The correct answer is (2) Particle delivery system (biolistic “gene gun” method).

Why the particle delivery system works for chloroplast transformation

The particle delivery system (biolistics or gene gun) accelerates microscopic metal particles coated with DNA so they physically penetrate cell walls and membranes and lodge directly in organelles such as chloroplasts. Once DNA reaches the stroma, homologous recombination between flanking plastid sequences in the vector and the chloroplast genome integrates the transgene into plastid DNA. This direct physical delivery to chloroplasts makes biolistics the routine, textbook method for chloroplast (plastid) transformation in many crops.

Explanation of each option

1. Agrobacterium-mediated transformation
   Agrobacterium naturally transfers T-DNA mainly into the nuclear genome, where it integrates by illegitimate recombination. Experimental Agrobacterium-based plastid transformation is rare and not a standard method for routine chloroplast engineering. So this is not the typical choice for introducing genes into chloroplast DNA.

2. Particle delivery system (biolistics) – Correct
   Biolistic transformation is widely used for plastid engineering because DNA-coated particles can directly reach chloroplasts inside cells without needing protoplasts or specialized uptake pathways. This method is standard for generating stably transformed chloroplast genomes in higher plants.

3. Permeabilization
   Chemical permeabilization (e.g., PEG treatment) is mainly used for protoplast transformation, most often targeting the nuclear genome. While PEG-mediated plastid transformation has been demonstrated in some systems, it is less robust and not the classic, most likely method referenced in exam questions compared with the particle gun.

4. Electroporation
   Electroporation can introduce DNA into protoplasts or bacteria by creating transient pores in membranes, again usually aimed at nuclear transformation. Chloroplast-targeted electroporation is technically possible but is not the standard or most widely adopted strategy in higher-plant chloroplast engineering.

SEO‑oriented introduction (for article use)

In plant biotechnology, stable chloroplast genome transformation is most reliably achieved using the particle delivery (gene gun) system, which shoots DNA-coated microprojectiles directly into chloroplasts, enabling homologous recombination with plastid DNA. In contrast, Agrobacterium-mediated transformation, simple permeabilization, and standard electroporation are primarily used for nuclear transformation or specialized protoplast systems, making the particle delivery system the preferred answer for this question.