	US 11,925,166 B2
	Cacao cell suspension protocol
Claudia Yanet Garcia, Itabuna (BR); Dahyana Britto, Ilhéus (BR); Jean-Philippe Marelli, Fort Lauderdale, FL (US); and Lidiane Dos Santos Silva, Itajuípe (BR)
Assigned to Mars, Incorporated, McLean, VA (US)
Appl. No. 17/617,215
Filed by Mars, Incorporated, McLean, VA (US)
PCT Filed Jun. 5, 2020, PCT No. PCT/US2020/036356 § 371(c)(1), (2) Date Dec. 7, 2021, PCT Pub. No. WO2020/247771, PCT Pub. Date Dec. 10, 2020.
Claims priority of provisional application 62/858,571, filed on Jun. 7, 2019.
Prior Publication US 2022/0232790 A1, Jul. 28, 2022
Int. Cl. A01H 6/60 (2018.01); A01H 4/00 (2006.01); A01H 5/10 (2018.01); A01H 5/12 (2018.01)

CPC A01H 6/60 (2018.05) [A01H 4/002 (2021.01); A01H 4/005 (2013.01); A01H 5/10 (2013.01)]

13 Claims

1. A method for preparing a Theobroma cell suspension culture, comprising:

obtaining an explant from a young Theobroma leaf, wherein the Theobroma is Theobroma cacao;

sterilizing the explant;

inducing friable callus from the explant on a callus induction medium, wherein the callus induction medium comprises:

a base plant medium;

1-naphthaleneacetic acid (NAA) at a concentration of 1 mg/L; and

6-benzylaminopurine (BAP) at a concentration from 0.5 mg/L to 1 mg/L;

proliferating the induced callus on a callus proliferation medium, wherein the callus proliferation medium comprises: the base plant medium and tidiazuron, and

wherein the base plant medium comprises:

woody plant medium salts;

vitamins;

L-glutamine;

myo-inositol; and

sucrose; and

suspending the proliferated callus in a suspension medium, wherein the suspension medium comprises:

salts;

vitamins;

L-glycine;

sucrose;

penicillin;

2,4 -dichlorophenoxyacetic acid (2,4-D); and

kinetin.