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1. WO2020182857 - TRANSCRIPTION FACTOR-MEDIATED PROMOTION OF DIRECT SOMATIC EMBRYOGENESIS

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Claims

1. A recombinant protein for use in a liquid culture medium for photo-autotrophic micropropagation of Cannabis Sativa L, the recombinant protein comprising a fusion of a growth induction part and a uptake enhancement part,

wherein said growth induction part comprises an Arabinogalactan protein and/or a plant transcription factor associated with plant growth and development,

wherein said uptake enhancement part comprises a cell penetrating peptide sequence and a nuclear localization signal,

wherein said nuclear localization signal is encoded by the peptide sequence SEQ ID NO 1

2. The recombinant protein of claim 1 , wherein said cell penetrating peptide sequence comprising a sequence of 7 to 25 arginines forming a poly-arginine transduction domain.

3. The recombinant protein of any of the previous claims, wherein said growth induction part comprises said plant transcription factor, said plant transcription factor comprising one or more of: Baby Boom, BBM, Leafy Cotyledon, LEC, Wuschel, WUS, RWP-RK Domain-Containing 4, RKD4, Wound Induced Dedifferentiation 1 , WIND1 , and

Plethora2, PLT2.

4. The recombinant protein of claim 3, wherein said plant transcription factor comprises AP2-like ethylene- responsive transcription factor BBM from Arabidopsis Thaliana, or an analog to said AP2-like ethylene- responsive transcription factor BBM from Arabidopsis Thaliana, wherein said analog is codon-optimized for a host species.

5. The recombinant protein of claim 4, wherein said host species is Nicotiana Benthamiana.

6. The recombinant protein of any of the previous claims, wherein said recombinant protein is encoded by the nucleotide sequence SEQ ID NO 2.

7. A plant cell culture medium comprising a recombinant protein in accordance with any of the previous claims. 8. The plant cell culture medium of claim 7, comprising the recombinant protein in a concentration in the range of

0.05 mg/L to 5 mg/L.

9. The plant cell culture medium of claim 7 or claim 8, further comprising salts and/or vitamins.

10. The plant cell culture medium of any of the claims 7 to 9, comprising one or more of the following: 6- benzylaminpurine, Ethephon, Kinetin, Putrescine, Spermidine, Hydrogen Peroxide, 6-(Y,Y- dimethylallylamino)purine, Gibberellic Acid or Gibberellin, and Naphthaleneacetic Acid.

11. The plant cell culture medium of any of the claims 7 to 10, wherein said medium is a liquid culture medium for photo-autotrophic micropropagation.

12. A method for micropropagation of Cannabis Sativa L., the method comprising:

obtaining explant material from a Cannabis Sativa L. plant,

- providing primary and/or secondary embryos by inducing embryogenesis in the explant material in a plant cell culture medium in accordance with any of the claims 7 to 11.

13. The method of claim 12, wherein providing primary and/or secondary embryos comprises a photoautotrophic method in which the explant material is exposed to light for stimulating direct somatic embryogenesis.

14. The method of claim 13, wherein said exposure to light comprises the exposure to light from a photo-synthetical ly active radiation (PAR) source, in the wavelength range of 400 nm to 700 nm, and at a photosynthetic photon flux between 100 and 200 pmol rrr2s-1.