This disclosure pertains to a novel platform for genetic engineering of chloroplasts. The disclosure provides episomal DNA vectors containing a chloroplast origin of replication. These vectors remain extra-plastomic and sustainably and autonomously replicate in chloroplasts of the plant cells transformed with the vectors and in the plants regenerated from the transformed plant cells. The episomal DNA vectors do not contain any sequence that shares sequence homology with the plastome DNA and, thus, do not get integrated into the plastome DNA. The vectors can also comprise one or more genes of interest that confer desirable characteristics to the transformed plant cells. The disclosure also provides methods of transforming plant cells with the episomal DNA vectors and regenerating from the transformed plant cells plants having desirable characteristics. The vectors and methods disclosed herein provide a significant advancement in speed, flexibility, and prospects of introducing genes into plant cells for effective metabolic engineering.

Provided herein are methods and compositions for modifying cells. Provided herein are methods and compositions for modifying an organism of a cell. Provided herein are methods and compositions for introducing polynucleotides and/or polypeptides into a nucleus of a cell.

Methods and compositions for plastid transformation and regeneration or development of transplastomic plants are provided. Embryo explants may be excised from seeds, and their meristematic tissue may be transformed directly without initiation of any callus phase before and/or after transformation. The present methods may be performed with fewer culturing steps relative to conventional methods, thereby enabling more rapid and efficient production of targeted transplastomic events in plants.

Provided are methods for plastid genome editing and development of plants, plant cells, plant parts, and seeds comprising edited plastid genomes. Compositions for transformation of plastid genomes are further provided. Specifically, the plastid genome is modified to comprise the replacement of a targeted endogenous plastid genome sequence with a modified version of the target plastid sequence, where the modified plastid genome sequence has been designed to deliberately reduce its homology to the targeted endogenous plastid genome sequence and, in some cases, encode a protein containing one or more mutations.

Phosphite dehydrogenase (ptxD) expression was established as a selectable marker for nuclear and chloroplast genetic selection in Picochlorum renovo and Picochlorum celeri Phosphite was used as a sole phosphorus source in P. renovo and P. celeri. Growth on phosphite led to comparable growth and composition relative to phosphate.

The present invention provides an organelle transformant screening method. More specifically, the present invention provides an organelle transformant screening method utilizing inhibition of a lipid synthetic pathway.

Use of aprotinin as a carrier to produce a recombinant protein, polypeptide or peptide in algae The present invention relates to the use of aprotinin as a carrier to produce a recombinant protein, polypeptide or peptide in algae, in particular microalgae, wherein aprotinin and said recombinant protein, polypeptide or peptide are fused together to form a fusion protein. It also relates to a method to produce a recombinant protein, polypeptide or peptide in algae, wherein said method comprises genetic transformation of algae, in particular microalgae, with a recombinant nucleic acid sequence encoding a fusion protein, wherein said fusion protein comprises aprotinin and said recombinant protein, polypeptide or peptide. It further relates to a recombinant algae comprising a recombinant nucleic acid sequence encoding a fusion protein, wherein said fusion protein comprises aprotinin and a recombinant protein, polypeptide or peptide. The use of said recombinant algae, for producing said fusion protein is also contemplated.

The present invention is directed to a transformed plant and a method for producing and using the same, or components thereof.

The present invention lies in the field of plant protection, in particular in the field of controlling plant pests and pathogens that affect plants. The present invention relates to a plant comprising a plastid comprising a double-stranded RNA (dsRNA) capable of silencing at least one target gene of a pest of a plant or of an agent causing a disease of a plant. The present invention further relates to such a transplastomic plant, wherein said dsRNA comprises two (separate) complementary single-stranded RNA strands. The present invention further relates to a plastid as comprised in the plant of the invention and to a plant cell comprising said plastid. Moreover, the present invention relates to a method of producing a plant of the invention and to a method of controlling a pest of a plant or a plant disease-causing agent or of protecting a plant from said pest or agent. Furthermore, the present invention relates to the use of a dsRNA for controlling a pest of a plant or a plant disease-causing agent or for protecting a plant from said pest or agent.

Emerging evidence indicates that diminished activity of the vasoprotective axis of the renin-angiotensin system, constituting angiotensin converting enzyme2 (ACE2) and its enzymatic product, angiotensin-(1-7) [Ang-(1-7)] contribute to pulmonary hypertension (PH). However, clinical success for long-term delivery of ACE2 or Ang-(1-7) would require stability and ease of administration to increase patient compliance. Chloroplast expression of therapeutic proteins enables their bioencapsulation within plant cells to protect from acids and gastric enzymes; fusion to a transmucosal carrier facilitates effective systemic absorption. Oral feeding of rats with bioencapsulated ACE2 or Ang-(1-7) attenuated monocrotaline (MCT)-induced increase in right ventricular systolic pressure, decreased pulmonary vessel wall thickness and improved right heart function in both prevention and reversal protocols. Furthermore, combination of ACE2 and Ang-(1-7) augmented the beneficial effects against cardio-pulmonary pathophysiology induced by MCT administration. Experiments have also been performed which indicate that this approach is also suitable for the treatment or inhibition of experimental uveitis and autoimmune uveoretinitis These studies provide proof-of-concept for a novel low-cost oral ACE2 or Ang-(1-7) delivery system using transplastomic technology for pulmonary and ocular disease therapeutics.