain secondary embryos; and
(ii) optionally transferring the secondary embryos from (i) into a development medium and culturing them with exposure to light for a period sufficient to produce secondary embryos by direct somatic embryogenesis;
(c) transferring the secondary embryos from step (b-1) to a medium configured to cause maturation of the secondary embryos.

2 . A process according to claim 1 , wherein in step (a-1) the first culture medium comprises a higher concentration of 6-Benzylaminopurine (BAP) than the second culture medium.

3 . A process according to claims 1 - 2 wherein a further step is carried out after step (b-1) above and before step (c), in which the products of step (b-1) are transferred to a medium suitable for initiation, expression and production of new direct secondary somatic embryos and incubated therein with exposure to light until the number of secondary embryos therein is increased.

4 . A process according to claims 1 - 3 wherein the explant material for use in step (a-1) comprises petals, staminodes, and epicotyls of primary embryos of Theobroma cacao L.

5 . A process according to claims 1 - 4 wherein the medium used in at least one of steps (a-1), (b-1) or (c) is a solid medium.

6 . A process according to claims 1 - 5 wherein the medium used in at least one of steps (a-1), (b-1) or (c) is a liquid medium.

7 . A process according to claims 1 - 6 wherein the temperature during steps (a-1) and/or (b-1) and/or (c) is in the range of from 23-29° C.

8 . A process according to claims 1 - 7 wherein during step (a) and step (b), cultures are in the light for at least 50% of each 24 hour period.

9 . A process according to claim 8 wherein the photoperiod is a 16 hour:8 hour (light:dark) with a photosynthetic active radiation (PAR) of 50-60 μmol/m 2 ·sec. x

10 . A process for propagating Theobroma cacao L. comprising:
(a-2) obtaining Theobroma cacao L. explant material;
(i) placing the explant material into a callus formation medium in the dark for a period sufficient to obtain callus formation; and
(ii) transferring the callus from 1) into an embryo development medium and culturing in the dark for a period sufficient to produce primary embryos by indirect somatic embryogenesis;
(b-2) removing tissue of primary embryos;
(i) placing the tissue into an induction medium with exposure to light for a period sufficient to obtain secondary embryos; and
(ii) optionally transferring the secondary embryos from (i) into a development medium and culturing them with exposure to light for a period sufficient to produce secondary embryos by direct somatic embryogenesis;
(c) transferring the secondary embryos from step (b-2) to a medium configured to cause maturation of the secondary embryos.

11 . A process according to claim 10 wherein a further step is carried out after step (b-2) above and before step (c), in which the products of step (b-2) are transferred to a medium suitable for initiation, expression and production of new direct secondary somatic embryos and incubated therein with exposure to light until the number of secondary embryos therein is increased.

12 . A process according to claims 10 - 11 wherein the explant material for use in step (a-2) comprises petals, staminodes, and epicotyls of primary embryos of Theobroma cacao L.

13 . A process according to claims 10 - 12 wherein the medium used in at least one of steps (a-2), (b-2) or (c) is a solid medium.

14 . A process according to claims 10 - 13 wherein the medium used in at least one of steps (a-2), (b-2) or (c) is a liquid medium.

15 . A process according to claims 10 - 14 wherein the temperature during steps (a-2) and/or (b-2) and/or (c) is in the range of from 23-29° C.

16 . A process according to claims 10 - 15 wherein during step (b-2), cultures are in the light for at least 50% of each 24 hour period.

17 . A process according to claim 16 wherein the photoperiod is a 16 hour:8 hour (light:dark) with a photosynthetic active radiation (PAR) of 50-60 μmol/m 2 ·sec.

18 . A process for propagating Theobroma cacao L. according to any one of the preceding claims, further comprising pre-germinating, optionally germinating, and developing embryos obtained by the propagation methods into plantlets.

19 . A process for preparing direct secondary somatic embryos of Theobroma cacao L. comprising culturing primary somatic embryos in an induction media comprising BAP, followed by developing embryos in a development media comprising gibberellic acid.

20 . The process of claim 19 , in which the induction and development media also include sugar.

21 . The process of claims 19 - 20 , in which the BAP is present in an amount of 0.5 mg/L to 1.5 mg/L.

22 . The process of claims 19 - 21 , in which the gibberellic acid is present in an amount of 0.2 g/L to about 0.8 g/L.

23 . The process of claims 20 - 22 , in which the sugar is sucrose, and is present in an amount of from 20 g/L to 80 g/L.

24 . The process of claims 19 - 23 , in which BAP is present in an amount of 1.0 mg/L, gibberellic acid is present in an amount of 0.6 g/L, and sucrose is present in an amount of 40 g/L.

25 . The process of any one of the previous claims wherein there is a further step of propogating the embryos or plantlets into plants bearing fruit.

26 . The process of claim 25 wherein the fruit is removed from the original plant.

27 . The process of claim 26 wherein the fruit is processed into a cocoa product.

28 . The process of claim 27 wherein the cocoa product is cocoa liquor, chocolate, compound chocolate, chocolate-like substance, cocoa powder or cocoa butter.

29 . The process of any one of claim 26 to claim 28 wherein the fruit is partially or fully depulped.

30 . The process of claim 29 wherein the partially or fully depulped fruit is fermented using microbiological processes or chemical processes.

31 . The process of any one of claim 26 or claim 30 wherein the fruit are dried by exposing the fruit to the sun or an artificial dryer including but not limited to a Samoan dryer, Buttner dryer, platform dryer.

32 . The process of claim 31 wherein the fruit are further roasted.

33 . The process of claim 32 wherein the roasted fruit are winnowed to generate a nib.

34 . The process of claim 33 wherein the nib is converted to cocoa liquor or cocoa butter.

35 . The process of claim 34 wherein the nib or cocoa liquor is converted to cocoa powder.

36 . The process of claim 33 or 34 wherein the liquor and/or cocoa butter

Assignees

• Mars Inc

Inventors

• Garcia Rojas Claudia Yanet
• Dias Cristiano Villela
• Marelli Jean-Philippe

Classifications

• A23G1/30

  Cocoa products, e.g. chocolate; Substitutes therefor · CPC title

• A01H4/005 Primary

  Methods for micropropagation; Vegetative plant propagation using cell or tissue culture techniques · CPC title